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Direct immersion solid-phase microextraction coupled with gas chromatography-mass spectroscopy was used to create chemical fingerprints of annual ryegrass cultivars ( Lolium rigidum ). Extracts made of the inflorescences of four cultivars and one accession of annual ryegrass were assessed to identify differential metabolites between those resistant to and susceptible to bacterial galls associated with annual ryegrass toxicity (ARGT). Numerous compounds were identified. Principal component analysis showed distinct clustering of metabolites from disease-resistant and disease-susceptible cultivars. Partial least-squares-discriminant analysis identified sterols, esters, aldehydes, and terpenes that correlated with resistance to galls formation. Esters, sterols, phenols, heterocyclics, fatty acids, organofluorides, and siloxanes were predominant in resistant genotypes, whereas alcohols, aldehydes, terpenes, and hydrocarbons were predominant in susceptible genotypes. The identification of differentially expressed metabolites provides potential chemical markers to guide breeding strategies for ARGT resistance in ryegrass.

Polycyclic aromatic hydrocarbons (PAHs) constitute a category of persistent organic contaminants that possess the potential to induce carcinogenic, teratogenic, and mutagenic consequences. Our previous findings have revealed that plant roots actively take up PAHs through co-transport with protons, and auxin can promote PAHs uptake by wheat roots. It remains unclear whether nitric oxide (NO), a signaling molecule involved in numerous physiological processes in plants and downstream of auxin, can affect PAHs uptake by plant roots. In our study, 50 μmol/L sodium nitroprusside (SNP) significantly enhanced phenanthrene uptake after 4 h of exposure. After the addition of SNP (50 μmol/L), the H + flux on root surface increased, and H + -ATPase activity was activated, indicating that exogenous NO promotes phenanthrene uptake by plant roots via activating H + -ATPase. By studying the effects of 50 μmol/L cyclic guanosine monophosphate (cGMP), 5 mmol/L Ca 2+ , and 50 μmol/L adenosine monophosphate (AMP) on phenanthrene uptake by ryegrass roots and measuring root calcium-dependent protein kinases (CDPK) activity, we demonstrated that exogenous NO promotes phenanthrene uptake through the signaling pathway of NO, cGMP, Ca 2+ , CDPK, 14-3-3 protein and H + -ATPase. The results contribute significant insights into elucidating the underlying mechanisms of NO modulating PAHs absorption by plant roots, thereby offering crucial strategies for advancing food safety measures and enhancing the phytoremediation potential of soils and waters contaminated with PAHs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Lipids and Ca 2+ are involved as intermediate messengers in temperature-sensing signaling pathways. Arbuscular mycorrhizal (AM) symbiosis is a mutualistic symbiosis between fungi and terrestrial plants that helps host plants cope with adverse environmental conditions. Nonetheless, the regulatory mechanisms of lipid- and Ca 2+ -mediated signaling pathways in mycorrhizal plants under cold and heat stress have not been determined. The present work focused on investigating the lipid- and Ca 2+ -mediated signaling pathways in arbuscular mycorrhizal (AM) and non-mycorrhizal (NM) roots under temperature stress and determining the role of Ca 2+ levels in AM symbiosis and temperature stress tolerance in perennial ryegrass (Lolium perenne L.) Compared with NM plants, AM symbiosis increased phosphatidic acid (PA) and Ca 2+ signaling in the roots of perennial ryegrass, increasing the expression of genes associated with low temperature (LT) stress, including LpICE1, LpCBF3, LpCOR27, LpCOR47, LpIRI, and LpAFP, and high temperature (HT) stress, including LpHSFC1b, LpHSFC2b, LpsHSP17.8, LpHSP22, LpHSP70, and LpHSP90, under LT and HT conditions. These effects result in modulated antioxidant enzyme activities, reduced lipid peroxidation, and suppressed growth inhibition caused by LT and HT stresses. Furthermore, exogenous Ca 2+ application enhanced AM symbiosis, leading to the upregulation of Ca 2+ signaling pathway genes in roots and ultimately promoting the growth of perennial ryegrass under LT and HT stresses. These findings shed light on lipid and Ca 2+ signal transduction in AM-associated plants under LT and HT stresses, emphasizing that Ca 2+ enhances cold and heat tolerance in mycorrhizal plants., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

This study aimed to examine the potential of soil amendment-assisted phytoremediation using ryegrass in reclaiming abandoned gold mine soil in southwestern Ghana, with a specific focus on the soil contamination hazards associated with metals and metalloids. A pot experiment lasting 60 days was carried out to assess the efficacy of soil amendments, such as compost, iron oxide, and poultry manure, in mitigating environmental hazards. Three soil contamination indices (soil contamination = CF, enrichment factor = ER, and pollution load index = PLI) were used to calculate the extent of soil contamination, enrichment, and pollution of the sites with Co, Hg, Ni, Mo, Se, Sb, and Pb. The findings show that Hg made the greatest contribution (with a maximum soil CF of 18.0) to the overall PLI, with a maximum value of 74.4. The sites were averagely and consequently enriched with toxic elements in the decreasing order: Ni (ER = 33.3) > Mo (20.5) > Sb (14.1) > Pb (11.0) > Hg (7.9) > Se (2.1). The bioaccumulation factor (BCF > 1) suggests that ryegrass has the ability to phytostabilize Co, Hg, Mo, and Ni. This means that the plant may store these elements in its roots, potentially decreasing their negative effects on the environment and human health. Ultimately, the addition of combined manure with iron oxides might have augmented the sequestration of these metals in the root. The elements may have accumulated through sorption on manure or Fe surfaces, dissolution from watering the plants in the pot, or mineralization of organic manure. Thus, ryegrass has shown potential for phytostabilisation of Co, Hg, Mo, and Ni when assisted with a combination of manure and iron oxides; and can consequently mitigate the environmental and human health impacts.

Background: Low rumen pH is proposed to be a major mechanism for low methane (CH 4 ) emissions from sheep fed forage rape. However, it is difficult to separate this from other in vivo factors, such as rumen passage rate. The objective of this study was to determine the effect of pH alone on CH 4 production in vitro using different pH buffers. Ryegrass, white clover and forage rape were incubated in vitro using three different incubation buffers with starting pH values of 5.5, 6.2 and 6.8., Results: Decreasing pH reduced overall in vitro CH 4 emission relative to fermented hexoses (CH 4 /FHex) by up to 54% and overall fermentation by 40%. pH also changed fermentation profiles where the acetate + butyrate to propionate + valerate ratio decreased when pH decreased. Within the three forages, forage rape led to the lowest CH 4 /FHex, but only in pH 5.5 and 6.2 buffer, and this was enhanced when the pH fell below 6., Conclusion: Reducing pH in vitro decreased CH 4 production and overall fermentation across all forages. The lower pH reached by forage rape compared to ryegrass and white clover appears to drive the lower CH 4 production relative to the extent of fermentation from forage rape compared to the other forages. © 2024 The Author(s). Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2024 The Author(s). Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)

Dissolved organic matter (DOM) regulates plant behavior in both agricultural and environmental fields. However, the regulatory mechanisms by which DOM influences soil-plant system interactions during the phytoremediation of Cd-contaminated soils remain unclear. Therefore, this study investigated the enhanced effect of kitchen compost-derived DOM on the Cd remediation capability of ryegrass across three phases of phytoremediation. The main pathways and mechanisms of DOM-assisted phytoremediation were identified through the analysis of changes in soil microbial communities and metabolism functions. The results revealed that DOM increased the bioavailability of soil Cd and significantly enhanced the Cd enrichment capacity of ryegrass, regardless of the application rate. The application of 20 % DOM to soil with a 20 mg/kg Cd content increased the bioconcentration factors of ryegrass roots and shoots by up to 38.19 and 11.08 times, respectively, compared with the control group. The direct or indirect optimizing effects of DOM on Cd fraction transformation, microbial communities, and their metabolism functions significantly enhanced the Cd enrichment capacity of ryegrass. Notably, DOM exhibited dual effects on ryegrass growth, mainly influenced by changes in soil physicochemical properties, optimization of microbial communities, and alterations in nitrogen metabolic functions. Additionally, the Cd reserves in ryegrass, which serve as a vital indicator of phytoremediation, exhibited a positive response to DOM. This study provides insights into the various reinforcing roles of kitchen compost-derived DOM in Cd-contaminated soil phytoremediation. These findings support the development of effective agronomic strategies for precise Cd regulation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The challenge of soil salinization and alkalization, with its significant impact on crop productivity, has raised growing concerns with global population growth and enhanced environmental degradation. Although arbuscular mycorrhizal fungi (AMF) and calcium ions (Ca 2+ ) are known to enhance plant resistance to stress, their combined effects on perennial ryegrass' tolerance to salt and alkali stress and the underlying mechanisms remain poorly understood. This study aimed to elucidate the roles of Arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis and exogenous Ca 2+ application in molecular and physiological responses to salt-alkali stress. AM symbiosis and exogenous Ca 2+ application enhanced antioxidant enzyme activity and non-enzymatic components, promoting reactive oxygen species (ROS) scavenging and reducing lipid peroxidation while alleviating oxidative damage induced by salt-alkali stress. Furthermore, they enhanced osmotic balance by increasing soluble sugar content (Proportion of contribution of the osmotic adjustment were 34∼38 % in shoots and 30∼37 % in roots) under salt stress and organic acid content (Proportion of contribution of the osmotic adjustment were 32∼36 % in shoots and 37∼42 % in roots) under alkali stress. Changes in organic solute and inorganic cation-anion contents contributed to ion balance, while hormonal regulation played a role in these protective mechanisms. Moreover, the protective mechanisms involved activation of Ca 2+ -mediated signaling pathways, regulation of salt-alkali stress-related genes (including LpNHX1 and LpSOS1), increased ATPase activity, elevated ATP levels, enhanced Na + extrusion, improved K + absorption capacity, and a reduced Na + /K + ratio, all contributing to the protection of photosynthetic pigments and the enhancement of photosynthetic efficiency. Ultimately, the combined application of exogenous Ca 2+ and AMF synergistically alleviated the inhibitory effects of salt-alkali stress on perennial ryegrass growth. This finding suggested that exogenous Ca 2+ may participate in the colonization of perennial ryegrass plants by R. irregularis, while AM symbiosis may activate Ca 2+ pathways. Consequently, the combined treatment of AM and Ca 2+ is beneficial for enhancing plant regulatory mechanisms and increasing crop yield under salt-alkali stress., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Considering both electrokinetic remediation and phytoremediation have limitations, an electrokinetic phytoremediation (EP) system was constructed to obtain efficient and environmentally friendly remediation results. This study indicates that the electric field can promote the absorption of Cd by ryegrass with little impact on soil physicochemical properties under the condition of rotary switching electrodes, and the accumulation of Cd in the aboveground and underground parts of ryegrass increased by 145.2% and 93.7%, respectively. The DC electric field combined with ryegrass under rotary switching electrode mode proved to be the optimal condition for the remediation of Cd contaminated soil with a remediation efficiency of 66.7%. Moreover, the rotary switching of the electrodes alleviated the suppression of the growth of ryegrass by the DC electric field. During the EP remediation process, the electric field promoted the transformation of the residue state of Cd to the other forms, which accelerated the desorption rate of Cd from the soil and facilitated the migration of Cd into plants. In conclusion, EP is a green and efficient remediation technology for heavy metal contaminated soil with good application prospects., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Genome mining in association with the OSMAC (one strain, many compounds) approach provides a feasible strategy to extend the chemical diversity and novelty of natural products. In this study, we identified the biosynthetic gene cluster (BGC) of restricticin, a promising antifungal agent featuring a reactive primary amine, from the fungus Aspergillus sclerotiorum LZDX-33-4 by genome mining. Combining heterologous expression and the OSMAC strategy resulted in the production of a new hybrid product ( 1 ), along with N -acetyl-restricticin ( 2 ) and restricticinol ( 3 ). The structure of 1 was determined by spectroscopic data, including optical rotation and electronic circular dichroism (ECD) calculations, for configurational assignment. Compound 1 represents a fusion of restricticin and phytotoxic cichorin. The biosynthetic pathway of 1 was proposed, in which the condensation of a primary amine of restricticin with a precursor of cichorine was postulated. Compound 1 at 5 mM concentration inhibited the growth of the shoots and roots of Lolium perenne , Festuca arundinacea , and Lactuca sativa with inhibitory rates of 71.3 and 88.7% for L. perenne , 79.4 and 73.0% for F. arundinacea , and 58.2 and 52.9% for L. sativa. In addition, compound 1 at 25 μg/mL showed moderate antifungal activity against Fusarium fujikuroi and Trichoderma harzianum with inhibition rates of 22.6 and 31.6%, respectively. These results suggest that heterologous expression in conjunction with the OSMAC approach provides a promising strategy to extend the metabolite novelty due to the incorporation of endogenous metabolites from the host strain with exogenous compounds, leading to the production of more complex compounds and the acquisition of new physiological functions.

Background: The potential of phytoremediation using garlic monoculture (MC) and intercropping (IC) system with perennial ryegrass to enhance the uptake of cadmium (Cd), chromium (Cr), and lead (Pb) were investigated., Results: Positive correlations were found between MC and IC systems, with varying biomass. Production of perennial ryegrass was affected differently depending on the type of toxic metal present in the soil. Root growth inhibition was more affected than shoot growth inhibition. The total biomass of shoot and root in IC was higher than MC, increasing approximately 3.7 and 2.9 fold compared to MC, attributed to advantages in root IC crop systems. Photosystem II efficiency showed less sensitivity to metal toxicity compared to the control, with a decrease between 10.07-12.03%. Among gas exchange parameters, only Cr significantly affected physiological responses by reducing transpiration by 69.24%, likely due to leaf chlorosis and necrosis., Conclusion: This study exhibited the potential of garlic MC and IC with perennial ryegrass in phytoremediation. Although the different metals affect plant growth differently, IC showed advantages over MC in term biomass production., (© 2024. The Author(s).)

The perennity of grassland species such as Lolium perenne greatly depends on their ability to regrow after cutting or grazing. Refoliation largely relies on the mobilization of fructans in the remaining tissues and on the associated sucrose synthesis and transport towards the basal leaf meristems. However, nothing is known yet about the sucrose synthesis pathway. Sucrose Phosphate Synthase (SPS) and Sucrose Synthase (SuS) activities, together with their transcripts, were monitored during the first hours after defoliation along the leaf axis of mature leaf sheaths and elongating leaf bases (ELB) where the leaf meristems are located. In leaf sheaths, which undergo a sink-source transition, fructan and sucrose contents declined while SPS and SuS activities increased, along with the expression of LpSPSA, LpSPSD.2, LpSuS1, LpSuS2, and LpSuS4. In ELB, which continue to act as a strong carbon sink, SPS and SuS activities increased to varying degrees while the expression of all the LpSPS and LpSuS genes decreased after defoliation. SPS and SuS both contribute to refoliation but are regulated differently depending on the source or sink status of the tissues. Together with fructan metabolism, they represent key determinants of ryegrass perennity and, more generally, of grassland sustainability., (© 2024 Scandinavian Plant Physiology Society.)

While plant toxicity reduction remains the primary metric for judging the success of metal immobilization in soil, the suitability of microorganisms as universal indicators of its effectiveness in various contaminated soils remains a point of contention. This study assessed the sensitivity of microbial bioindicators in monitoring metal immobilization success in smelter-impacted soils. It compared plants and microorganisms as indicators of the efficiency of natural Fe-Mn nodules from the Gulf of Finland in immobilizing metals in soils contaminated by a Ni/Cu smelter, on the Kola Peninsula, Murmansk region, Russia. Perennial ryegrass (Lolium perenne) was grown on nodule-amended and control soils. Plant responses in the smelter-impacted soils proved to be sensitive and robust indicators of successful metal immobilization. However, microbial responses exhibited a more complex story. Despite the observed reductions in soluble metal concentrations, shoot metal contents in ryegrass, and significant improvements in plant growth, certain microbial bioindicators were unresponsive to metal immobilization success brought about by the addition of Fe-Mn nodules. Among microbial bioindicators studied, community-level physiological profiling, microbial biomass carbon, and basal respiration were sensitive indicators of metal immobilization success, whereas the number of saprotrophic, oligotrophic, and Fe-oxidizing bacteria and fungi, the biomass of bacteria and fungi, and enzymatic activity were less robust indicators. Interestingly, the correlations between different microbial responses measured were weak or even negative. Some microbial responses also exhibited negative correlations with plant biomass. These findings underscore the need for further research on comparative evaluations of plants and microorganisms as reliable indicators of metal immobilization efficacy in polluted environments., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Phytoremediation has emerged as a common technique for remediating Cd pollution in farmland soil. Moreover, phosphorus, an essential element for plants, can alter the pectin content of plant cell walls and facilitate the accumulation of Cd in plant tissues, thereby enhancing phytoremediation efficiency. Therefore, pot experiments were conducted in order to investigate the effect of phosphorus levels on Cd extraction, phosphorus transformation and phosphorus-related genes during phytoremediation. The results revealed that an optimal application of suitable phosphate fertilizers elevated the soil's pH and electrical conductivity (EC), facilitated the conversion of soil from insoluble phosphorus into available forms, augmented the release of pertinent enzyme activity, and induced the expression of phosphorus cycling-related genes. These enhancements in soil conditions significantly promoted the growth of ryegrass. When applying phosphorus at a rate of 600 mg/kg, ryegrass exhibited plant height, dry weight, and chlorophyll relative content that were 1.27, 1.26, and 1.18 times higher than those in the control group (P 0 ), while the Cd content was 1.12 times greater than that of P 0 . The potentially toxic elements decline ratio and bioconcentration factor were 42.86% and 1.17 times higher than those of P 0 , respectively. Consequently, ryegrass demonstrated the highest Cd removal efficiency under these conditions. Results from redundancy analysis (RDA) revealed a significant correlation among pH, total phosphorus, heavy metal content, phosphorus forms, soil enzyme activity, and phosphorus-related genes. In conclusion, this study suggests applying an optimal amount of suitable phosphate fertilizers can enhance restoration efficiency, leading to a reduction in soil Cd content and ultimately improving the safety of crop production in farmlands., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

This study focused on the effects of urea humate-based porous materials (UHPM) on soil aggregates, plant physiological characteristics, and microbial diversity to explore the effects of UHPM on the phytoremediation of petroleum-contaminated soil. The compositions of soil aggregates, ryegrass (Lolium perenne) biomass, plant petroleum enrichment capacity, and bacterial communities in soils with and without UHPM were investigated. The results showed that UHPM significantly increased soil aggregate content by 0.25 mm-5 mm, resulting in higher fertilizer holding capacity, erosion resistance capacity, and plant biomass and microbial number than the soil without UHPM mixed. In addition, UHPM decreased the absorption of petroleum by plants in the soil while increasing the abundance of degrading bacteria and petroleum-degrading-related genes in the soil, thereby promoting the removal of hard-to-degrade petroleum components. RDA showed that, compared with the unimproved soil, each soil indicator was positively correlated with a high abundance of degrading bacteria in the improved soil and was significant. UHPM can be regarded as a petroleum-contaminated soil remediation agent that combines slow nutrient release with soil improvement effects., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Insitu stabilization and phytoextraction are considered as two convenient and effective technologies for the remediation of toxic elements (TEs) in soils. However, the effectiveness of these two remediation technologies together on the bioavailability and phytoextraction of TEs in field trials has not been explored yet. Specifically, the remediation potential of fly ash (FA; as stabilizing agent) and ryegrass (as a TE accumulator) intercropped with a target crop for soil polluted with multiple TEs has not been investigated yet, particularly in long-term field trials. Therefore, in this study, a six-month combined remediation field experiment of FA stabilization and/or ryegrass intercropping (IR) was carried out on the farmland soils contaminated with As, Cd, Cr, Cu, Hg, Ni, Pb and Zn where Zanthoxylumbungeanum (ZB) trees as native crops were grown for years. The treatments include soil cultivated alone with ZB untreated- (control) and treated-with FA (FA), produced by burning lignite in Shaanxi Datong power plant, China, soil cultivated with ZB and ryegrass untreated- (IR) and treated-with FA (FA + IR). This was underpinned by a large-scale survey in Daiziying (China), which showed that the topsoils were polluted by Cd, Cu, Hg and Pb, and that Hg and Pb contents in the Zanthoxylumbungeanum fruits exceeded their allowable limits. The TEs contents in the studied FA were lower than their total element contents in the soil. The DTPA-extractable TEs contents of the remediation modes were as follows: FA < FA + IR < IR < control. Notably, TEs contents in the ZB fruits were lowest under the FA + IR treatment, which were decreased by 27.6% for As, 42.3% for Cd, 16.7% for Cr, 30.5% for Cu, 23.1% for Hg, 15.5% for Ni, 33.2% for Pb and 38.1% for Zn compared with the control treatment. Whereas the FA + IR treatment enhanced TEs contents in ryegrass shoots and roots, and the TEs contents in ryegrass shoots were below their regulatory limits for fodder crops. The findings confirmed that the combined remediation strategy, i.e., FA (with low content of TEs) stabilization effect and intercropping of ZB (target crop) and ryegrass (accumulating plant) could provide a prospective approach to produce target plants within safe TEs thresholds with greater economic benefits, while remediating soils polluted with multiple TEs and mitigating the potential ecological and human health risk. Those results are of great applicable concern., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Cadmium (Cd) and arsenic (As) are two highly toxic heavy metals and metalloids that coexist in many situations posing severe threats to plants. Our investigation was conducted to explore the different regulatory mechanisms of ryegrass (Lolium perenne L.) responding to individual and combined Cd and As stresses in hydroponics. Results showed that the ryegrass well-growth phenotype was not affected by Cd stress of 10 mg·L -1 . However, As of 10 mg·L -1 caused rapid water loss, proline surge, and chlorosis in shoots, suggesting that ryegrass was highly sensitive to As. Transcriptomic analysis revealed that the transcription factor LpIRO2 mediated the upregulation of ZIP1 and YSL6 that played an important role in Cd tolerance. We found that the presence of As caused the overexpression of LpSWT12, a process potentially regulated by bHLH14, to mitigate hyperosmolarity. Indoleacetic acid (IAA) and abscisic acid (ABA) contents and expression of their signaling-related genes were significantly affected by As stress rather than Cd. We predict a regulatory network to illustrate the interaction between transporters, transcription factors, and signaling transduction, and explain the antagonism of Cd and As toxicity. This present work provides a research basis for plant protection from Cd and As pollution., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Sustainable technologies for the recovery of rare earth elements (REE) from waste need to be developed to decrease the volume of ore mining extractions and its negative environmental consequences, while simultaneously restoring previously impacted lands. This is critical due to the extensive application of REE in everyday life from electronic devices to energy and medical technologies, and the dispersed distribution of REE resources in the world. REE recovery by plants has been previously studied but the feasibility of REE phytoextraction from a poorly soluble solid phase (i.e., nanoparticles) by different plant species has been rarely investigated. In this study, the effect of biostimulation and bioaugmentation on phytorecovery of REE nanoparticles (REE-NP) was investigated by exposing ryegrass seeds to REE-NP in hydroponic environments. This was studied in two sets of experiments: bioaugmentation (using CeO 2 nanoparticles and Methylobacterium extorquens AM1 pure culture), and biostimulation (using CeO 2 or Nd 2 O 3 nanoparticles and endogenous microorganisms). Addition of M. extorquens AM1 in bioaugmentation experiment including 500 mg/L CeO 2 nanoparticles could not promote the nanoparticles accumulation in both natural and surface-sterilized treatments. However, it enhanced the translocation of Ce from roots to shoots in sterile samples. Moreover, another REE-utilizing bacterium, Bacillus subtilis, was enriched more than M. extorquens in control samples (no M. extorquens AM1), and associated with 52% and 14% higher Ce extraction in both natural (165 μg/g dried-plant ) and surface-sterilized samples (136 μg/g dried-plant ), respectively; showing the superior effect of endogenous microorganisms' enrichment over bioaugmentation in this experiment. In the biostimulation experiments, up to 705 μg/g dried-plant Ce and 19,641 μg/g dried-plant Nd could be extracted when 500 mg/L REE-NP were added. Furthermore, SEM-EDS analysis of the surface and longitudinal cross-sections of roots in Nd 2 O 3 treatments confirmed surface and intracellular accumulation of Nd 2 O 3 -NP. These results demonstrate stimulation of endogenous microbial community can lead to an enhanced REE phytoaccumulation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Pollution by heavy metals (HMs) has become a global problem for agriculture and the environment. In this study, the effects of pristine biochar and biochar modified with manganese dioxide (BC@MnO 2 ) and zinc oxide (BC@ZnO) nanoparticles on the immobilization and bioavailability of Pb, Cd, Zn, and Ni in soil under ryegrass (Lolium perenne L.) cultivation were investigated. The results of SEM-EDX, FTIR, and XRD showed that ZnO and MnO 2 nanoparticles were successfully loaded onto biochar. The results showed that BC, BC@MnO 2 and BC@ZnO treatments significantly increased shoots and roots dry weight of ryegrass compared to the control. The maximum dry weight of root and shoot (1.365 g pot -1 and 4.163 g pot -1 , respectively) was reached at 1% BC@MnO 2 . The HMs uptake by ryegrass roots and shoots decreased significantly after addition of amendments. The lowest Pb, Cd, Zn and Ni uptake in the plant shoot (13.176, 24.92, 32.407, and 53.88 µg pot -1 , respectively) was obtained in the 1% BC@MnO 2 treatment. Modified biochar was more successful in reducing HMs uptake by ryegrass and improving plant growth than pristine biochar and can therefore be used as an efficient and cost effective amendment for the remediation of HMs contaminated soils. The lowest HMs translocation (TF) and bioconcentration factors were related to the 1% BC@MnO 2 treatment. Therefore, BC@MnO 2 was the most successful treatment for HMs immobilization in soil. Also, a comparison of the TF values of plant showed that ryegrass had a good ability to accumulate all studied HMs in its roots, and it is a suitable plant for HMs phytostabilization., (© 2024. The Author(s).)

Complex rhizoremediation is the main mechanism of phytoremediation in organic-contaminated soil. Low molecular weight organic acids (LMWOAs) in root exudates have been shown to increase the bioavailability of contaminants and are essential for promoting the dissipation of contaminants. The effects of root exudates on the dissipation of organophosphate esters (OPEs) in soil are unclear. Consequently, we studied the combined effects of root exudates, soil enzymes and microorganisms on OPEs (tri (1-chloro-2-propyl) phosphate (TCPP) and triphenyl phosphate (TPP)) dissipation through pot experiments. Oxalic acid (OA) was confirmed to be the main component of LMWOAs in root exudates of ryegrass. The existence of OA increased the dissipation rate of OPEs by 6.04%-25.50%. Catalase and dehydrogenase activities were firstly activated and then inhibited in soil. While, urease activity was activated and alkaline phosphatase activity was inhibited during the exposure period. More bacteria enrichment (e.g., Sphingomonas, Pseudomonas, Flavisolibacter, Pontibacter, Methylophilus and Massilia) improved the biodegradation of OPEs. In addition, the transformation paths of OPEs hydrolysis and methylation under the action of root exudates were observed. This study provided theoretical insights into reducing the pollution risk of OPEs in the soil., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Given the high impact of traditional mining, the recovery of rare earth elements (REEs) from hazardous waste materials could become an option for the future in accordance with the principles of the circular economy. In this work, the technical feasibility of REEs recovery from metal mine tailings has been explored using electrokinetic-assisted phytoremediation with ryegrass (Lolium perenne L.). Phytoextraction combined with both AC current and DC current with reversal polarity was applied (1 V cm -1 , 8 h day -1 ) to real mine tailings containing a total concentration of REEs (Sc, Y, La, Ce, Pr, and Nd) of around 146 mg kg -1 . Changes in REEs geochemical fractionation and their concentrations in the soil pore water showed the mobilization of REEs caused by plants and electric current; REE availability was increased to a higher extent for combined electrokinetic-assisted phytoextraction treatments showing the relevant role of plants in the process. Our results demonstrated the initial hypothesis that it is feasible to recover REEs from real metal mining waste by phytoextraction and that the performance of this technology can be significantly improved by applying electric current, especially of the AC type, which increased REE accumulation in ryegrass in the range 57-68% as compared to that of the treatment without electric field application., (© 2024. The Author(s).)

Perennial ryegrass (Lolium perenne) is a widely used cool season turfgrass with outstanding turf quality and grazing tolerance. High temperature is the key factor restricting the distribution of perennial ryegrass in temperate and sub-tropic regions. In this study, we found that one HEAT SHCOK TRANSCRIPTION FACOTR (HSF) class A gene from perennial ryegrass, LpHSFA3, was highly induced by heat stress. LpHSFA3 is localized in nucleus and functions as a transcription factor. Ectopic overexpression of LpHSFA3 in Arabidopsis improved thermotolerance and rescued heat sensitive deficiency of athsfa3 mutant. Overexpression of LpHSFA3 in perennial ryegrass enhanced heat tolerance and increased survival rate in summer season as evidenced by decreased EL and MDA, increased number of green leaves and total chlorophyll content. LpHSFA3 binds to the HSE region in LpHSFA2a promoter to constitutively activate the expression of LpHSFA2a and downstream heat stress responsive genes. Ectopic overexpression of LpHSFA2a consequently rescued thermal sensitivity of athsfa3 mutant and enhanced thermotolerance of athsfa2 mutant. Perennial ryegrass protoplasts with overexpression of LpHSFA3 and LpHSFA2a exhibited induction of similar subsets of heat responsive genes. These results indicated that transcription factor LpHSFA3 functions as positive regulator of LpHSFA2a to improve thermotolerance of perennial ryegrass, providing further evidence to understand the regulatory networks of plant heat stress response., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Phytostabilization of metal-contaminated soils can be enabled or improved by biochar application. However, biochar-aided effects vary on biochar types, and little attention has been paid to plant management (time and cutting) to enhance phytostabilization efficiency in synergy with biochar. Therefore, biochars derived from pig manure (PM), Japanese knotweed (JK), and a mixture of both (P1J1) were applied to Pb and As mining soil with ryegrass cultivation to assess the biochar-induced effects on plant growth, dissolved organic matter (DOM), As and Pb mobility, and bioaccumulation within a phytostabilization strategy. Additional treatments involving the combined biochar (P1J1) and ryegrass were conducted to explore the influence of sequential cutting and growing time on facilitating phytostabilization efficacy. Biochar applications promoted plant growth, progressively increasing over time, but were not enhanced by cutting. Short and long-wavelength humic-like DOM substances identified in the soil pore water after biochar application varied depending on the biochar types used, providing evidence for the correlation among DOM changes, biochar origin, and metal immobilization. Biochar-treated soils exhibited reduced Pb availability and enhanced As mobility, with P1J1 stabilizing Pb significantly similar to PM while causing less As mobilization as JK did. The mobilized As did not result in increased plant As uptake; instead, all biochar-added plants showed a significant decrease in As and Pb concentrations compared to those without biochar. Soil available As decreased while available Pb increased with time, and cutting did not influence soil As behavior but did reduce soil Pb release. Nevertheless, plant As and Pb concentrations decreased over time, whereas those in multiple-cut plants were generally higher than those without cuts. Biochar, especially P1J1, along with growth time, holds promise in promoting plant biomass, reducing plant Pb and As concentrations, and minimizing the migration of PbAs within the soil., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous at relatively high concentrations by atmospheric deposition, and they are threatening to the environment. In this study, the toxicity of naphthalene on tall fescue and its potential responding mechanism was first studied by integrating approaches. Tall fescue seedlings were exposed to 0, 20, and 100 mg L -1 naphthalene in a hydroponic environment for 9 days, and toxic effects were observed by the studies of general physiological studies, chlorophyll fluorescence, and root morphology. Additionally, Ultra Performance Liquid Chromatography - Electrospray Ionization - High-Resolution Mass Spectrometry (UPLC-ESI-HRMS) was used to depict metabolic profiles of tall fescue under different exposure durations of naphthalene, and the intrinsic molecular mechanism of tall fescue resistance to abiotic stresses. Tall fescue shoots were more sensitive to the toxicity of naphthalene than roots. Low-level exposure to naphthalene inhibited the electron transport from the oxygen-evolving complex (OEC) to D1 protein in tall fescue shoots but induced the growth of roots. Naphthalene induced metabolic change of tall fescue roots in 12 h, and tall fescue roots maintained the level of sphingolipids after long-term exposure to naphthalene, which may play important roles in plant resistance to abiotic stresses., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

In agroecosystems, herbicides are the predominant anthropogenic selection pressure for agriculture weed species. While weeds are the primary target, herbicides can have adverse impacts on non-target plant beneficial microorganisms. We aimed to investigate the influence of a foliar endophytic fungus (Epichloë occultans) on the sensitivity of Lolium multiflorum to a graminicide herbicide (diclofop-methyl) during both plant ontogeny and progeny. Susceptible individuals to diclofop-methyl with and without endophyte were pre-exposed to the auxin 2,4-D herbicide. This herbicide is known to stimulate the metabolic detoxification mechanism (CYP-450) of diclofop-methyl. Regardless of the endophyte, 2,4-D pre-treatment increased mother plant survival to nearly 100 % under diclofop treatment but not in the progeny. Furthermore, maternal plant exposure to 2,4-D reduced endophyte transmission to the seeds and from seed-to-seedlings. Our findings suggest that, despite a reduction in diclofop-methyl sensitivity during the ontogeny of mother plants, 2,4-D-mediated induction of likely CYP-450 metabolism is not intergenerationally transmitted and shows detrimental effects on the symbiotic endophyte persistence., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Based on the growth-promoting effect of plant growth promoting bacteria on plants and the mobilization of Cd by citric acid, an experiment was designed in which the combined treatment of Bacillus megaterium and citric acid promoted ryegrass to repair Cd-contaminated soil. This study aimed to evaluate the effects of different treatments on the antioxidant enzyme activity, photosynthesis intensity, Cd accumulation, and rhizosphere cadmium migration under cadmium contamination conditions. And the soil morphology and structure changes were studied by infrared spectroscopy FourierTransformInfrared(FT-IR) and scanning electron microscope Energy Dispersive Spectrometer(SEM-EDS) before and after different treatments. The results show that the combined treatment of Bacillus megaterium and citric acid significantly improved the oxidative stress defense and plant photosynthesis and increased of rye biomass. rye biomass 1.28 times higher than CK treatment. Joint treatment significantly increased the amount of shoot accumulation of Cd, 2.31 times higher than CK treatment, increased the migration and accumulation of cadmium. FTIR and SEM-EDS also showed that the organic constituents such as O-H, C-O and C-N in soils as a major mechanism for mobilization of the heavy metal Cd. Thus, the combined treatment of Bacillus megaterium and citric acid can promote plant growth, improve the damage to ryegrass caused by single organic acid addition, and improve the plant extraction efficiency, which is a feasible way to repair Cd-contaminated soil through activated extraction system.

Italian ryegrass (Lolium multiflorum L.) is an invasive species widely spread in croplands worldwide. The intensive use of glyphosate has resulted in the selection of resistance to this herbicide in Italian ryegrass. This work characterized the response to glyphosate of Italian ryegrass populations from the South and Southwest regions of Paraná, Brazil. A total of 44 Italian ryegrass populations were collected in farming areas, and were classified for glyphosate resistance with 75% of populations resistant to gloyphosate. Of these, 3 resistant (VT05AR, MR20AR and RN01AR) and three susceptible (VT07AS, MR05AS and RN01AS) of these populations were selected to determine the resistance level and the involvement of the target site mechanisms for glyphosate resistance. Susceptible populations GR 50 ranged from 165.66 to 218.17 g.e.a. ha -1 and resistant populations from 569.37 to 925.94, providing RI ranging from 2.88 and 4.70. No mutation in EPSPS was observed in the populations, however, in two (MR20AR and RN02AR) of the three resistant populations, an increase in the number of copies of the EPSPs gene (11 to 57×) was detected. The number of copies showed a positive correlation with the gene expression (R 2  = 0.86) and with the GR 50 of the populations (R 2  = 0.81). The increase in EPSPS gene copies contributes to glyphosate resistance in Italian ryegrass populations from Brazil., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Perennial ryegrass (Lolium perenne L.) is an outstanding turfgrass and forage cultivated in temperate regions worldwide. However, poor tolerance to extreme cold, heat, or drought limits wide extension and cultivation. DEHYDRATION-RESPONSIVE ELEMENT BINDING FACTOR1s (DREB1s) play a vital role in enhancing plant tolerance to abiotic stress, specifically for low-temperature stress. In this study, a total of 24 LpDREB1 family members were identified from the released genome of perennial ryegrass. Phylogenetic analysis showed that the LpDREB1 genes are divided into 7 groups that have close relationships with rice homologues. Conserved motif analysis revealed that members within the same group have similar conserved motif compositions. All LpDREB1s lack introns, and the promoter sequences of LpDREB1 genes contain multiple cis-acting elements associated with stress response, phytohormone signal transduction and plant growth and development. The majority of LpDREB1 genes were upregulated by drought, submergence, heat and cold stress treatments, including LpDREB1H2. Further investigation showed that LpDREB1H2 is localized in the nucleus. Overexpression of LpDREB1H2 in Arabidopsis induced the expression of cold-responsive (COR) genes, increased the levels of osmotic adjusting substances, and enhanced antioxidant enzyme activities, thus improving the cold tolerance of Arabidopsis. This study lays a foundation for further understanding the function of LpDREB1 genes in perennial ryegrass and provides insights for plant stress tolerance breeding., (© 2024 Scandinavian Plant Physiology Society.)

Phosphorus (P) is critical to plants in metal-contaminated soils because it participates in various biochemical reactions during plant growth. However, the mechanisms of P in mitigating the toxicity of heavy metals to ryegrass root is still veiled. In this study, the physiological and biochemical dynamics of the ryegrass root under various cadmium (Cd) and P conditions were investigated in a hydroponic system. Cd stress decreased the length of the ryegrass root, but P application enhanced the root elongation to reduce the Cd concentration in the root. Both Cd and P dosages were positively correlated with hemicellulose 1 content, pectin content, and PME activity, while having a negative effect on cellulose content. Moreover, the addition of 80 mg L -1 P increased the contents of pectin and hemicellulose 1 by 2.5 and 5.8% even with 4 mg L -1 Cd. In addition, P supply increased pectin methylesterbase activity under Cd stress, which further changed the extra-cytoplasmic structures and cell wall composition. Thus, exogenous P promoted the immobilization of Cd onto the cell wall and protected protoplast primarily through indirectly regulating the binding capacity of the root cell wall for Cd., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Deyi Hou reports financial support was provided by National Key Research and Development Program of China. Hui Jia reports financial support was provided by National Natural Science Foundation of China. Zhengtao Shen reports financial support was provided by National Natural Science Foundation of China. Hui Jia reports financial support was provided by Natural Science Foundation of Jiangsu Province. Zhengtao Shen reports financial support was provided by Natural Science Foundation of Jiangsu Province. Hui Jia reports financial support was provided by China Postdoctoral Research Foundation., (Copyright © 2023 Elsevier Masson SAS. All rights reserved.)

In recent years, heavy metal pollution has become a global environmental problem and poses a great threat to the health of people and ecosystems. Therefore, strategies for the effective remediation of Cd from contaminated soil are urgently needed. In this study, ryegrass was utilized as a remediation plant, and its remediation potential was enhanced through the application of Citric Acid (CA) in conjunction with Bacillus megaterium (B. megaterium). The P3 treatment (CA + Bacillus megaterium) exhibited a significantly higher efficiency in promoting cadmium extraction by ryegrass, resulting in a 1.79-fold increase in shoot cadmium accumulation compared to the control group (CK) with no Bacillus megaterium or CA. Moreover, the P3 treatment led to an increased abundance of Actinobacteriota , Acidobacteriota , and Patescibacteria in the rhizosphere. The concentration of amino derivatives (such as betaine, sulfolithocholylglycine, N-alpha-acetyl-lysine, glycocholic acid, arginyl-threonine) showed significant upregulation following the P3 treatment. In summary, this study proposes a viable approach for phytoremediation of soil contaminated with cadmium by harnessing the mobilizing abilities of soil bacteria.

C-repeat binding factor (CBF) subfamily genes encoding transcriptional activators are members of the AP2/ERF superfamily. CBFs play important roles in plant tolerance to abiotic stress. In this study, we identified and analyzed the structure, phylogeny, conserved motifs, and expression profiles of 12 CBFs of the grass species Lolium perenne cultured under abiotic stress. The identified LpCBFs were grouped into three phylogenetic clades according to their protein structures and motif organizations. LpCBF expression was differentially induced by cold, heat, water deficit, salinity, and abscisic acid, among which cold treatment induced LpCBF gene expression significantly. Furthermore, association network analysis indicated that different proteins, including certain stress-related proteins, potentially interact with LpCBFs. Altogether, these findings will enhance our understanding of LpCBFs protein structure and function in the regulation of L. perenne stress responses. Our results will provide valuable information for further functional research of LpCBF proteins in L. perenne stress resistance.

Reclaimed water (RW) has been extensively used for irrigation in agriculture, yet the occurrence of antibiotics in real RW, and their toxicity, uptake dynamics and metabolic fate still needs comprehensive exploration. In this study, we investigated the residual concentrations of nineteen antibiotics in the RW from four wastewater treatment plants, and determined their toxicity on plant at environment-relevant concentration. Total found concentrations of these antibiotics ranged from 623.66 ng L -1 to 1536.96 ng L -1 , which decreased 10.3 and 19.4 % of roots' length and weight. Uptake dynamics analysis of the most hazardous antibiotic, norfloxacin (NFX) showed increasing amounts in the roots and leaves up to 3087.71 μg g -1 . Ryegrass also can remove >80 % of 100 μg L -1 NFX being achieved by biodegradation through ring cleavage, decarboxylation, defluorination, hydrogenation, methylation and oxidation. Toxicity assessment of the identified byproducts showed their more toxic effect on fish, daphnia and algae. This study extended our understanding of the fate of antibiotics in plants during irrigation with reclaimed water, and emphasized its safety and pollutants' biomagnification concerns., Competing Interests: Declaration of competing interest The authors declare no conflicts of interests., (Copyright © 2023 Elsevier B.V. All rights reserved.)

In order to investigate the impact of herbaceous root development on soil slope stability in expansive soil areas, the research was conducted in the soil slope experimental area of Yaoshi Town, Shangzhou District, Shangluo City. Three types of herbaceous plants, namely Lolium perenne, Medicago, and Cynodon dactylon, were planted to examine their influence on slope stability. The results indicated that Lolium perenne had significantly higher root length density and root surface area density compared to Cynodon dactylon and Medicago. However, the root weight density of Cynodon dactylon was found to be highest. The roots of Lolium perenne, Cynodon dactylon, and Medicago were predominantly observed in diameter ranges of 0 < L ≤ 1.0 mm, 0 < L ≤ 2.5 mm, and 2.5 < L ≤ 3.0 mm, respectively. The roots of herbaceous plants have the ability to enhance water retention in soil, resist hydraulic erosion of slope soil, and reduce soil shrinkage and swelling. During the initial phase of herbaceous planting, there is an accelerated process of organic carbon mineralization in the soil. The roots of herbaceous plants play a crucial role in soil consolidation and slope protection. They achieve this by dispersing large clastic particles, binding small particles together, altering soil porosity, enhancing soil water retention, and reducing soil water infiltration. It was found that Lolium perenne and Medicago, which have well-developed roots, exhibited superior slope protection effects. These findings contribute to the theoretical understanding for the implementation of green ecological protection technology on soil slopes., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Cao et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Lolium multiflorum grass is the major pollen allergen source in the southern region of Brazil, but most of its allergens remain poorly characterized. The aim of this study was to investigate antibody reactivity to L. multiflorum crude and carboxymethyl-ligand extracts in allergic patients and healthy individuals. Ion exchange carboxymethyl (CM) chromatography (CM-Sepharose) was used to isolate proteins (S2) from L. multiflorum crude extract (S1), which were assessed by SDS-PAGE. S1- and S2-specific IgE and IgG4 levels were measured by ELISA using sera from 55 atopic and 16 non-atopic subjects. Reactive polypeptide bands in S1 and S2 were detected by immunoblotting, and the most prominent bands in S2 were analyzed by mass spectrometry (MS-MS). Similar IgE and IgG4 levels were observed to both S1 (IgE median absorbance: 1.22; IgG4 median absorbance: 0.68) and S2 (IgE median absorbance: 1.26; IgG4 median absorbance: 0.85) in atopic subjects. S1 and S2 had positive correlations for IgE and IgG4 (IgE: r=0.9567; IgG4: r=0.9229; P<0.0001) levels. Homology between S1 and S2 was confirmed by IgE (84%) and IgG4 (83%) inhibition. Immunoblotting revealed that the 29-32 kDa band was recognized by 100% of atopic subjects in both S1 and S2. MS-MS analysis identified similarity profile to groups 1 and 5 grass allergens. This study revealed that carboxymethyl-ligand fraction played an important role for pollen allergy diagnosis by containing clinically relevant allergens and constituted a promising candidate for allergen-specific immunotherapy.

The application of exogenous growth-regulating substances is an effective technique to enhance plant stress tolerance. Here, a hydroponic experiment was conducted to investigate the effects of exogenous basal application of 0.1 mmol·L -1 spermidine (Spd) on both the physiology and molecular biology of ryegrass root systems under varying degrees (0, 5, and 10 mg·L -1 ) of cadmium (Cd) stress using ryegrass as the test plants. The results of physiological studies revealed that Cd stress significantly reduced the physiological functions of the ryegrass root system, whereas the addition of Spd effectively alleviated the negative effects caused by Cd. The most significant effect was on the root soluble protein content, which increased by 90.91% and 158.35% compared with 5 mg·L -1 and 10 mg·L -1 Cd alone. Spd also inhibited the accumulation of oxidative stress products malondialdehyde (MDA) and hydrogen peroxide (H 2 O 2 ) by increasing the ascorbic acid (ASA) and glutathione (GSH) content and peroxidase (POD) activity, whereas the effects on root activity and superoxide dismutase (SOD) activity were not significant. The results of molecular biology studies demonstrated that 10 mg·L -1 Cd stress caused differential expression of a large number of genes in ryegrass roots, and the number of differentially expressed genes, differential significance, and differential multiplicity were significantly reduced after the application of exogenous Spd. The most significant part of the GO enrichment analysis shifted from responding to organic cyclic compounds and aldehyde/ketone group transferase activity to responding to trivalent iron ions and 2'-deoxymugineic-acid 2'-dioxygenase activity. Single gene expression heat map analysis revealed that exogenous Spd upregulated the expression of genes encoding zinc-iron transporter protein and 2'-deoxymugineic-acid 2'-dioxygenase, which improved the uptake and utilization of iron by the root system. In conclusion, the application of certain concentrations of Spd could effectively regulate the response of ryegrass roots to Cd stress, enhance its tolerance physiology, and mitigate the toxic effects of Cd.

The various grass-induced epichloëcyclins of the Epichloë spp. are ribosomally synthesized and post-translationally modified peptides (RiPPs), produced as small, secreted cyclopeptides from a single gene, gigA. Here, four clustered and coregulated genes ( gigA , gigB , gigC , and kexB ) with predicted roles in epichloëcyclin production in Epichloë festucae were evaluated through gene disruption. Subsequent chemical analysis indicates that GigB is a DUF3328 domain-containing protein associated with cyclization of epichloëcyclins; GigC is a methyltransferase enzyme responsible for N-methylation of desmethylepichloëcyclins; and KexB is a subtilisin-like enzyme, partly responsible for the propeptide cleavage of epichloëcyclin intermediates. Symbiotic effects on the host phenotype were not observed for gigA , gigC , or kexB mutants, although Δ gigB infection correlated with increased host tiller height and biomass, while only Δ kexB exhibited an effect on endophyte morphology. Disrupting epichloëcyclin biosynthesis showed negligible influence on the biosynthesis of E. festucae -associated alkaloids. Epichloëcyclins may perform other secondary metabolism functions in Epichloë and other fungi.

The use of urine-derived fertilizers has several economic and environmental advantages. However, there is concern that pharmaceutical residues present in urine could enter the food chain after plant uptake and pose potential risks to human and animal health. A pot experiment was conducted to evaluate the uptake of nine target antiretroviral drugs (ARVDs) by pepper (Capsicum annum), ryegrass (Lolium perenne) and radish (Raphanus sativus) grown in two soils of contrasting texture and organic matter content and fertilized with stored urine, nitrified urine concentrate (NUC), and struvite. Nevirapine was the only ARVD detected in crops grown with NUC and struvite on both soils, but the concentrations were below the limit of quantification. Plants fertilized with stored urine absorbed lamivudine, ritonavir, stavudine, emtricitabine, nevirapine, and didanosine, while abacavir, efavirenz and zidovudine were not detected. The ARVDs detected in the soils after harvest were significantly higher in the soil with high organic matter and clay content. To assess direct human exposure the estimated daily dietary intake (DDI) of ARVDs by consumption of the pepper and radish fertilized with stored urine was compared with the Threshold of Toxicological Concern (TTC) values based on the Cramer classification tree. The calculated DDI values for all ARVDs were about 300-3000 times lower than the TTC values for class III compounds. Therefore, daily consumption of these crops fertilized with stored urine does not pose a health risk to the consumer. Future research is required to assess the impact of ARVD metabolites, which may be more harmful to human health than the parent compounds., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Cadmium (Cd) and polycyclic aromatic hydrocarbons (PAHs) are prominent soil contaminants found in industrial sites, and their combined effects on plants are not yet fully understood. To investigate the mechanisms underlying the co-exposure of Cd and PAHs and identify key biomarkers for their co-effects, an integrated analysis of metabolomics, transcriptomics, and proteomics was conducted on ryegrass leaves cultivated in soil. In nontarget metabolomics analysis, nine differentially expressed metabolites that were specifically induced by the compound exposure were identified. When combined with the analysis of differentially expressed genes and proteins, it was determined that the major pathways involved in the response to the co-stress of Cd and PAHs were linoleic acid metabolism and phenylpropanoid biosynthesis. The upregulation of 12,13-dihydroxy-9Z-octadecenoic acid and the downregulation of sinapyl alcohol were identified as typical biomarkers, respectively. Compared to scenarios of single exposures, the compound exposure to Cd and PAHs disrupted the oxidation of linoleic acid, leading to alterations in the profiles of linoleate metabolites. Additionally, it intensified hydroxylation, carboxylation, and methylation processes, and interfered with reactions involving coenzyme A, thus inhibiting lignin production. As a result, oxidative stress was elevated, and the cell wall defense system in ryegrass was weakened. The findings of this study highlight the ecological risks associated with unique biological responses in plants co-exposed to Cd and PAHs in polluted soils., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Endophytic fungi have been demonstrated to produce bioactive secondary metabolites, some of which promote plant growth. Three endophytic fungi isolated from healthy plants living in dehesas of Extremadura (Spain) were identified and evaluated for their ability to produce phytohormone-like substances, antioxidant activity, total polyphenol content, phosphate solubilization ability and siderophore and ammonia production. The filtrates and extracts produced by the three endophytes were applied to Lolium multiflorum seeds and seedlings under both in vitro and greenhouse conditions, to analyse their influence on plant growth traits such as germination, vigour index, chlorophyll data, number and length of leaves and roots, and dry weight. All three endophytes, which were identified as Fusarium avenaceum, Sarocladium terricola and Xylariaceae sp., increased the germination of L. multiflorum seeds by more than 70%. Shoot and root length, plant dry weight and the number of roots were positively affected by the application of fungal filtrates and/or extracts, compared with controls. The tentative HPLC-MS identification of phytohormone-like substances, such as gibberellin A2 and zeatin, or the antioxidant acetyl eugenol, may partially explain the mechanisms of L. multiflorum plant growth promotion after the application of fungal filtrates and/or extracts., (© 2023. The Author(s).)

Forage nutrient contents are an important factor explaining the dry matter intake (DMI), average daily gain (ADG), and methane emissions (CH 4 ) of ruminants fed indoors. However, for grazing animals, the forage nutrient contents might be limited in explaining such response variables. We aimed to verify the explanatory power of forage nutrient contents and sward structure on daily intake, performance, and CH 4 emissions by sheep and beef cattle grazing different grassland types in southern Brazil. We analyzed data from five grazing trials using sheep and beef cattle grazing on Italian ryegrass (Lolium multiflorum), mixed Italian ryegrass and black oat (Lolium multiflorum + Avena strigosa), pearl millet (Pennisetum americanum), and multispecies native grassland. We used mixed models, including the forage nutrient contents [crude protein (CP), neutral detergent fiber (NDF), and acid detergent fiber (ADF)], sward structure (sward height and herbage mass) and their interactions, as fixed effects and trial, season, methodologies, animal species, grassland type, and paddock, as random effects. The model for DMI (kg DM/LW 0.75 ) had an adjusted coefficient of determination (R 2 adj ) of 71.6 %, where 11.3, 23.1, and 37.2 % of the R 2 adj were explained by the forage nutrient contents, sward structure, and their interaction, respectively. The ADG (kg/LW 0.75 ) model presented an R 2 adj of 74.2 %, with 12.5 % explained by forage nutrient contents, 29.3 % by sward structure, and 32.4 % by their interaction. The daily CH 4 emission (g/LW 0.75 ) model had a lower adjusted coefficient of determination (R 2 adj = 47.6 %), with 16.8 % explained by forage nutrient contents and 30.8 % explained by sward structure, but no effect of the interaction. Our results show that in grazing ecosystems, the forage nutrient contents explain a small fraction, and the greater explanatory power for DMI, ADG, and CH 4 emissions models is related to sward structure descriptors, such as sward height and herbage mass. Moreover, the interaction between these variables explains most of the variation. In conclusion, forage nutrient contents and sward structure have different influences on DMI, ADG, and CH 4 emissions by grazing ruminants. Because of its relevance to daily CH 4 emissions, offering an optimal sward structure to grazing animals is a major climate-smart strategy to improve animal production and mitigate CH 4 emissions in pastoral ecosystems., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Perennial ryegrass is a grass species used to establish lawns in urban areas where pollution is a major environmental problems. Cadmium (Cd) and nickel (Ni) contribute significantly to these pollutants and may cause photosynthetic limitation. The main objective of this work was to perform a comprehensive analysis of photosynthetic efficiency of perennial ryegrass seedlings under Cd and Ni stress. Some of the main indices of photosynthetic efficiency (prompt and delayed chlorophyll-a fluorescence signals and modulated reflectance at 820 nm) were compared with growth parameters. Two cultivars were tested: 'Niga' and 'Nira'. A decrease in photosystem (PS) II and PSI activity was observed. This was due to an increase in nonradiative dissipation of the PSII antenna, a decrease in PSII antenna size, or a decrease in the number of photosynthetic complexes with fully closed PSII RCs. Efficiency of electron transport was decreased. The effect on the modulated reflectance signal could indicate a restriction in electron flow from PSII to PSI. The correlation between photosynthetic efficiency parameters, such as Area, Fo, Fm, and Fv, and growth parameters, confirmed that some photosynthetic efficiency parameters can be used as indicators for early detection of heavy metal effects., (© 2023. The Author(s).)

Sobemoviruses encode serine-like 3C proteases (Pro) that participate in the processing and maturation of other virus-encoded proteins. Its cis and trans activity is mediated by the naturally unfolded virus-genome-linked protein (VPg). Nuclear magnetic resonance studies show a Pro-VPg complex interaction and VPg tertiary structure; however, information regarding structural changes of the Pro-VPg complex during interaction is lacking. Here, we solved a full Pro-VPg 3D structure of ryegrass mottle virus (RGMoV) that demonstrates the structural changes in three different conformations due to VPg interaction with Pro. We identified a unique site of VPg interaction with Pro that was not observed in other sobemoviruses, and observed different conformations of the Pro β2 barrel. This is the first report of a full plant Pro crystal structure with its VPg cofactor. We also confirmed the existence of an unusual previously unmapped cleavage site for sobemovirus Pro in the transmembrane domain: E/A. We demonstrated that RGMoV Pro in cis activity is not regulated by VPg and that in trans , VPg can also mediate Pro in free form. Additionally, we observed Ca 2+ and Zn 2+ inhibitory effects on the Pro cleavage activity.

The effect of Pseudomonas aeruginosa (P. aeruginosa) on the phytoremediation efficiency of ryegrass on soil contaminated with nonylphenol (NP) and cadmium (Cd) was investigated by pot experiments. Pseudomonas aeruginosa application stimulated the adsorption of Cd by ryegrass and facilitated the biodegradation of NP in the soil. Exogenous P. aeruginosa inoculation increased the activities of urease, dehydrogenase, and polyphenol oxidase in the soil of the T4 treatment by 38.5%, 50.0%, and 56.5% compared to that of the T2 treatment, respectively. There was a significant positive correlation between the activities of dehydrogenase and polyphenol oxidase and the NP removal rate (P < 0.001). The relative abundances of beneficial microorganisms (such as Sphingomonas, Lysobacter, Streptomyces, Chloroflexia, Deltaproteobacteria, and Alphaproteobacteria) were increased as a result of P. aeruginosa inoculation. These microorganisms play important roles in nutrient cycling, Cd adsorption, and NP degradation. Additionally, P. aeruginosa was not the dominate bacterial species at the end of the experiment. According to this study, P. aeruginosa application improved the phytoremediation efficiency of ryegrass on soil contaminated with NP and Cd, with a minimal risk of alien microbial invasion., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Expansins are cell-wall loosening proteins involved in plant cell expansion and elongation. Objectives of this study were to identify expansins related to leaf elongation in a perennial grass species and determine the relationship between the expression of expansin genes and leaf elongation. A total of 20 expansin genes were identified in tall fescue (Festuca arundinacea), out of which nine genes belonged to the EXPA- and 11 to the EXPB subfamily. Two genotypes ("TF007" and "TF116") with different growth rates were used to determine the correlation between expansins and leaf growth. Among the 20 expansins, 16 were differentially expressed in the leaf growth zone in "TF007" and "TF116." The further analysis of gene expression in different leaf segments of "TF007" and "TF116" revealed that the expression level of FaEXPB16 was positively correlated with leaf elongation rate, and "TF007" had a higher leaf elongation rate than "TF116" due to the greater expression level of FaEXPB16. FaEXPA7 exhibited significantly higher expression level in leaves of the rapid-growing genotypes than the slow-growing genotypes, suggesting that FaEXPA7 acts as a positive regulator for leaf elongation. FaEXPA7 also exhibited its highest expression level in the cell division zone located in the leaf base. FaEXPB3, FaEXPB4-2, and FaEXPB11-2 showed a negative correlation with the leaf elongation rate in "TF007" and "TF116" and were highly expressed in leaves of the slow-growing genotypes. As promoting or repressing factors for leaf growth, these five expansins could be used as candidate genes in developing the rapid or slow-growing perennial grass species., (© 2023 Scandinavian Plant Physiology Society.)

A process of plant recovery after drought cessation is a complex trait which has not been fully recognized. The most important organ associated with this phenomenon in monocots, including forage grasses, is the crown tissue located between shoots and roots. The crown tissue is a meristematic crossroads for metabolites and other compounds between these two plant organs. Here, for the first time, we present a metabolomic and lipidomic study focused on the crown tissue under drought and recovery in forage grasses, important for agriculture in European temperate regions. The plant materials involve high (HDT) and low drought-tolerant (LDT) genotypes of Festuca arundinacea, and Lolium multiflorum/F. arundinacea introgression forms. The obtained results clearly demonstrated that remodeling patterns of the primary metabolome and lipidome in the crown under drought and recovery were different between HDT and LDT plants. Furthermore, HDT plants accumulated higher contents of primary metabolites under drought in the crown tissue, especially carbohydrates which could function as osmoprotectants and storage materials. On the other hand, LDT plants characterized by higher membranes damage under drought, simultaneously accumulated membrane phospholipids in the crown and possessed the capacity to recover their metabolic functions after stress cessation to the levels observed in HDT plants., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Heavy metals such as beryllium (Be) have been identified as toxic for plants with a negative impact on plant growth. Therefore, there is an urgent need for environmentally friendly techniques to reduce Be toxicity on plant growth and productivity. To this end, arbuscular mycorrhizal fungi (AMF) are widely applied to induce plant growth and stress tolerance. However, how AMF-plant symbiosis can support plants under Be stress has not been studied. Accordingly, we investigated the physiological and biochemical responses of AMF inoculated ryegrass and chickpea plants to Be stress. The associated changes in Be uptake and accumulation, photosynthesis, oxidative stress, carbon and nitrogen metabolism were studied. Soil contamination with Be induced higher Be accumulation, particularly in ryegrass, which consequentially reduced plant growth and photosynthesis. However, photorespiration and oxidative damage (H 2 O 2 accumulation, lipid oxidation, and LOX activity) were increased, mainly in ryegrass. In both plant species, AMF inoculation reduced Be accumulation and mitigated growth inhibition and oxidative damage, but to a more extent in ryegrass. This could be explained by improved photosynthesis as well as the upregulation of osmoprotectants i.e., sucrose and proline biosynthesis pathways. The increase in proline level was consistent with higher nitrogen (N) metabolism as reflected by N level and nitrate reductase. Species-specific responses were recorded and supported by principal component analysis. This study provided insight into the mechanism of AMF's impact on Be-stressed ryegrass and chickpea plants. Hence, the current research suggested that AMF inoculation could be used as a viable strategy to mitigate Be phytotoxicity in ryegrass and chickpea plants., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier Ltd.)

Perennial ryegrass (Lolium perenne) is an important turf grass and forage grass with moderately tolerant to salinity stress. Aspergillus aculeatus has been documented to involved in salt stress response of perennial ryegrass, while the A. aculeatus-mediated molecular mechanisms are unclear. Therefore, to investigate the molecular mechanisms underlying A. aculeatus-mediated salt tolerance, the comprehensive transcriptome analysis of the perennial ryegrass roots was performed. Twelve cDNA libraries from roots were constructed after 12 h of plant-fungus cocultivation under 300 mM salt stress concentrations. A total of 21,915 differentially expressed genes (DEGs) were identified through pairwise comparisons. Enrichment analysis revealed that potentially important A. aculeatus-induced salt responsive genes belonging to specific categories, such as hormonal metabolism (auxin and salicylic acid metabolism related genes), secondary metabolism (flavonoid's metabolism related genes) and transcription factors (MYB, HSF and AP2/EREBP family). In addition, weighted gene co-expression network analysis (WGCNA) showed that blue and black modules were significantly positively correlated with the peroxidase activity and proline content, then the hub genes within these two modules were further identified. Taken together, we found the categories of A. aculeatus-induced salt responsive genes, revealing underlying fungus-induced molecular mechanisms of salt stress response in perennial ryegrass roots. Besides, fungus-induced salt-tolerant hub genes represent a foundation for further exploring the molecular mechanisms., (© 2022 Scandinavian Plant Physiology Society.)

Biotechnological strategies have become effective in the remediation of polluted soils as they are cost-effective and do not present a risk of secondary pollution. However, using a single bioremediation technique (microorganism or plant) is not suitable for achieving a high remediation rate of polluted saline-alkali soils with heavy metals. Therefore, the present study aims to assess the effects and mechanisms of combined ryegrass and Fusarium incarnatum on the zinc (Zn)-polluted saline-alkali soil over 45 days. According to the obtained results, the combined Fusarium incarnatum-ryegrass showed the highest remediation rate of 49.35% after 45 days, resulting in a significantly lower soil Zn concentration than that observed in the control group. In addition, the inoculation of Fusarium incarnatum showed a positive effect on the soil EPS secretion. The soil protein contents ranged from 0.035 to 0.055 mg/kg, while the soil polysaccharide contents increased from 0.25 to 0.61 mg/g. The soil microbial flora and ryegrass showed resistance to saline and alkaline stresses through the secretion of extracellular polysaccharides. The three-dimensional fluorescence spectrum (3D-EEM) confirmed that EPS in the soil was mainly a fulvic acid-like substance. The fluorescein diacetate (FDA) hydrolase activity in the saline-alkali soil was first increased due to the effect of Fusarium incarnatum and then decreased to a minimum value of 96 μg/(g·h). In addition, the Fusarium incarnatum inoculation improved the diversity and richness of soil fungi. Although the Fusarium incarnatum inoculation had a slight effect on the germination of ryegrass, it increased the biomass and enrichment coefficient. The results revealed a translocation factor (TF) value of 0.316 at 45 days after ryegrass sowing, showing significant enrichment of the soil Zn heavy metal zinc in the ryegrass roots., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier Ltd.)

Perennial ryegrass (Lolium perenne) is a cool-season turf and forage grass. Heat shock transcription factors (HSFs) play an important role in regulating plant abiotic stress. However, HSFs in perennial ryegrass have rarely been characterized. Here, 25 LpHSFs were identified from the perennial ryegrass genome. Phylogenetic analysis showed that the LpHSFs could be classified into 12 subclasses. Gene structure analysis showed that 22 LpHSFs have only one intron. Cis-acting elements analysis revealed that the promoter of 15 LpHSFs contained hormone-responsive and abiotic stress-responsive elements. Expression profile analysis indicated that 24 LpHSFs were differentially expressed under submerge, drought, heat, and cold stresses. In addition, a subclass C2 gene, LpHSFC2b, was significantly induced by abiotic stresses. The LpHSFC2b protein is localized to the nucleus, and heterologous expression of LpHSFC2b in Arabidopsis improves plant thermotolerance. This study provides insights useful for the breeding of stress tolerance in perennial ryegrass., (© 2022 Scandinavian Plant Physiology Society.)

Under favourable conditions, perennial ryegrass (Lolium perenne) engineered to accumulated high lipid (HL) carbon sink in their leaves was previously shown to also enhance photosynthesis and growth. The greater aboveground biomass was found to be diminished in a dense canopy compared to spaced pots. Besides, the underlying genetic regulatory network linking between leaf lipid sinks and these physiological changes remains unknown. In this study, we demonstrated that the growth advantage was not displayed in HL Lolium grown in spaced pots under low lights. Under standard lights, analysis of differentiating transcripts in HL Lolium reveals that the plants had elevated transcripts involved in lipid metabolism, light capturing, photosynthesis, and sugar signalling while reduced expression of genes participating in sugar biosynthesis and transportation. The plants also had altered several transcripts involved in mitochondrial oxidative respiration and redox potential. Many of the above upregulated or downregulated transcript levels were found to be complemented by growing the plants under low light. Overall, this study emphasizes the importance of carbon and energy homeostatic regulatory mechanisms to overall productivity of the HL Lolium through photosynthesis, most of which are significantly impacted by low irradiances., Competing Interests: The authors have declared that no competing interests exist.

The magnetic field can alter the hydrogen-bond structure and polarity characteristics of water; therefore, we hypothesize that magnetized water can affect plant physiological functions, including metal detoxification and excretion. In this study, the amount of Cd excreted on the leaves of Festuca arundinacea was estimated using magnetized water and normal water irrigation patterns. Irrigation with magnetized water improved the shoot dry weight and Cd content in F. arundinacea by 13.6% and 52.8%, respectively, compared to the control. Magnetized water irrigation also increased antioxidant enzyme activities in plant leaves, thereby alleviating the oxidative damage. The concentration of ferritin was 0.91 folds higher than that of the control, increasing the Fe sequestration and detoxification capacity of F. arundinacea. The amount of Cd excreted was significantly higher under magnetized water irrigation, thereby increasing the annual Cd removal by 109.7% from soil by leaf washing compared with that of the control. In contrast, F. arundinacea irrigated with magnetized water excreted 38.1% less Fe owing to the increase in ferritin levels, compared with that of the control. This study suggests a novel pathway of Cd phytoremediation by rinsing excreted Cd from the leaf surface without harvesting and replanting F. arundinacea., (Copyright © 2022 Elsevier B.V. All rights reserved.)