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Utilizing and improving the productivity of reclaimed land are highly significant for alleviating the problem of food production shortage in China, and the integrated rice-frog farming model can improve soil fertility. However, there are few studies on the use of integrated rice-frog farming technology to improve the fertility of reclaimed land and increase its efficiency in food production. Therefore, this study was conducted to evaluate the effects of the rice-frog co-cropping mode on the soil fertility and microbial diversity of reclaimed land. A rice monoculture group (SF), low-density rice-frog co-cropping group (SD, 5000 frogs/mu, corresponds to 8 frogs/m2), and high-density rice-frog co-cropping group (SG, 10,000 frogs/mu, corresponds to 15 frogs/m2) were established and tested. The contents of total nitrogen, soil organic matter, available potassium, and available phosphorus of the soil in the SG group were significantly higher than those in the SF group (p < 0.05) in the mature stage of rice. Compared with the SF group, the SD and SG groups improved the soil microbial diversity and changed the structure of the microbial community. This study indicates that compared with the rice monoculture mode, the rice-frog co-cropping pattern can improve the soil fertility, as well as microbial diversity, of reclaimed land.

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Plants synthesize a plethora of chemical defence compounds, which vary between evolutionary lineages. We hypothesize that plants evolved the ability to utilize defence compounds synthesized and released by neighbouring heterospecific plants. In two experiments, we incubated clover (Trifolium repens L.) seedlings with individual benzoxazinoid (BX) compounds (2,4-dihydroxy-1,4-benzoxazin-3-one, 2-hydroxy-1,4-benzoxazin-3-one, benzoxazolinone, and 6-methoxy- benzoxazolin-2-one), a group of bioactive compounds produced by cereals, to allow clover BX uptake. Subsequently, we transplanted the seedlings into soil and quantified BX root and shoot content and invasion of root-knot nematodes in clover roots up to 8 weeks after transplantation. We show that clover root uptake of BXs substantially enhanced clover's resistance against the root-knot nematode Meloidogyne incognita. This effect lasted up to 6 weeks after the clover roots were exposed to the BXs. BXs were absorbed by clover roots, and then translocated to the shoots. As a result of clover metabolization, we detected the parent BXs and a range of their transformation products in the roots and shoots. Based on these novel findings, we envisage that co-cultivation of crop species with complementary and transferable chemical defence systems can add to plant protection.

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Mine driven trace elements' pollution entails environmental risks and causes soil infertility. In the last decades, in situ techniques such as phytostabilization have become increasingly important as ways to tackle these negative impacts. The aim of this study was to test the individual and combined effects of different aided phytostabilization techniques using substrate from barren tailings of a Cu mine, characterized by extreme infertility (high acidity and deficiency of organic matter and nutrients). The experiment analyzed the growth of Populus nigra L. planted alone (P) or in co-cropping with Trifolium repens L. (PT), in pots containing mine soil amended with compost (1, 10, compost, soil, w/w) non inoculated (NI) or inoculated with plant growth promoting rhizobacteria (PGP), mycorrhizae (MYC) or a combination of bacterial and fungal inocula (PGPMYC). Non-amended, non-planted and non-inoculated reference ports were also prepared. Plants were harvested after 110 days of plant development and several biometric and phytopathological parameters (stem height, aerial biomass, root biomass, wilting, chlorosis, pest and death) and macro and micronutrient composition were determined. The growth substrate was analyzed for several physicochemical (pH, CECe, and exchangeable cations, total C and N, P Olsen and availability of trace elements) and microbiological (community level physiological profiles: activity, richness and diversity) parameters. The use of the amendment, P. nigra plantation, and inoculation with rhizobacteria were the best techniques to reduce toxicity and improve soil fertility, as well as to increase the plant survival and growth. Soil bacterial functional diversity was markedly influenced by the presence of plants and the inoculation with bacteria, which suggests that the presence of plant regulated the configuration of a microbial community in which the inoculated bacteria thrive comparatively better. The results of this study support the use of organic amendments, tolerant plants, and plant growth promoting rhizobacteria to reduce environmental risk and improve fertility of soils impacted by mining.

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Phytoremediation shows great promise as a plant-based alternative to conventional clean-up methods that are prohibitively expensive. As part of an integrated strategy, the selection of well-adapted plant species as well as planting and management techniques could determine the success of a long-term program. Herein, we conducted an experiment under semi-controlled conditions to screen different plants species with respect to their ability to phytoremediate Zn-contaminated soil excavated from a contaminated site following a train derailment and spillage. The effect of nitrilotriacetic acid (NTA) application on the plants and soil was also comprehensively evaluated, albeit we did not find its use relevant for field application. In less than 100 days, substantial Zn removal occurred in the soil zone proximal to the roots of all the tested plant species. Three perennial herbaceous species were tested, namely, Festuca arundinacea, Medicago sativa, and a commercial mix purposely designed for revegetation; they all showed strong capacity for phytostabilization at the root level but not for phytoextraction. The Zn content in the aboveground biomass of willows was much higher. Furthermore, the degree of growth, physiological measurements, and the Zn extraction yield indicated Salix purpurea 'Fish Creek' could perform better than Salix miyabeana, 'SX67', in situ. Therefore, we suggest implementing an S. purpurea­perennial herbaceous co-cropping strategy at this decade-long-abandoned contaminated site or at similar disrupted landscapes.

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The rice-crayfish co-cropping pattern is a traditional method for the intensive utilization of rice fields. In recent years, this pattern has been over-developed in many countries and regions, especially in China, because of its simple agronomic technology and high economic benefits. However, little is known about the potential ecological problems regarding soil microorganisms caused by the over-utilization of this pattern. The results show that rice-crayfish co-cropping, when over-utilized for a long time, reduced soil microbial richness and diversity compared with rice monocropping. A decrease in bacterial abundance in the nitrogen cycle and an increase in bacterial abundance in the carbon cycle led to a decrease in the nitrogen cycle function and an increase in the carbon cycle function. In an analysis of bacteria that are sensitive to cropping patterns, it was found that in the rice-crayfish co-cropping, the relative abundances of sensitive OTUs from Firmicutes (Bacillus and Clostridium) and Chloroflexi (Anaerolineaceae) were significantly higher during the entire growth period than those observed in the rice monocropping pattern, while the relative abundances of sensitive OTUs from Nitrospirae (Nitrospira), Gemmatimonadetes (Gemmatimonas), and Actinobacteria (Nocardioides) were significantly lower than those observed in the rice monocropping pattern. A network analysis shows that growth-period-sensitive OTUs drive the co-occurrence network modules, although the OTUs also have positive and negative correlations among modules but a positive synergistic effect on the regulation of soil nutrients. In addition, OTUs that were sensitive at the booting stage and filling stage were the key microbial groups in the rice-crayfish co-cropping and rice monocropping networks, respectively. Understanding the classifications and functions of sensitive microbes present during the rice growth period is the basis for formulating a microbial flora management strategy for the rice-crayfish co-cropping pattern, which is of great significance for adjusting agricultural management measures and controlling current soil microbial ecological problems.

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Co-cropping is an eco-friendly strategy to improve the phytoremediation capacity of plants growing in soils contaminated with heavy metals such as cadmium (Cd). This study was conducted to investigate the effects of co-cropping Indian mustard (Brassicajuncea) and silage maize (Zeamays) and applying peat on the phytoremediation of a Cd-contaminated acid paddy soil via characterizing plant growth and Cd uptake in pot experiments. There were six planting patterns (Control: no plants; MI-2 and MI-4: mono-cropping of Indian mustard at low and high densities, respectively; MS: mono-cropping of silage maize; CIS-2 and CIS-4: co-cropping of Indian mustard at low and high densities with silage maize, respectively) and two application rates of peat (NP: 0; WP: 30 g kg-1). When Indian mustard and silage maize were co-cropped, the shoot biomass of Indian mustard plants per pot was significantly (p < 0.05) lower than that obtained in the mono-cropping systems, with a substantial reduction (55-72%) in the same plant density group. The shoot biomass of silage maize plants in the mono-cropping systems did not differ significantly from that in the co-cropping systems regardless of the density of Indian mustard. The growth-promoting effect of the peat application was more pronounced in Indian mustard than silage maize. Under the low density of Indian mustard, the co-cropping systems significantly (p < 0.05) decreased Cd uptake by silage maize. Additionally, soil amendment with peat significantly (p < 0.05) increased shoot Cd removal rate and Cd translocation factor value in the co-cropping systems. Taken together, the results demonstrated that silage maize should be co-cropped with Indian mustard at an appropriate density in Cd-polluted soils to achieve simultaneous remediation of Cd-contaminated soils (via Indian mustard) and production of crops (here, silage maize). Peat application was shown to promote the removal of Cd from soil and translocation of Cd into shoots and could contribute to enhanced phytoremediation of Cd-contaminated acid paddy soil.

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Phytoremediation coupled with co-cropping is assumed to be good for safety utilization and remediation of heavy metal contaminated farmland, which can ensure farmers' income without increasing health risks for human. In this study, the effects on plant cadmium (Cd) accumulation and health risk of consuming the vegetable plant were compared between monoculture and co-cropping of cauliflower (Brassica oleracea) with two ecotypes of Sedum alfredii in a moderately (0.82 mg kg-1) Cd contaminated greenhouse vegetable field. The results showed that co-cropping with S. alfredii raised Cd concentration in edible part of cauliflower with slightly growth promotion. The health risk of consuming cauliflower to different groups of people have been evaluated by calculating Hazard Quotient (HQ) and all HQ value were less than 1.0, which indicated that eating co-cropped cauliflower would not cause health risks to adults and children. Besides, the Cd concentration of hyperaccumulating ecotype (HE) of S. alfredii was 27.3 mg kg-1 in monoculture and it increased to 51.2 mg kg-1 after co-cropping with cauliflower, suggesting that the co-cropping system promoted HE Cd absorption capacity. Therefore, the "Phytoextraction Coupled with Agro-safe-production" (PCA) model of cauliflower and HE can serve as an alternative sustainable strategy in the Cd moderate polluted greenhouse.

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Arbutus unedo (the strawberry tree) is a Mediterranean shrub which forms arbutoid mycorrhizae with a variety of Asco- and Basidiomycetes. After the discovery of the mycorrhizal symbiosis between A. unedo and Tuber borchii, in this study, arbutoid mycorrhizae were synthetized in greenhouse with Tuber aestivum and Tuber melanosporum. Six months after inoculation, both species colonized the roots of all inoculated A. unedo seedlings, but mature mycorrhizae were only observed after 12 months. Ultrastructure analysis of Tuber arbutoid mycorrhizae was described for the first time, showing, as observed in typical endosymbiosis, a rearrangement of host cells and the creation of an interface compartment with both truffle species. Immunolabelling experiments suggested that pectins are not present in the interface matrix surrounding the intracellular hyphae. Thus, the ability to establish symbiosis with A. unedo seems to be a common feature in the genus Tuber, opening up the possibility to use this plant for mycorrhization with valuable truffles. This could represent an important economic opportunity in Mediterranean areas by combining the production of truffles, edible fruits and valued honey.

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Phytoremediation by co-cropping may be a promising approach to produce safe crops while remediating the soil. However, the effects of plant interaction, especially stress response, remain unclear. The aims of this study were to investigate the effect of co-cropping on plant growth, stress response and lead (Pb) uptake in soybean and Tagetes minuta, and to assess the feasibility of agricultural production in Pb-polluted soils. A pot experiment was conducted to study the effect of co-cropping vs monocrop at three soil Pb concentrations. The following parameters were analyzed: biomass, Pb content in plants, and stress response indicators (chlorophylls, proteins, sugars, malondialdehyde, glutathione S-transferase activity, carotenes and antioxidant power). Results showed that in co-cropping, both species were benefited in polluted soils, since biomass and stress response were improved. T. minuta reduced adverse effects of Pb on soybean by improving grain quality and even survival in polluted soils, where soybean in monocrop grew only up to early vegetative stages. This effect was related to a 50% reduction in lipid peroxidation for soybean in co-cropping along with a sharp increase in the antioxidant response. In addition, co-cropping enhanced Pb accumulation in T. minuta (45% higher), as well as content of chlorophylls and carotenes (66% and 42% of increment, respectively) and glutathione S-transferase activity (two times higher) in the highly polluted soil. Our results showed that rhizosphere interactions can help enhance tolerance to Pb toxicity in both species, allowing soybean production in highly polluted soils without posing health risk from grain consumption.

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Human industrial activities have left millions of hectares of land polluted with trace element metals and persistent organic pollutants (POPs) around the world. Although contaminated sites are environmentally damaging, high economic costs often discourage soil remediation efforts. Phytoremediation is a potential green technology solution but can be challenging due to the diversity of anthropogenic contaminants. Co-cropping could provide improved tolerance to diverse soil challenges by taking advantage of distinct crop capabilities. Co-cropping of three species with potentially complementary functions, Festuca arundinacea, Salix miyabeana and Medicago sativa, perform well on diversely contaminated soils. Here, rhizosphere microbiomes of each crop in monoculture and in all co-cropping combinations were compared using 16S rRNA gene amplification, sequencing and differential abundance analysis. The hyperaccumulating F. arundinacea rhizosphere microbiome included putative plant growth promoting bacteria (PGPB) and metal tolerance species, such as Rhizorhapis suberifaciens, Cellvibrio fibrivorans and Pseudomonas lini. The rhizosphere microbiome of the fast-growing tree S. miyabeana included diverse taxa involved in POP degradation, including the species Phenylobacterium panacis. The well-characterised nitrogen-fixing M. sativa microbiome species, Sinorhizobium meliloti, was identified alongside others involved in nutrient acquisition and putative yet-to-be-cultured Candidatus saccharibacteria (TM7-1 group). The majority of differentially abundant rhizosphere-associated bacterial species were maintained in co-cropping pairs, with pairs having higher numbers of differentially abundant taxa than monocultures in all cases. This was not the case when all three crops were co-cropped, where most host-specific bacterial species were not detected as differentially abundant, indicating the potential for reduced rhizosphere functionality. The crops cultivated in pairs here retained rhizosphere microbiome bacteria involved in these monoculture ecosystem services of plant growth promotion, POP tolerance and degradation, and improved nutrient acquisition. These findings provide a promising outlook of the potential for complementary co-cropping strategies for phytoremediation of the multifaceted anthropogenic pollution which can disastrously affect soils around the world.

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Truffles are sequestrate hypogeous fungi, and most form ectomycorrhizal (ECM) associations with trees. Truffles belonging to the genus Tuber (Pezizales, Ascomycota), "true truffles," associate with diverse plant hosts, including economically important species such as pecan (Carya illinoinensis). Morphological and phylogenetic studies delimited several major lineages of Tuber, which include many cryptic and undescribed species. One of these, the Maculatum clade, is a speciose group characterized by relatively small, light-colored ascomata that have alveolate-reticulate spores. Here, we describe two new species in the Maculatum clade, Tuber brennemanii and T. floridanum (previously identified as Tuber sp. 36 and Tuber sp. 47). We delineate these two species by phylogenetic analyses of nuc ITS1-5.8S-ITS2 (= ITS) and partial 28S rDNA (= LSU), and through morphological analysis. A recent collection of T. floridanum from a pecan orchard in Brazil indicates that this species was introduced there on the roots of pecan seedlings. Systematic studies of ascomata and ECM fungal communities indicate that these species are geographically widespread and common ECM symbionts of pecans and other members of the Fagales, particularly in sites with disturbed soils and nutrient enrichment.

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The genus Alyssum (Brassicaceae) contains Ni hyperaccumulators (50), many of which can achieve 30 g kg(-1) Ni in dry leaf. Some Alyssum hyperaccumulators are viable candidates for commercial Ni phytoremediation and phytomining technologies. It is not known whether these species secrete organic and/or amino acids into the rhizosphere to solubilize Ni, or can make use of such acids within the soil to facilitate uptake. It has been hypothesized that in fields with mixed plant species, mobilization of metals by phytosiderophores secreted by Graminaceae plants could affect Alyssum Ni, Fe, Cu, and Mn uptake. We co-cropped the Ni hyperaccumulator Alyssum murale, non-hyperaccumulator A. montanum and perennial ryegrass in a natural serpentine soil. All treatments had standard inorganic fertilization required for ryegrass growth and one treatment was compost amended. After 4 months A. murale leaves and stems contained 3600 mg kg(-1) Ni which did not differ significantly with co-cropping. Overall Ni and Mn concentrations were significantly higher in A. murale than in A. montanum or L. perenne. Copper was not accumulated by either Alyssum species, but L. perenne accumulated up to 10 mg kg(-1). A. montanum could not compete with either A. murale or ryegrass, and neither Alyssum species survived in the compost-amended soil. Co-cropping with ryegrass reduced Fe and Mn concentrations in A. murale but not to the extent of either increasing Ni uptake or affecting plant nutrition. The hypothesized Alyssum Ni accumulation in response to phytosiderophores secreted by co-cropped grass did not occur. Our data do not support increased mobilization of Mn by a phytosiderophore mechanism either, but the converse: mobilization of Mn by the Alyssum hyperaccumulator species significantly increased Mn levels in L. perenne. Tilling soil to maximize root penetration, adequate inorganic fertilization and appropriate plant densities are more important for developing efficient phytoremediation and phytomining approaches.

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Polluted agricultural soils are a serious problem for food safety, with phytoremediation being the most favorable alternative from the environmental perspective. However, this methodology is generally time-consuming and requires the cessation of agriculture. Therefore, the purpose of this study was to evaluate two potential phytoextractor plants (the native species Bidens pilosa and Tagetes minuta) co-cropped with lettuce growing on agricultural lead-polluted soils. The concentrations of Pb, as well as of other metals, were investigated in the phytoextractors, crop species, and in soils, with the potential risk to the health of consumers being estimated. The soil parameters pH, EC, organic matter percentage and bioavailable lead showed a direct relationship with the accumulation of Pb in roots. In addition, the concentration of Pb in roots of native species was closely related to Fe (B. pilosa, r = 0.81; T. minuta r = 0.75), Cu (T. minuta, r = 0.93), Mn (B. pilosa, r = 0.89) and Zn (B. pilosa, r = 0.91; T. minuta, r = 0.91). Our results indicate that the interaction between rhizospheres increased the phytoextraction of lead, which was accompanied by an increase in the biomass of the phytoextractor species. However, the consumption of lettuce still revealed a toxicological risk from Pb in all treatments.

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In a field experiment on multi-metal contaminated soil, we investigated the efficiency of Cd, Pb, Zn, and Cu removal by only mixture of chelators (MC) or combining with FeCl3. After washing treatment, a co-cropping system was performed for heavy metals to be extracted by Sedum alfredii and to produce safe food from Zea mays. We analyzed the concentration of heavy metals in groundwater to evaluate the leashing risk of soil washing with FeCl3 and MC. Results showed that addition of FeCl3 was favorable to the removal of heavy metals in the topsoil. Metal leaching occurred mainly in rain season during the first co-cropping. The removal rates of Cd, Zn, Pb, and Cu in topsoil were 28%, 53%, 41%, and 21% with washing by FeCl3+MC after first harvest. The application of FeCl3 reduced the yield of S. alfredii and increased the metals concentration of Z. mays in first harvest. However, after amending soil, the metals concentration of Z. mays in FeCl3+MC treatment were similar to that only washing by MC. The grains and shoots of Z. mays were safe for use in feed production. Soil washing did not worsen groundwater contamination during the study period. But the concentration of Cd in groundwater was higher than the limit value of Standard concentrations for Groundwater IV. This study suggests that soil washing using FeCl3 and MC for the remediation of multi-metal contaminated soil is potential feasibility. However, the subsequent measure to improve the washed soil environment for planting crop is considered.

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The presence and composition of soil microbial communities has been shown to have a large impact on plant-plant interactions and consequently plant diversity and composition. The goal of the present study was to evaluate impact of arbuscular mycorrhizal fungi (AMF) and nitrogen-fixing bacteria, which constitutes an essential link between the soil and the plant's roots. A greenhouse pot experiment was conducted to evaluate the feasibility of using selected microbes to improve Hieracium pilosella and Medicago sativa growth on Zn-Pb-rich site. Results of studies revealed that biomass, the dry mass of shoots and roots, increased significantly when plants were inoculated with mycorrhizal fungi and nitrogen-fixing bacteria. The addition of Azospirillum sp. and Nostoc edaphicum without mycorrhiza suppressed plant growth. Single bacterial inoculation alone does not have a positive effect on M. sativa growth, while co-inoculation with AMF improved plant growth. Plant vitality (expressed by the performance index) was improved by the addition of microbes. However, our results indicated that even dry heat sterilization of the substratum created imbalanced relationships between soil-plant and plants and associated microorganisms. The studies indicated that AMF and N2-fixers can improve revegetation of heavy metal-rich industrial sites, if the selection of interacting symbionts is properly conducted.

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