Nitrogen fertilization modulates rice phyllosphere functional genes and pathogens through fungal communities.

The phyllosphere is a vital yet often neglected habitat hosting diverse microorganisms with various functions. However, studies regarding how the composition and functions of the phyllosphere microbiome respond to agricultural practices, like nitrogen fertilization, are limited. This study investigated the effects of long-term nitrogen fertilization with different levels (CK, N90, N210, N330) on the functional genes and pathogens of the rice phyllosphere microbiome. Results showed that the relative abundance of many microbial functional genes in the rice phyllosphere was significantly affected by nitrogen fertilization, especially those involved in C fixation and denitrification genes. Different nitrogen fertilization levels have greater effects on fungal communities than bacteria communities in the rice phyllosphere, and network analysis and structural equation models further elucidate that fungal communities not only changed bacterial-fungal inter-kingdom interactions in the phyllosphere but also contributed to the variation of biogeochemical cycle potential. Besides, the moderate nitrogen fertilization level (N210) was associated with an enrichment of beneficial microbes in the phyllosphere, while also resulting in the lowest abundance of pathogenic fungi (1.14 %). In contrast, the highest abundance of pathogenic fungi (1.64 %) was observed in the highest nitrogen fertilization level (N330). This enrichment of pathogen due to high nitrogen level was also regulated by the fungal communities, as revealed through SEM analysis. Together, we demonstrated that the phyllosphere fungal communities were more sensitive to the nitrogen fertilization levels and played a crucial role in influencing phyllosphere functional profiles including element cycling potential and pathogen abundance. This study expands our knowledge regarding the role of phyllosphere fungal communities in modulating the element cycling and plant health in sustainable agriculture.

Seeds Act as Vectors for Antibiotic Resistance Gene Dissemination in a Soil-Plant Continuum.

Though the evidence for antibiotic resistance spread via plant microbiome is mounting, studies regarding antibiotic resistome in the plant seed, a reproductive organ and important food resource, are still in their infancy. This study investigated the effects of long-term organic fertilization on seed bacterial endophytes, resistome, and their intergenerational transfer in the microcosm. A total of 99 antibiotic resistance genes (ARGs) and 26 mobile genetic elements (MGEs) were detected by high-throughput quantitative PCR. The amount of organic fertilizer applied was positively correlated to the number and relative abundance of seed-associated ARGs and MGEs. Moreover, the transmission of ARGs from the rhizosphere to the seed was mainly mediated by the shared bacteria and MGEs. Notably, the rhizosphere of progeny seedlings derived from seeds harboring abundant ARGs was found to have a higher relative abundance of ARGs. Using structural equation models, we further revealed that seed resistome and MGEs were key factors affecting the ARGs in the progeny rhizosphere, implying the seed was a potential resistome reservoir for rhizosphere soil. This study highlights the overlooked role of seed endophytes in the dissemination of resistome in the soil-plant continuum, and more attention should be paid to plant seeds as vectors of ARGs within the "One-Health" framework.

Phloem iron remodels root development in response to ammonium as the major nitrogen source.

Plants use nitrate and ammonium as major nitrogen (N) sources, each affecting root development through different mechanisms. However, the exact signaling pathways involved in root development are poorly understood. Here, we show that, in Arabidopsis thaliana, either disruption of the cell wall-localized ferroxidase LPR2 or a decrease in iron supplementation efficiently alleviates the growth inhibition of primary roots in response to NH4+ as the N source. Further study revealed that, compared with nitrate, ammonium led to excess iron accumulation in the apoplast of phloem in an LPR2-dependent manner. Such an aberrant iron accumulation subsequently causes massive callose deposition in the phloem from a resulting burst of reactive oxygen species, which impairs the function of the phloem. Therefore, ammonium attenuates primary root development by insufficiently allocating sucrose to the growth zone. Our results link phloem iron to root morphology in response to environmental cues.

Variations of earthworm gut bacterial community composition and metabolic functions in coastal upland soil along a 700-year reclamation chronosequence.

Most ecosystem functions attributed to earthworms are mediated by their internal microbiomes, and these are sensitive to disturbances in the external environment. However, few studies have focused on the response of the earthworm gut microbiome to soil chronosequence. Here, we used 16S rRNA high-throughput sequencing and high-throughput quantitative PCR to investigate the variations in bacterial communities and functional gene abundance in earthworm (Lumbricina sp.) guts and upland soils under 700 years of cultivation. Our results indicated that 700 years of upland cultivation significantly shaped bacterial communities and increased functional traits of microbes in earthworm guts, which were more sensitive to cultivation age compared to the surrounding soils. The earthworm gut bacterial community changed rapidly over the first 300 years of cultivation and then changed slowly in the following centuries. Along with the cultivation age, we also observed that the earthworm gut microbiota was successive towards a copiotrophic strategy (e.g., Xanthobacteraceae, Nocardioidaceae, Hyphomicrobiaceae, and Bacillaceae) and higher potential functions (e.g., ureC, nirS, nosZ, phoD, and pqqC). Furthermore, canonical correspondence analysis further revealed that soil pH, C:N ratio, soil organic carbon, and total nitrogen were key abiotic drivers shaping earthworm gut bacterial communities. Taken together, this study reveals the succession of bacterial communities and potential functions in earthworm guts within 700 years of upland cultivation, which may provide a broader space for us to rationally exploit and utilize the interactions between soil and earthworm gut microbiotas to benefit the soil nutrient cycling process.

STOP1 activates NRT1.1-mediated nitrate uptake to create a favorable rhizospheric pH for plant adaptation to acidity.

Protons (H+) in acidic soils arrest plant growth. However, the mechanisms by which plants optimize their biological processes to diminish the unfavorable effects of H+ stress remain largely unclear. Here, we showed that in the roots of Arabidopsis thaliana, the C2H2-type transcription factor STOP1 in the nucleus was enriched by low pH in a nitrate-independent manner, with the spatial expression pattern of NITRATE TRANSPORTER 1.1 (NRT1.1) established by low pH required the action of STOP1. Additionally, the nrt1.1 and stop1 mutants, as well as the nrt1.1 stop1 double mutant, had a similar hypersensitive phenotype to low pH, indicating that STOP1 and NRT1.1 function in the same pathway for H+ tolerance. Molecular assays revealed that STOP1 directly bound to the promoter of NRT1.1 to activate its transcription in response to low pH, thus upregulating its nitrate uptake. This action improved the nitrogen use efficiency (NUE) of plants and created a favorable rhizospheric pH for root growth by enhancing H+ depletion in the rhizosphere. Consequently, the constitutive expression of NRT1.1 in stop1 mutants abolished the hypersensitive phenotype to low pH. These results demonstrate that STOP1-NRT1.1 is a key module for plants to optimize NUE and ensure better plant growth in acidic media.

Knockout of FER decreases cadmium concentration in roots of Arabidopsis thaliana by inhibiting the pathway related to iron uptake.

Identifying the genes that affect cadmium (Cd) accumulation in plants is a prerequisite for minimizing dietary Cd uptake from contaminated edible parts of plants by genetic engineering. This study showed that Cd stress inhibited the expression of FERONIA (FER) gene in the roots of wild-type Arabidopsis. Knockout of FER in fer-4 mutants downregulated the Cd-induced expression of several genes related to iron (Fe) uptake, including IRT1, bHLH38, NRAMP1, NRAMP3, FRO2 andFIT. In addition, the Cd concentration in fer-4 mutant roots reduced to approximately half of that in the wild-type seedlings. As a result, the Cd tolerance of fer-4 was higher. Furthermore, increased Fe supplementation had little effect on the Cd tolerance of fer-4 mutants, but clearly improved the Cd tolerance of wild-type seedlings, showing that the alleviation of Cd toxicity by Fe depends on the action of FER. Taken together, the findings demonstrate that the knockout of FER might provide a strategy to reduce Cd contamination and improve the Cd tolerance in plants by regulating the pathways related to Fe uptake.

How can fertilization regimes and durations shape earthworm gut microbiota in a long-term field experiment?

The positive roles of earthworms on soil functionality has been extensively documented. The capacity of the earthworm gut microbiota on decomposition and nutrient cycling under long-term fertilization in field conditions has rarely been studied. Here, we report the structural, taxonomic, and functional responses of Eisenia foetida and Pheretima guillelmi gut microbiota to different fertilization regimes and durations using 16S rRNA gene-based Illumina sequencing and high-throughput quantitative PCR techniques. Our results revealed that the core gut microbiota, especially the fermentative bacteria were mainly sourced from the soil, but strongly stimulated with species-specificity, potential benefits for the host and soil health. The functional compositions of gut microbiota were altered by fertilization with fertilization duration being more influential than fertilization regimes. Moreover, the combination of organic and inorganic fertilization with the longer duration resulted in a higher richness and connectivity in the gut microbiota, and also their functional potential related to carbon (C), nitrogen, and phosphorus cycling, particularly the labile C decomposition, denitrification, and phosphate mobilization. We also found that long-term inorganic fertilization increased the abundance of pathogenic bacteria in the P. guillelmi gut. This study demonstrates that understanding earthworm gut microbiota can provide insights into how agricultural practices can potentially alter soil ecosystem functions through the interactions between soil and earthworm gut microbiotas.

Fates of Antibiotic Resistance Genes in the Gut Microbiome from Different Soil Fauna under Long-Term Fertilization.

Applying organic fertilizers has been well documented to facilitate the dissemination of antibiotic resistance genes (ARGs) in soil ecosystems. However, the role of soil fauna in this process has been seldom addressed, which hampers our ability to predict the fate of and to manage the spread of ARGs. Here, using high-throughput quantitative polymerase chain reaction (HT-qPCR), we examined the effect of long-term (5-, 8-, and 10-year) fertilization treatments (control, inorganic fertilizers, and mixed fertilizers) on the transfer of ARGs between soil, nematodes, and earthworms. We found distinct fates for ARGs in the nematodes and earthworms, with the former having higher enriched levels of ARGs than the latter. Fertilization impacted the number and abundance of ARGs in soil, and fertilization duration altered the composition of ARGs. Shared ARGs among soil, nematodes, and earthworm guts supported by a fast expectation-maximization microbial source tracking analysis demonstrated the trophic transfer potential of ARGs through this short soil food chain. The transfer of ARGs was reduced by fertilization duration, which was mainly ascribed to the reduction of ARGs in the earthworm gut microbiota. This study identified the transfer of ARGs in the soil-nematode-earthworm food chain as a potential mechanism for a wider dissemination of ARGs in the soil ecosystem.

Ammonium aggravates salt stress in plants by entrapping them in a chloride over-accumulation state in an NRT1.1-dependent manner.

Global climate change has exacerbated flooding in coastal areas affected by soil salinization. Ammonium (NH4+) is the predominant form of nitrogen in flooded soils, but the role played by NH4+ in the plant response to salt stress has not been fully clarified. We investigated the responses of Arabidopsis thaliana, Oryza sativa, and Nicotiana benthamiana plants fed with NH4+. All species were hypersensitive to NaCl stress and accumulated more Cl- and less Na+ than those fed with NO3-. Further investigation of A. thaliana indicated that salt hypersensitivity induced by the presence of NH4+ was abolished by removing the Cl- but was not affected by the removal of Na+, suggesting that excess accumulation of Cl- rather than Na+ is involved in NH4+-conferred salt hypersensitivity. The expression of nitrate transporter NRT1.1 protein was also up-regulated by NH4+ treatment, which increased root Cl- uptake due to the Cl- uptake activity of NRT1.1 and the absence of uptake competition from NO3-. Knockout of NRT1.1 in plants decreased their root Cl- uptake and retracted the NH4+-conferred salt hypersensitivity. Our findings revealed that NH4+-aggravated salt stress in plants is associated with Cl- over-accumulation through the up-regulation of NRT1.1-mediated Cl- uptake. These findings suggest the significant impact of Cl- toxicity in flooded coastal areas, an issue of ecological significance.

Partial replacement of inorganic phosphorus (P) by organic manure reshapes phosphate mobilizing bacterial community and promotes P bioavailability in a paddy soil.

The optimization of more sustainable fertilization practice to relieve phosphorus (P) resource scarcity and increase P fertilizer utilization, a better understanding of the regulatory roles of microbes in P mobilization is urgently required to reduce P input. The genes phoD and pqqC are responsible for regulating organic and inorganic P mobilization, respectively. Using high-throughput sequencing, the corresponding bacterial communities harbored by these genes were determined. We conducted a 4-year rice-rice-crop rotation to investigate the responses of phoD- and pqqC-harboring bacterial communities to the partial replacement of inorganic P fertilizer by organic manure with reduced P input. The results showed that a combination of organic and inorganic fertilization maintained high rice yield, and also produced a more complex and stable phosphate mobilizing bacterial community, which contributed to phosphatase activities more than their gene abundances in the model analysis. Compared with the conventional mineral fertilization, organic-inorganic fertilization with the reduced P input slightly increased pqqC gene abundance while significantly enhanced the abundance of phoD-harboring bacteria, especially the genera Bradyrhizobium and Methylobacterium known as potential organic P mineralizers which can maintain high rice production. Moreover, the increased pH was the most impactful factor for the phoD- and pqqC-harboring bacterial communities, by promoting microbial P turnover and greatly increasing bioavailable P pools (H2O-Pi and NaHCO3-Pi, NaOH-Pi) in this P-deficient paddy soil. Hence, our study demonstrated that the partial replacement of mineral P with organic manure could reshape the inorganic phosphate solubilizing and alkaline-phosphomonoesterase encoding bacterial communities towards more resilient and effective to the high P utilization and productivity over intense cultivation, providing insights into the potential of soil microbes in the efficient management of agricultural P fertilization.

Nitrate transporter 1.1 alleviates lead toxicity in Arabidopsis by preventing rhizosphere acidification.

Identification of the mechanisms that control lead (Pb) concentration in plants is a prerequisite for minimizing dietary uptake of Pb from contaminated crops. This study examines how nitrate uptake by roots affects Pb uptake and reveals a new resistance strategy for plants to cope with Pb contamination. We investigated the interaction between nitrate transporter (NRT)-mediated NO3- uptake and exposure to Pb in Arabidopsis using NRT-related mutants. Exposure to Pb specifically stimulated NRT1.1-mediated nitrate uptake. Loss of function of NRT1.1 in nrt1.1-knockout mutants resulted in greater Pb toxicity and higher Pb accumulation in nitrate-sufficient growth medium, whereas no difference was seen between wild-type plants and null-mutants for NRT1.2, NRT2.1, NRT2.2, NRT2.4, and NRT2.5. These results indicate that only NRT1.1-mediated NO3- uptake alleviated Pb toxicity in the plants. Further examination indicated that rhizosphere acidification, which favors Pb entry to roots by increasing its availability, is prevented when NRT1.1 is functional and both NO3- and NH4+ are present in the medium.

Increased risk of phosphorus and metal leaching from paddy soils after excessive manure application: Insights from a mesocosm study.

Livestock manure has gradually become an alternative fertilizer for maintaining soil fertility, whereas excessive application of manure leads to the release of phosphorus (P) and toxic metals that may cause complex environmental risks. To investigate the accumulation and migration of P within soil profiles, a mesocosm experiment was conducted to analyze the content and leaching of soil P, metals, and dissolved organic carbon after different fertilization treatments, including control (no fertilizer, CK), chemical fertilizer (CF), chemical fertilizer combined low (CF + LPM) and high (CF + HPM) rate of manure application. Results showed that a high rate of manure application significantly enhanced the accumulation of total soil P (by ~14%) and P availability (easily-available P, by ~24%; Olsen-P, by ~20%) in topsoil, and also increased the content of easily-available organic P (EA-Po) in both topsoil and subsoil compared to the CK treatment. The migration of dissolved inorganic and organic P (DIP and DOP) in leachate within soil profiles was strengthened by manure application. Moreover, significant positive correlations between P, metals, and dissolved organic carbon (DOC) in leachate indicated that downward co-migration occurred within the soil profiles, and also suggested that excessive manure application can intensify the risk of P loss by increasing the migration of manure-derived DOC. Overall, our findings provide insights into P accumulation and migration within soil profiles after excessive manure application, which is useful for predicting the potential risk of P and metal leaching from paddy soils.

Sulfide alleviates cadmium toxicity in Arabidopsis plants by altering the chemical form and the subcellular distribution of cadmium.

Several sulfur compounds are thought to play important roles in the plant tolerance to cadmium (Cd), but the role of inorganic sulfide in Cd tolerance remains largely unknown. In this study, we found that Cd exposure increased the accumulation of soluble sulfide in Arabidopsis plants. When exogenous sulfide, in the form of NaHS, was foliarly applied, Cd-induced growth inhibition and oxidative stress were alleviated. In addition, although the foliar application of sulfide did not affect the total Cd levels, it significantly decreased the soluble Cd fractions in plants. Furthermore, foliar applications of sulfide decreased Cd distribution in the cytoplasm and organelles, but increased Cd retention in the cell wall, which is a less sensitive compartment. These results suggest that the Cd-induced accumulation of soluble sulfide alleviates Cd toxicity in plants by inactivating Cd and sequestering it into the cell wall.

Effects of combined application of nitrogen fertilizer and biochar on the nitrification and ammonia oxidizers in an intensive vegetable soil.

Soil amended with single biochar or nitrogen (N) fertilizer has frequently been reported to alter soil nitrification process due to its impact on soil properties. However, little is known about the dynamic response of nitrification and ammonia-oxidizers to the combined application of biochar and N fertilizer in intensive vegetable soil. In this study, an incubation experiment was designed to evaluate the effects of biochar and N fertilizer application on soil nitrification, abundance and community shifts of ammonia-oxidizing bacteria (AOB) and ammonia oxidizing archaea (AOA) in Hangzhou greenhouse vegetable soil. Results showed that single application of biochar had no significant effect on soil net nitrification rates and ammonia-oxidizers. Conversely, the application of only N fertilizer and N fertilizer + biochar significantly increased net nitrification rate and the abundance of AOB rather than AOA, and only AOB abundance was significantly correlated with soil net nitrification rate. Moreover, the combined application of N fertilizer and biochar had greater effect on AOB communities than that of the only N fertilizers, and the relative abundance of 156 bp T-RF (Nitrosospira cluster 3c) decreased but 60 bp T-RF (Nitrosospira cluster 3a and cluster 0) increased to become a single predominant group. Phylogenetic analysis indicated that all the AOB sequences were grouped into Nitrosospira cluster, and most of AOA sequences were clustered within group 1.1b. We concluded that soil nitrification was stimulated by the combined application of N fertilizer and biochar via enhancing the abundance and shifting the community composition of AOB rather than AOA in intensive vegetable soil.

Alleviation of proton toxicity by nitrate uptake specifically depends on nitrate transporter 1.1 in Arabidopsis.

Protons in acid soil are highly rhizotoxic to plants, but the mechanism of tolerance of plants to protons is largely unknown. Nitrate uptake by root cells is accompanied by the uptake of protons. Therefore, nitrate uptake transporters (NRTs) may be involved in plant tolerance to proton toxicity. We investigated the root nitrate uptake response to proton stress in Arabidopsis and its association with proton tolerance using NRT-related mutants and pharmacological methods. Lack of NRT1.1 in knockout nrt1.1 mutants led to impaired proton tolerance in nitrate-sufficient growth medium, whereas no difference was seen between wild-type plants and NRT1.2-, NRT2.1-, NRT2.2-, and NRT2.4-null mutants. Another nrt1.1 point mutant, which is defective in nitrate uptake but has a normal nitrate-sensing function, also had impaired proton tolerance compared with the wild-type plant. Furthermore, proton stress induced NRT1.1-mediated nitrate uptake. These results indicate that NRT1.1-conferred proton tolerance depends on nitrate uptake activity. In addition, the rooting medium was alkalified by wild-type plants, but not by knockout nrt1.1 mutants, and in pH-buffered medium, there were no differences in proton tolerance between wild-type plants and knockout nrt1.1 mutants. We conclude that NRT1.1-mediated nitrate uptake plays a crucial role in plant proton tolerance by alkalifying the rhizosphere.

Increased Sucrose Accumulation Regulates Iron-Deficiency Responses by Promoting Auxin Signaling in Arabidopsis Plants.

Previous studies have identified that auxins acts upstream of nitric oxide in regulating iron deficiency responses in roots, but the upstream signaling molecule of auxins remains unknown. In this study, we showed that Fe deficiency increased sucrose (Suc) level in roots of Arabidopsis (Arabidopsis thaliana). Exogenous application of Suc further stimulated Fe deficiency-induced ferric-chelate-reductase (FCR) activity and expression of Fe acquisition-related genes FRO2, IRT1, and FIT in roots. The opposite patterns were observed in the dark treatment. In addition, FCR activity and expression of Fe acquisition-related genes were higher in the Suc high-accumulating transgenic plant 35S::SUC2 but were lower in the Suc low-accumulating mutant suc2-5 compared with wild-type plants under Fe-deficient conditions. Consequently, Fe deficiency tolerance was enhanced in 35S::SUC2 but was compromised in suc2-5. Exogenous Suc also increased root ß-glucuronidase (GUS) activity in auxin-inducible reporter DR5-GUS transgenic plants under Fe deficiency. However, exogenous Suc failed to increase FCR activity and expression of Fe acquisition-related genes in the auxin transport-impaired mutants aux1-7 and pin1-1 as well as in the wild-type plants treated with an auxin transport inhibitor under Fe deficiency. In summary, we found that increased Suc accumulation is required for regulating Fe deficiency responses in plants, with auxins acting downstream in transmitting the Fe deficiency signal.

Elevation of NO production increases Fe immobilization in the Fe-deficiency roots apoplast by decreasing pectin methylation of cell wall.

Cell wall is the major component of root apoplast which is the main reservoir for iron in roots, while nitric oxide (NO) is involved in regulating the synthesis of cell wall. However, whether such regulation could influence the reutilization of iron stored in root apoplast remains unclear. In this study, we observed that iron deficiency elevated NO level in tomato (Solanum lycopersicum) roots. However, application of S-nitrosoglutathione, a NO donor, significantly enhanced iron retention in root apoplast of iron-deficient plants, accompanied with a decrease of iron level in xylem sap. Consequently, S-nitrosoglutathione treatment increased iron concentration in roots, but decreased it in shoots. The opposite was true for the NO scavenging treatment with 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). Interestingly, S-nitrosoglutathione treatment increased pectin methylesterase activity and decreased degree of pectin methylation in root cell wall of both iron-deficient and iron-sufficient plants, which led to an increased iron retention in pectin fraction, thus increasing the binding capacity of iron to the extracted cell wall. Altogether, these results suggested that iron-deficiency-induced elevation of NO increases iron immobilization in root apoplast by decreasing pectin methylation in cell wall.

Oxalate synthesis in leaves is associated with root uptake of nitrate and its assimilation in spinach (Spinacia oleracea L.) plants.

BACKGROUND: Excessive accumulation of oxalate in numerous vegetables adversely affects their quality as food. While it is known that nitrate could effectively stimulate oxalate accumulation in many vegetables, little information is available about the mechanism of nitrate-induced oxalate accumulation. In this study, we examined the association of oxalate synthesis with nitrate uptake and assimilation in two genotypes of spinach (Spinacia oleracea L.), Heizhenzhu and Weilv. RESULTS: Increasing nitrate levels resulted in enhanced synthesis of oxalate, as well as increased root uptake of nitrate and leaf activities of nitrate reductase (NR) and glutamine synthetase (GS) for both genotypes. Correlation analysis revealed that oxalate accumulation in spinach leaves was positively related with rate of nitrate uptake by roots, as well as leaf activities of NR and GS. Addition of plasmalemma H(+)-ATPase inhibitor sodium vanadate (Na3VO4) significantly decreased leaf oxalate accumulation in both genotypes. Presence of NR or GS inhibitors led to reduction of leaf oxalate contents, GS/NR activities and decreased nitrate uptake rate. Significantly higher levels of nitrate root uptake, leaf NR and GS activities were observed in the high-oxalate genotype Heizhenzhu than in Weilv. CONCLUSION: Oxalate synthesis in leaves of spinach is not only positively associated with root uptake of nitrate, but also with its assimilation within the plants.

Inhibition of nitrate transporter 1.1-controlled nitrate uptake reduces cadmium uptake in Arabidopsis.

Identification of mechanisms that decrease cadmium accumulation in plants is a prerequisite for minimizing dietary uptake of cadmium from contaminated crops. Here, we show that cadmium inhibits nitrate transporter 1.1 (NRT1.1)-mediated nitrate (NO3 (-)) uptake in Arabidopsis (Arabidopsis thaliana) and impairs NO3 (-) homeostasis in roots. In NO3 (-)-containing medium, loss of NRT1.1 function in nrt1.1 mutants leads to decreased levels of cadmium and several other metals in both roots and shoots and results in better biomass production in the presence of cadmium, whereas in NO3 (-)-free medium, no difference is seen between nrt1.1 mutants and wild-type plants. These results suggest that inhibition of NRT1.1 activity reduces cadmium uptake, thus enhancing cadmium tolerance in an NO3 (-) uptake-dependent manner. Furthermore, using a treatment rotation system allowing synchronous uptake of NO3 (-) and nutrient cations and asynchronous uptake of cadmium, the nrt1.1 mutants had similar cadmium levels to wild-type plants but lower levels of nutrient metals, whereas the opposite effect was seen using treatment rotation allowing synchronous uptake of NO3 (-) and cadmium and asynchronous uptake of nutrient cations. We conclude that, although inhibition of NRT1.1-mediated NO3 (-) uptake by cadmium might have negative effects on nitrogen nutrition in plants, it has a positive effect on cadmium detoxification by reducing cadmium entry into roots. NRT1.1 may regulate the uptake of cadmium and other cations by a common mechanism.

Short-term alteration of nitrogen supply prior to harvest affects quality in hydroponic-cultivated spinach (Spinacia oleracea).

BACKGROUND: Quality-associated problems, such as excessive in planta accumulation of oxalate, often arise in soillessly cultivated spinach (Spinacia oleracea). Maintaining a higher level of ammonium (NH4⁺) compared to nitrate (NO3⁻) during the growth period can effectively decrease the oxalate content in hydroponically cultivated vegetables. However, long-term exposure to high concentrations of NH4⁺ induces toxicity in plants, and thus decreases the biomass production. Short-term application of NH4⁺ before harvesting in soilless cultivation may provide an alternative strategy to decrease oxalate accumulation in spinach, and minimise the yield reduction caused by NH4⁺ toxicity. RESULT: The plants were pre-cultured in 8 mmol L⁻¹ NO3⁻ nutrient solution. Next, 6 days before harvest, the plants were transferred to a nutrient solution containing 4 mmol L⁻¹ NO3⁻ and 4 mmol L⁻¹ NH4⁺. This new mix clearly reduced oxalate accumulation, increased levels of several antioxidant compounds, and enhanced antioxidant capacity in the edible parts of spinach plants, but it did not affect biomass production. However, when the 8 mmol L⁻¹ NO3⁻ was shifted to either nitrogen-free, 4 mmol L⁻¹ NH4⁺ or 8 mmol L⁻¹ NH4⁺ treatments, although some of the quality indexes were improved, yields were significantly reduced. CONCLUSIONS: Short-term alteration of nitrogen supply prior to harvest significantly affects quality and biomass of spinach plants, and we strongly recommend to simultaneously use NO3⁻ and NH4⁺ in hydroponic cultivation, which improves vegetable quality without decreasing biomass production.