Impact of nitrogen fertilizer type and application rate on growth, nitrate accumulation, and postharvest quality of spinach.

Background: A balanced supply of nitrogen is essential for spinach, supporting both optimal growth and appropriate nitrate (NO3 -) levels for improved storage quality. Thus, choosing the correct nitrogen fertilizer type and application rate is key for successful spinach cultivation. This study investigated the effects of different nitrogen (N) fertilizer type and application rates on the growth, nitrate content, and storage quality of spinach plants. Methods: Four fertilizer types were applied at five N doses (25, 50, 200, and 400 mg N kg-1) to plants grown in plastic pots at a greenhouse. The fertilizer types used in the experiment were ammonium sulphate (AS), slow-release ammonium sulphate (SRAS), calcium nitrate (CN), and yeast residue (YR). Spinach parameters like Soil Plant Analysis Development (SPAD) values (chlorophyll content), plant height, and fresh weight were measured. Nitrate content in leaves was analyzed after storage periods simulating post-harvest handling (0, 5, and 10 days). Results: The application of nitrogen fertilizer significantly influenced spinach growth parameters and nitrate content. The YRx400 treatment yielded the largest leaves (10.3 ± 0.5 cm long, 5.3 ± 0.2 cm wide). SPAD values increased with higher N doses for AS, SRAS, and CN fertilizers, with AS×400 (58.1 ± 0.8) and SRAS×400 (62.0 ± 5.8) reaching the highest values. YR treatments showed a moderate SPAD increase. Fresh weight response depended on fertilizer type, N dose, and storage period. While fresh weight increased in all fertilizers till 200 mg kg-1 dose, a decrease was observed at the highest dose for AS and CN. SRAS exhibited a more gradual increase in fresh weight with increasing nitrogen dose, without the negative impact seen at the highest dose in AS and CN. Nitrate content in spinach leaves varied by fertilizer type, dose, and storage day. CNx400 resulted in the highest NO3 - content (4,395 mg kg-1) at harvest (Day 0), exceeding the European Union's safety limit. This level decreased over 10 days of storage but remained above the limit for CN on Days 0 and 5. SRAS and YR fertilizers generally had lower NO3 - concentrations throughout the experiment. Storage at +4 °C significantly affected NO3 - content. While levels remained relatively stable during the first 5 days, a substantial decrease was observed by Day 10 for all fertilizers and doses, providing insights into the spinach's nitrate content over a 10-day storage period. Conclusion: For rapid early growth and potentially higher yields, AS may be suitable at moderate doses (200 mg kg-1). SRAS offers a more balanced approach, promoting sustained growth while potentially reducing NO3 - accumulation compared to AS. Yeast residue, with its slow nitrogen release and consistently low NO3 - levels, could be a viable option for organic spinach production.

Salt stress amelioration and nutrient strengthening in spinach (Spinacia oleracea L.) via biochar amendment and zinc fortification: seed priming versus foliar application.

Soil salinity is a major nutritional challenge with poor agriculture production characterized by high sodium (Na+) ions in the soil. Zinc oxide nanoparticles (ZnO NPs) and biochar have received attention as a sustainable strategy to reduce biotic and abiotic stress. However, there is a lack of information regarding the incorporation of ZnO NPs with biochar to ameliorate the salinity stress (0, 50,100 mM). Therefore, the current study aimed to investigate the potentials of ZnO NPs application (priming and foliar) alone and with a combination of biochar on the growth and nutrient availability of spinach plants under salinity stress. Results demonstrated that salinity stress at a higher rate (100 mM) showed maximum growth retardation by inducing oxidative stress, resulted in reduced photosynthetic rate and nutrient availability. ZnO NPs (priming and foliar) alone enhanced growth, chlorophyll contents and gas exchange parameters by improving the antioxidant enzymes activity of spinach under salinity stress. While, a significant and more pronounced effect was observed at combined treatments of ZnO NPs with biochar amendment. More importantly, ZnO NPs foliar application with biochar significantly reduced the Na+ contents in root 57.69%, and leaves 61.27% of spinach as compared to the respective control. Furthermore, higher nutrient contents were also found at the combined treatment of ZnO NPs foliar application with biochar. Overall, ZnO NPs combined application with biochar proved to be an efficient and sustainable strategy to alleviate salinity stress and improve crop nutritional quality under salinity stress. We inferred that ZnO NPs foliar application with a combination of biochar is more effectual in improving crop nutritional status and salinity mitigation than priming treatments with a combination of biochar.

Gene Regulatory Network Controlling Flower Development in Spinach (Spinacia oleracea L.).

Spinach (Spinacia oleracea L.) is a dioecious, diploid, wind-pollinated crop cultivated worldwide. Sex determination plays an important role in spinach breeding. Hence, this study aimed to understand the differences in sexual differentiation and floral organ development of dioecious flowers, as well as the differences in the regulatory mechanisms of floral organ development of dioecious and monoecious flowers. We compared transcriptional-level differences between different genders and identified differentially expressed genes (DEGs) related to spinach floral development, as well as sex-biased genes to investigate the flower development mechanisms in spinach. In this study, 9189 DEGs were identified among the different genders. DEG analysis showed the participation of four main transcription factor families, MIKC_MADS, MYB, NAC, and bHLH, in spinach flower development. In our key findings, abscisic acid (ABA) and gibberellic acid (GA) signal transduction pathways play major roles in male flower development, while auxin regulates both male and female flower development. By constructing a gene regulatory network (GRN) for floral organ development, core transcription factors (TFs) controlling organ initiation and growth were discovered. This analysis of the development of female, male, and monoecious flowers in spinach provides new insights into the molecular mechanisms of floral organ development and sexual differentiation in dioecious and monoecious plants in spinach.

Utilization of organic-residues as potting media: Physico-chemical characteristics and their influence on vegetable production.

Soilless agriculture is acknowledged worldwide because it uses organic leftovers as a means of supporting intensive and efficient plant production. However, the quality of potting media deteriorates because of lower nutrient content and excessive shrinkage of most organic materials. A current study was undertaken to identify the optimal blend of locally available organic materials with desirable qualities for use as potting media. Therefore, different ingredients, viz., Pinus roxburghii needles, sugarcane bagasse, and farmyard manure were used alone or in combination as potting media to test their suitability by growing spinach as a test crop. Results showed that an increase in Pinus roxburghii needles and sugarcane bagasse decreased medium pH and electrical conductivity. Higher pH and electrical conductivity were recorded for the treatments having a higher farmyard manure ratio (≥50%) in combination. Except for pine needles 100%, pH and electrical conductivity were in the recommended range. The growth attributes include, leaves plant-1, shoot length, fresh- and dry shoot weight along with plant macronutrients (nitrogen, phosphorous, and potassium) and micronutrients (iron, copper, manganese, and zinc) content were higher in treatment pine needles 50%+farmyard manure 50% followed by pine needles 25%+farmyard manure 50%+sugarcane bagasse 25%. Moreover, the particular treatment of pine needles 50%+farmyard manure 50% exhibited the highest concentrations of macro- (nitrogen, phosphorus, and potassium) as well as micronutrients (iron, copper, manganese, and zinc) in the potting media following the harvest. This study highlights the potential of utilizing agro-industrial litter/waste as a soilless growing medium for spinach production under greenhouse conditions. When employed in appropriate proportions, this approach not only addresses disposal concerns but also proves effective for sustainable cultivation. Further research is needed to investigate the use of these wastes as potting media by mixing various particle-size ingredients.

The mechanism behind tenuazonic acid-mediated inhibition of plant plasma membrane H+-ATPase and plant growth.

The increasing prevalence of herbicide-resistant weeds has led to a search for new herbicides that target plant growth processes differing from those targeted by current herbicides. In recent years, some studies have explored the use of natural compounds from microorganisms as potential new herbicides. We previously demonstrated that tenuazonic acid (TeA) from the phytopathogenic fungus Stemphylium loti inhibits the plant plasma membrane (PM) H+-ATPase, representing a new target for herbicides. In this study, we further investigated the mechanism by which TeA inhibits PM H+-ATPase and the effect of the toxin on plant growth using Arabidopsis thaliana. We also studied the biochemical effects of TeA on the PM H+-ATPases from spinach (Spinacia oleracea) and A. thaliana (AHA2) by examining PM H+-ATPase activity under different conditions and in different mutants. Treatment with 200 µM TeA-induced cell necrosis in larger plants and treatment with 10 µM TeA almost completely inhibited cell elongation and root growth in seedlings. We show that the isoleucine backbone of TeA is essential for inhibiting the ATPase activity of the PM H+-ATPase. Additionally, this inhibition depends on the C-terminal domain of AHA2, and TeA binding to PM H+-ATPase requires the Regulatory Region I of the C-terminal domain in AHA2. TeA likely has a higher binding affinity toward PM H+-ATPase than the phytotoxin fusicoccin. Finally, our findings show that TeA retains the H+-ATPase in an inhibited state, suggesting that it could act as a lead compound for creating new herbicides targeting the PM H+-ATPase.

Effect of zinc nanoparticles seed priming and foliar application on the growth and physio-biochemical indices of spinach (Spinacia oleracea L.) under salt stress.

Salt stress is the major risk to the seed germination and plant growth via affecting physiological and biochemical activities in plants. Zinc nanoparticles (ZnNPs) are emerged as a key agent in regulating the tolerance mechanism in plants under environmental stresses. However, the tolerance mechanisms which are regulated by ZnNPs in plants are still not fully understood. Therefore, the observation was planned to explore the role of ZnNPs (applied as priming and foliar) in reducing the harmful influence of sodium chloride (NaCl) stress on the development of spinach (Spinacia oleracea L.) plants. Varying concentrations of ZnNPs (0.1%, 0.2% & 0.3%) were employed to the spinach as seed priming and foliar, under control as well as salt stress environment. The alleviation of stress was observed in ZnNPs-applied spinach plants grown under salt stress, with a reduced rise in the concentration hydrogen peroxide, melondialdehyde and anthocyanin contents. A clear decline in soluble proteins, chlorophyll contents, ascorbic acid, sugars, and total phenolic contents was observed in stressed conditions. Exogenous ZnNPs suppressed the NaCl generated reduction in biochemical traits, and progress of spinach plants. However, ZnNPs spray at 0.3% followed by priming was the most prominent treatment in the accumulation of osmolytes and the production of antioxidant molecules in plants.

High and low oxalate content in spinach: an investigation of accumulation patterns.

BACKGROUND: Oxalic acid is a common antinutrient in the human diet, found in large quantities in spinach. However, spinach is highly regarded by vegetable producers because of its nutritional content and economic value. One of the primary purposes of spinach-breeding programs is to improve the nutritional value of spinach by adjusting oxalate accumulation. Knowledge of the biosynthetic patterns of oxalic acid, and its different forms, is important for a better understanding of this process. RESULTS: We found three biosynthetic patterns of accumulation and concentration of oxalates. Two of them are related to the maximum type and one is related to the minimum type. We also developed a general model of variations in these compounds in the genotypes that were studied. CONCLUSION: This study introduced a unique type of spinach with high oxalate accumulation, which could be particularly suitable for consumption. This had the highest ratio of insoluble oxalate to soluble oxalate. It also accumulated more ascorbic acid (AA) than other types. Our findings in this study also indicate a small role for AA as a precursor to oxalate production in spinach, possibly confirming the significant role of glyoxylate as the most critical precursor in this plant. © 2021 Society of Chemical Industry.

Lysinibacillus xylanilyticus Strain GIC41 as a Potential Plant Biostimulant.

To identify Lysinibacillus strains with the potential to function as plant biostimulants, we screened 10 previously isolated Lysinibacillus strains from the rhizosphere and soil for their plant growth-promoting (PGP) effects. In vitro tests showed that all strains produced indole-3-acetic acid. In primary screening, the PGP effects of these strains were assessed on spinach seedlings grown on Jiffy-7 pellets; strains GIC31, GIC41, and GIC51 markedly promoted shoot growth. In secondary screening, the PGP efficacies of these three strains were examined using spinach seedlings grown in pots under controlled conditions. Only GIC41 exerted consistent and significant PGP effects; therefore, it was selected for subsequent experiments. The results of 6-week glasshouse experiments revealed that GIC41 markedly increased shoot dry weight by ca. 12-49% over that of the control. The impact of fertilization levels on the PGP efficacy of GIC41 was investigated using pot experiments. The application of a specific level of fertilizer was required for the induction of sufficient PGP effects by this strain. The phylogenetic ana-lysis based on the 16S rDNA sequence identified GIC41 as L. xylanilyticus. Collectively, these results show the potential of strain GIC41 to function as a plant biostimulant.

The first report of iron-rich population of adapted medicinal spinach (Blitum virgatum L.) compared with cultivated spinach (Spinacia oleracea L.).

Folk medicine such as herbal and natural products have been used for centuries in every culture throughout the world. The Chenopodiaceae family with more than 1500 species is dispersed worldwide. The Iranian wild spinach (Blitum virgatum L.) is an important traditional medicinal plant used for antiviral diseases such as pneumonia and other respiratory track infections. This plant is a mountainous herb and is growing upper than 3000 m. We performed a mass selection plant breeding program on wild populations of this Iranian wild spinach during 2013-2020. Based on experimental and field characteristics this plant was identified as B. virgatum, |abbaricum|, and related characteristics were prepared with reference to the International Union for the Protection of New Varieties of Plants (UPOV). Mass selection program resulted from an adapted population named as medicinal spinach (MSP) population. To compare the mineral content of the mass-selected population with cultivated spinach (Spinacia oleracea L. |Varamin 88|), both plants were planted in pots and fields under similar conditions. In five leaves stage, plant samples were taken from both leaf and crown sections and used for experimental analysis. Atomic absorption spectroscopy was used to determine the mineral content including iron (Fe), zinc (Z), manganese (Mn), and copper (Cu). Our results showed the selected medicinal spinach population (MSP) with about 509 ppm iron was an important iron-rich population with about 3.5-4 times more than the amount of iron in cultivated spinach in the same conditions. Because iron is an important essential element for blood production, respiration process, energy metabolisms, synthesis of collagen, and some neurotransmitters are needed for proper immune function, so the supply of absorbable adequate iron is very important. The reasons such as the prevalence of the COVID-19 pandemic, which affects the amount of exchangeable oxygen in the lungs and historical local evidences of the use of this plant (MSP) for pneumonia, could open new horizons for focusing on studies related to the use of ancestral human experiences in addition to scientifically modern research.

Auxin regulated metabolic changes underlying sepal retention and development after pollination in spinach.

BACKGROUND: Pollination accelerate sepal development that enhances plant fitness by protecting seeds in female spinach. This response requires pollination signals that result in the remodeling within the sepal cells for retention and development, but the regulatory mechanism for this response is still unclear. To investigate the early pollination-induced metabolic changes in sepal, we utilize the high-throughput RNA-seq approach. RESULTS: Spinach variety 'Cornel 9' was used for differentially expressed gene analysis followed by experiments of auxin analog and auxin inhibitor treatments. We first compared the candidate transcripts expressed differentially at different time points (12H, 48H, and 96H) after pollination and detected significant difference in Trp-dependent auxin biosynthesis and auxin modulation and transduction process. Furthermore, several auxin regulatory pathways i.e. cell division, cell wall expansion, and biogenesis were activated from pollination to early developmental symptoms in sepals following pollination. To further confirm the role auxin genes play in the sepal development, auxin analog (2, 4-D; IAA) and auxin transport inhibitor (NPA) with different concentrations gradient were sprayed to the spinach unpollinated and pollinated flowers, respectively. NPA treatment resulted in auxin transport weakening that led to inhibition of sepal development at concentration 0.1 and 1 mM after pollination. 2, 4-D and IAA treatment to unpollinated flowers resulted in sepal development at lower concentration but wilting at higher concentration. CONCLUSION: We hypothesized that sepal retention and development might have associated with auxin homeostasis that regulates the sepal size by modulating associated pathways. These findings advanced the understanding of this unusual phenomenon of sepal growth instead of abscission after pollination in spinach.

Insight into morphological and molecular variations across Iranian spinach landraces.

Spinach is salt-tolerant leafy vegetable stemmed from central Asia. It has been well adapted to different climates of Iran. We aimed to study genetic diversity between several landraces as a prerequisite for crop improvement programs using molecular markers including microsatellites (SSRs) and morphological traits. Genetic diversity was studied among 22 spinach landraces using morphological and molecular tools. We developed 17 genic and genomic SSR markers based on the information acquired from NCBI resources. Morphological evaluation indicated high variability for economic traits including leaf color, leaf thickness, leaf wrinkle and seed type, across Iranian landraces. The molecular results exhibited that 12 out of 17 primer pairs successfully amplified genomic DNA with explicit bands. The results verified that genic markers were superior to genomic markers to detect polymorphism and genetic diversity. In this regard, PIC for genomic and genic SSRs was in the range of 0.4616-0.6621 and 0.5188-0.7394, respectively. Polymorphic genic SSRs were identified to be directly and indirectly involved in biotic/abiotic stresses. High degree of polymorphism, which was detected across the landraces by genic SSRs, could assist us to select 11 landraces for the second experiment. The second experiment was designed to evaluate the response of selected landraces to salinity stress. The results confirmed genetic variability among the landraces in terms of salinity tolerance. A highly diverse germplasm of Iranian spinach based on molecular and morphological characteristics along with the tolerance to oxidative stress provides an ample opportunity for plant breeders to select superior genotypes.

Prospective usage of magnesium potassium phosphate cement combined with Bougainvillea alba derived biochar to reduce Pb bioavailability in soil and its uptake by Spinacia oleracea L.

Combining biochar (BR) with other immobilizing amendments has additive effects on Pb immobilization and been recognized to be effective for the restoration of Pb polluted soils. However, the impacts of different proportions between BR and a highly efficient Pb immobilizing agent called "magnesium potassium phosphate cement (MC)" have never been earlier investigated. This work aimed to investigate the consequences of BR and MC alone and their mixtures of 25:75, 50:50, and 75:25 ratios on Pb bioavailability, Pb immobilization index (Pb-IMMi), and enzymatic activities in Pb polluted soil. Furthermore, amendments effects on Pb distribution in spinach, growth, antioxidant capacity, biochemical, and nutritional spectrum were also investigated. We found that MC alone performed well to immobilize Pb in soil and reducing its distribution in shoots, but was less efficient to improve soil enzymatic activities and plant attributes. Conversely, the application of BR alone stimulated soil enzymatic activities, plant growth, and quality but was less effective to immobilize Pb in soil and reducing shoot Pb concentrations. The combinations of BR and MC of various ratios showed variable results. Interestingly, the most promising outcomes were obtained with BR50%+MC50% treatment which resulted in enhanced Pb-IMMi (73%), activities of soil enzymes, plant growth and quality, and antioxidant capacity, compared to control. Likewise, significant reductions in Pb concentrations in shoots (85%), roots (78%), extractable Pb (73%) were also obtained with BR50%+MC50% treatment, compared to control. Such outcomes point towards a cost-effective approach for reducing Pb uptake by the plants via using MC and BR at a 50:50 ratio.

Fine mapping and molecular marker development of the Fs gene controlling fruit spines in spinach (Spinacia oleracea L.).

KEY MESSAGE: The Fs gene, which controls spinach fruit spines, was fine mapped to a 0.27 Mb interval encompassing four genes on chromosome 3. There are two types of fruit of spinach (Spinacia oleracea L.), spiny and spineless, which are visually distinguishable by the spines of fruit coat. In spinach breeding, the fruit characteristic is an important agronomic trait that have impacts on "seed" treatment and mechanized sowing. However, the gene(s) controlling the fruit spiny trait have not been characterized and the genetic mechanism of this trait remained unclear. The objectives of the study were to fine map the gene controlling fruit spines and develop molecular markers for marker-assisted selection purpose. Genetic analysis of the spiny trait in segregating populations indicated that fruit spines were controlled by a single dominant gene, designated as Fs. Using a super-BSA method and recombinants analysis in a BC1 population, Fs was mapped to a 1.9-Mb interval on chromosome 3. The Fs gene was further mapped to a 0.27-Mb interval using a recombinant inbred line (RIL) population with 120 lines. From this 0.27 Mb region, four candidate genes were identified in the reference genome. The structure and expression of the four genes were compared between the spiny and spineless parents. A co-dominant marker YC-15 was found to be co-segregating with the fruit spines trait, which produced a 129-bp fragment specific to spiny trait and a 108-bp fragment for spineless fruit. This marker can predict spiny trait with a 94.8% accuracy rate when tested with 100 diverse germplasm, suggesting that this marker would be valuable for marker-assisted selection in spinach breeding.

Effect of salt stress on the growth, mineral contents, and metabolite profiles of spinach.

BACKGROUND: Increased soil salt concentration decreases productivity and changes the physiological and chemical properties of plants. Various omics technologies have been used to understand the salt response in plants but overall changes in the metabolite profiles of spinach (Spinacia oleracea L.) under salt stress have not been studied. In this article, therefore, the changes in mineral and metabolite profiles of spinach plants cultivated with different NaCl concentrations of 0-200 mmol L-1 in the irrigation water were analyzed to investigate the effect of salt stress on nutritional quality. RESULTS: Increasing NaCl concentration decreased plant growth due to mineral imbalance. The amounts of minerals (K+ , Ca2+ , and Fe2+ ) were reduced with increasing NaCl concentration, resulting in altered ratios of Na+ :K+ and Na+ :Ca2+ . The change in the mineral ratios due to NaCl irrigation led to a decrease in the height and an increase in the weight of spinach. Moreover, the profiles of 32 metabolites, including flavonoids, amino acids, acidic compounds, sugars, and lipid-related compounds, were altered by NaCl irrigation; most of them showed decreased levels. In particular, at 200 mmol L-1 NaCl, the levels of sucrose, glutamic acid, hexose sugars, and acidic compounds significantly decreased upon NaCl irrigation. Based on these metabolites, a salt-stress-related spinach metabolomic pathway was proposed. CONCLUSION: Sodium chloride irrigation increased mineral imbalance, resulting in decreased plant growth, and the levels of most metabolites involved in energy production, sensory quality, and health benefits decreased with NaCl irrigation. The results suggest that NaCl irrigation negatively affects the nutritional quality of spinach. © 2020 Society of Chemical Industry.

Binary mixture of nanoparticles in sewage sludge: Impact on spinach growth.

Present study carried out pot experiments and evaluated effects of single and binary mixture of nanoparticles (exposed via sludge as soil conditioner) on spinach plant. Exposure of Ag2O nanoparticles (NPs) (1 and 10 mg/kg soil-sludge) did not show significant reduction in plant as compared to control. On the other hand, TiO2 NPs (exposed as single and in binary mixture) resulted in significant increase in root length (29% and 37%) and fresh weight (60% and 48%) at highest exposure concentration. Total chlorophyll content decreased for Ag2O and binary mixture (7% and 4%, respectively) and increased for TiO2 (5%) at 10 mg/kg soil-sludge. The toxic interaction between Ag2O and TiO2 NPs was additive at both exposure concentrations. Ag2O NPs had higher tendency of root surface adsorption than TiO2 NPs. Metal content in spinach leaves at highest exposure concentration was Ag: 2.6 ± 0.55 mg/g plant biomass(for Ag2O NPs) and 1.02 ± 0.32 mg/g plant biomass (for Ag2O + TiO2 NPs) and for Ti: 1.12 ± 0.78 (for TiO2 NPs) mg/g plant biomass and 0.58 ± 0.41 mg/g (for Ag2O + TiO2 NPs). The inadvertent ingestion of NPs- contaminated spinach resulted in projected daily intake (DI) of Ag and Ti for different age-mass classes (child to adult) exceeding the oral reference dose for toxicity during oral ingestion. In conclusion, we report no acute toxicity of single and binary mixture of NPs to spinach but significant accumulation of Ag and Ti metals in spinach leaves. There are high chances that ingestion of spinach grown in such environment might lead to human health risks.

A novel chloroplast super-complex consisting of the ATP synthase and photosystem I reaction center.

Several 'super-complexes' of individual hetero-oligomeric membrane protein complexes, whose function is to facilitate intra-membrane electron and proton transfer and harvesting of light energy, have been previously characterized in the mitochondrial cristae and chloroplast thylakoid membranes. We report the presence of an intra-membrane super-complex dominated by the ATP-synthase, photosystem I (PSI) reaction-center complex and the ferredoxin-NADP+ Reductase (FNR) in the thylakoid membrane. The presence of the super-complex has been documented by mass spectrometry, clear-native PAGE and Western Blot analyses. This is the first documented presence of ATP synthase in a super-complex with the PSI reaction-center located in the non-appressed stromal domain of the thylakoid membrane.

Contamination of spinach at germination: A route to persistence and environmental reintroduction by Salmonella.

The effects of using contaminated seed and water on the persistence and internalization of Salmonella Newport in organic spinach cultivars- Lazio, Space, Emilia and Waitiki were studied. Seeds were contaminated by either immersing in a suspension of Salmonella and then sprouted or were sprouted in Salmonella contaminated water in the dark at 25 °C. After 5 days, germinated sprouts were analyzed for S. Newport population and internalization. Germinated sprouts were potted in soil and grown in a plant incubator for 4 weeks. Leaves, stems and roots were sampled for Salmonella population by plating on CHROMagar™. Plants surface-sterilized with chlorine were analyzed for internalized pathogen. Potting soil and water runoff were sampled for Salmonella after 4 weeks of plant growth. Contaminated seeds and irrigation water had S. Newport populations of 7.64±0.43 log CFU/g and 7.12±0.04 log CFU/ml, respectively. Sprouts germinated using contaminated water or seeds had S. Newport populations of 8.09±0.04 and 8.08±0.03 log CFU/g, respectively and had a Salmonella population that was significantly higher than other spinach tissues (P<0.05). Populations of S. Newport in leaves, stem and roots of spinach plants were as follows: contaminated seed- 2.82±1.69, 1.69±0.86, and 4.41±0.62 log CFU/ml; contaminated water- 3.56±0.90, 3.04±0.31, and 4.03±0.42 log CFU/ml of macerated tissue suspension, respectively. Internalization was observed in plants developing from contaminated seeds and in sprouts germinated using contaminated water. S. Newport populations of 2.82±0.70 log CFU/g and 1.76±0.46 log CFU/ml were recovered from soil and water runoff, respectively. The results indicate that contamination of spinach during germination can result in persistence, internalization and environmental reintroduction of Salmonella.

Biochar counteracts nitrification inhibitor DMPP-mediated negative effect on spinach (Spinacia oleracea L.) growth.

The use of nitrification inhibitors (NIs) such as 3,4-dimethylpyrazole phosphate (DMPP) has been suggested to diminish agricultural soil nitrate (NO3-) loss and increase nitrogen (N) use efficiency (NUE). However, the yield of ammonium (NH4+)-sensitive plants such as spinach (Spinacia oleracea L.) may be adversely affected by the application of NIs at high N levels and, on the other hand, the efficiency of the NIs may also be affected by soil amendments such as biochar. These two issues are still not adequately addressed. The aim of this study was to evaluate the effect of different N levels including DMPP or not in a calcareous soil with and without amendment of wheat straw biochar on spinach yield, NUE, nitrate concentration of spinach leaf, activity of enzymes nitrate reductase (NR) and nitrite reductase (NiR), and soil ammonium (NH4+) and NO3- concentration under greenhouse conditions. This experiment was carried out with different N rates factor at seven levels (un-fertilized, N0; fertilized with 50 mg N kg-1 soil, N50; fertilized with 75 mg N kg-1 soil, N75; fertilized with 100 mg N kg-1 soil, N100; fertilized with N50 + DMPP; fertilized with N75 + DMPP; and fertilized with N100 + DMPP) and biochar (BC) factor at two levels (0, 0%BC; and 2% (w/w), 2%BC) with six replications over a 56-day cultivation period of spinach. Results showed that the application of DMPP had no significant effect on the yield of spinach plant at low and medium levels of N (50 and 75 mg N kg-1 soil), but decreased the yield of this plant at the higher level of N (100 mg N kg-1 soil). However, application of BC decreased the negative effect of DMPP on spinach yield as the yield in spinach plants fertilized with N75 + DMPP and N100 + DMPP significantly increased. Both application of DMPP and addition of BC to soil decreased leaf NO3- concentration by 29.2% and 16.3% compared to control, respectively. Biochar compared to control decreased NR activity by 46.3%. With increasing N rate, NR and NiR activities increased, but DMPP decreased the activities of both enzymes. Biochar reduced the efficiency of DMPP as soil NH4+ concentration was higher in the treatments containing DMPP without BC at 56 days after planting. Biochar and DMPP could increase the quality of spinach plant through decreasing the leaf NO3- concentration. In general, wheat straw biochar counteracted DMPP-mediated negative effect on growth of spinach plant at high level of N by decreasing the efficiency of this inhibitor. These results provide the useful information for managing the application rate of N fertilizers including DMPP in biochar-amended soil.

Comparative nutrient leaching capability of cattle dung biogas digestate and inorganic fertilizer under spinach cropping condition.

A field experiment was used to determine leaching capability of biogas digestate used as soil amendment in comparison to inorganic fertilizer under spinach cropping condition. The biogas digestate used in this experiment was obtained from a biogas production plant that used cattle dung as feedstock. Spinach was cultivated under three treatments, namely, biogas digestate (BD), inorganic fertilizer (IN) and control (no fertilizer or biogas digestate). All the treatments were replicated three times. Irrometer soil solution access tubes were inserted in the middle of each plot for the collection of the nutrient leachates. The leaf area of spinach was significantly higher in IN treatment than BD and control at the harvest stage; BD produced higher fresh mass (479 g/plant) than IN (468 g/plant) and control (201 g/plant). The leachates of inorganic fertilizer showed higher nutrient concentration compared to biogas digestate and control treatments. Twenty-eight days after transplanting, the leachate consisted of 3670.7, 12.12 and 8.5 mg/l of ammonium and 408, 83 and 39 mg/l of phosphate at IN, BD and control treatments, respectively. The study demonstrates that cattle dung biogas digestate can be applied on soil as fertilizer for crops with little or no environmental consequences to water resources and still have the same product quality as inorganic fertilizer.

Yield advantage and cadmium decreasing of rice in intercropping with water spinach under moisture management.

Rice (Oryza sativa L.) intercropping with water spinach (Ipomoea aquatica Forsk) is an effective agricultural practice for safe crop production and for phytoremediation in cadmium-contaminated soil. A field and pot experiment were conducted to investigate the growth and cadmium absorption of rice intercropped with water spinach under different moisture management schemes (continuous flooding, interval flooding, and 75% field capacity). In the field experiment, the concentration of Cd in the grain of rice was significantly lower in the intercropping system than that permitted by the National Food Safety Standard of China (GB 2762-2017). Furthermore, the land equivalent ratio (1.42) was higher in the rice-water spinach intercropping system, indicating a significant advantage of the intercropping system in yield. At the same time, the bio-concentration amount (BCA) of Cd of rice and water spinach in intercropping system significantly increased by 17.99% and 31.98%, respectively (P＜0.05). However, the metal removal equivalent ratio (MRER) of Cd was 1.34, which showed the intercropping system of rice-water spinach had advantage in Cd removal. In the pot experiment, the total iron plaque concentration on the root surface of rice and the pH of the rhizosphere soil were higher under continuous flooding (TCF) than under the control conditions (75% field capacity, TCK), which could significantly decrease the available Cd in the rhizosphere soil and the accumulation of Cd in rice organs. So, this study demonstrated that iron plaque can obstruct and decrease the Cd absorbed by rice in a rice-water spinach intercropping system combined with water management. The intercropping rice with water spinach can achieve the goal of remediation while producing for farmland contaminated by Cd.