Assessing global-warming induced soil organic matter and iron oxides depletion: Empirical insights into sorption and uptake of atrazine by plants.

Recent pesticide use is alarmingly high and unregulated in several parts of the world. Pesticide fate in soil is controlled by sorption processes which affect the subsequent transport and chemical reactivity in the environment, as well as uptake by plants. Sorption processes are dependent on soil composition and properties, but these are beginning to be affected by global warming-linked factors leading to soil depletion. Thus, it is vital to decipher soils' response, especially in the sub-Sahara (SS), to the depletion of some inherent components in the presence of pesticides. This was ascertained by monitoring a model pesticide (atrazine) sorption and desorption on whole SS soil (WS), and the same soil whose organic matter (OMR) and iron oxides (IOR) were substantially depleted, as well as studying atrazine uptake from these soils by fast-growing vegetables. Organic matter depletion enhanced equilibrium in OMR. Sorption was enhanced at lower ambient pH, higher initial atrazine concentration, and higher temperature. Hysteresis was low resulting in high desorption. Overall, atrazine desorption of ≥65 % was observed; it was higher in OMR (≥95 %) since SOM enhanced hysteresis. Though sub-Saharan soils are rich in iron oxides, SOM played a significantly higher role in sorption than iron oxides in this soil. This result suggests a high potential for atrazine to leach into the aquifer in the sub-Saharan. Atrazine uptake experiment by waterleaf and spinach showed that it could be detected in soil after 63 d, and its presence significantly affected the growth of both vegetables especially in soils with depleted SOM and iron oxides, and at high (100 µg/kg) atrazine spiking. Spinach may be a higher atrazine accumulator than waterleaf. It may be concluded that waterleaf and spinach grown on atrazine-contaminated soils, especially on SOM/iron oxide-depleted soils, are likely to accumulate atrazine.

Root rot of spinach caused by Pythium myriotylum and P. aphanidermatum in Taiwan.

Spinach (Spinacia oleracea) is a commonly used green vegetable. During September and October in both 2022 and 2023, a vegetable nursery company located among paddy rice fields in Taichung City, Taiwan, reported significant failures in spinach seedling production in net-houses with mean outdoor temperatures of 28.7℃. Abnormal growth was observed in approximately 30% of the spinach seedlings in each batch (n = 2,000 to 3,000), with aboveground tissues showing stunting, yellowing, and wilt, and underground tissues displaying root rot. The symptoms resembled the spinach damping-off documented in Taiwan in extension articles but which lacked complete pathogen identification. A total of 110 plants from two batches were used for pathogen isolation by placing roots on water agar incubated at 25℃ or were examined for the presence of oospores in diseased roots. Eighty-one percent of these plants were associated with Pythium. Nine Pythium isolates were used in subsequent analyses. Genomic DNA from these isolates was subjected to amplification of ITS, ß-tubulin gene (TUB2), and cytochrome C oxidase subunit Ⅱ (COXII) gene with primer pairs ITS1 / ITS4, BT5 / BT6, and FM58 / FM66 (Villa et al. 2006). Sequences of ITS (PP209187-PP209195), TUB2 (PP212864-PP212872), and COXII (PP212855-PP212863) were deposited in GenBank. Four isolates (sp01, sp02, sp03, and sp04) were 100% identical to the neotype strain (CBS 118.80) of Pythium aphanidermatum (Edson) Fitzp. for the ITS (761 bp), TUB2 (583 bp), and COXII (547 bp). Five isolates (2sp, 3sp, ND2-4sp, D3-4sp, and ND3-3sp) were 99.87%, 100%, and 99% identical to the reference strain (CBS 254.70) of Pythium myriotylum Drechsler for the ITS (762 bp), TUB2 (602 bp), and COXII (556 bp), respectively. Phylogenetic analysis of Pythium isolates inferred from concatenated sequences of the three genes (LéVesque and De Cock 2004; Villa et al. 2006) revealed that the same four isolates grouped with the neotype strain of P. aphanidermatum, and the five isolates clustered with the reference strain of P. myriotylum, each with a 100% bootstrap support. Morphological features of isolates ND3-3sp and sp01 were used for identification. Isolate ND3-3sp produced inflated lobulate sporangia and aplerotic and smooth oospores (16.3 to 25.1 um; n = 30) attached with three to five antheridia, consistent with identification as P. myriotylum. Isolate sp01 produced inflated lobulate sporangia and aplerotic and smooth oospores (17.0 to 24.0 um; n= 30) attached with a single intercalary antheridium, agreeing with the morphology of P. aphanidermatum (Van der Plaats-Niterink 1981). To investigate the pathogenicity of the nine Pythium isolates on spinach, 20 mycelial agar discs (4 mm in diameter) from a 2-day-old V8 culture of each isolate were used to induce sporangia and zoospores in 20 ml sterilized water at 25℃ with a 12 h light / dark regime. A 1.5 ml zoospore suspension (6 × 103 zoospores / ml) was dropped into BVB growth substrate of two spinach seedlings in 2-week-old at 25℃ with 12 h light / dark regime, resulting in symptoms resembling those observed in commercial nurseries at 7 days post-inoculation (dpi). Each Pythium isolate inoculated 20 seedlings in 10 cells of a planting tray. At 14 dpi, disease incidences were 95 to 100% for P. myriotylum isolates and 60 to 85% for P. aphanidermatum isolates, while control plants treated with water showed no symptoms. Re-isolated pathogens from the inoculated plants were morphologically identical to the inoculated isolates, completing Koch's postulates. Results of the pathogenicity assay, along with molecular and morphological identification, conclude that the root rot of spinach was caused by P. myriotylum and P. aphanidermatum. The two oomycetes were not formally documented to cause spinach diseases in Taiwan. Although P. myriotylum has been isolated from spinach (Wang et al. 2003), its pathogenicity to spinach was not documented worldwide. Root rot of spinach caused by P. aphanidermatum has been reported in the United States (Bates and Stanghellini 1984), Korea (Cho and Shin 2004), and Italy (Garibaldi et al. 2015). These pathogens thrive in humid and hot weather (Littrell and McCarter, 1970). Producing spinach in cooler weather or in a temperature-controlled environment may help prevent severe occurrence of the disease.

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.

Guard Cell Transcriptome Reveals Membrane Transport, Stomatal Development and Cell Wall Modifications as Key Traits Involved in Salinity Tolerance in Halophytic Chenopodium quinoa.

A comparative investigation was conducted to evaluate transcriptional changes in guard cells (GCs) of closely related halophytic (Chenopodium quinoa) and glycophytic (Spinacia oleracea) species. Plants were exposed to 3 weeks of 250 mM sodium chloride treatment, and GC-enriched epidermal fragments were mechanically prepared. In both species, salt-responsive genes were mainly related to categories of protein metabolism, secondary metabolites, signal transduction and transport systems. Genes related to abscisic acid (ABA) signaling and ABA biosynthesis were strongly induced in quinoa but not in spinach GCs. Also, expression of the genes encoding transporters of amino acids, proline, sugars, sucrose and potassium increased in quinoa GCs under salinity stress. Analysis of cell-wall-related genes suggests that genes involved in lignin synthesis (e.g. lignin biosynthesis LACCASE 4) were highly upregulated by salt in spinach GCs. In contrast, transcripts related to cell wall plasticity Pectin methylesterase3 (PME3) were highly induced in quinoa. Faster stomatal response to light and dark measured by observing kinetics of changes in stomatal conductance in quinoa might be associated with higher plasticity of the cell wall regulated by PME3 Furthermore, genes involved in the inhibition of stomatal development and differentiation were highly expressed by salt in quinoa, but not in spinach. These changes correlated with reduced stomatal density and index in quinoa, thus improving its water use efficiency. The fine modulation of transporters, cell wall modification and controlling stomatal development in GCs of quinoa may have resulted in high K+/Na+ ratio, lower stomatal conductance and higher stomatal speed for better adaptation to salinity stress in quinoa.

Changes in morphological traits, anatomical and molecular alterations caused by gamma-rays and zinc oxide nanoparticles in spinach (Spinacia oleracea L.) plant.

Spinach seeds were irradiated with gamma-rays after that soaked in zinc oxide nanoparticles (ZnO-NPs) at 0.0, 50, 100 and 200 ppm for twenty-four hours at room temperature. Vegetative plant growth, photosynthetic pigments, and proline contents were investigated. Also, anatomical studies and the polymorphism by the SCoT technique were conducted. The present results revealed that the germination percentage was at the maximum values for the treatment of 100 ppm ZnO-NPs (92%), followed by 100 ppm ZnO-NPs + 60 Gy (90%). The application of ZnO-NPs resulted in an enhancement in the plant length. The maximum of chlorophylls and carotenoids content was recorded in the treatment, 100 ppm ZnO-NPs + 60 Gy. Meanwhile, the irradiation dose level (60 Gy) with all ZnO-NPs treatments increased proline content and reached its maximum increase to 1.069 mg/g FW for the treatment 60 Gy combined with 200 ppm ZnO-NPs. Also, the anatomical studies declared that there were variations between the treatments; un-irradiated and irradiated combined with ZnO-NPs plants which reveal that the leave epidermal tissue increased with 200 ppm ZnO-NPs in both the upper and lower epidermis. While irradiated plants with 60 Gy combined with 100 ppm ZnO-NPs gave more thickness of upper epidermis. As well as SCoT molecular marker technique effectively induced molecular alterations between the treatments. Where, SCoT primers targeted many new and missing amplicons that are expected to be associated with the lowly and highly expressed genes with 18.2 and 81.8%, respectively. Also, showed that the soaking in ZnO-NPs was helped for reducing molecular alteration rate, both spontaneous and induced by gamma irradiation. This nominates ZnO-NPs as potential nano-protective agents that can reduce irradiation-induced genetic damage.

Real-Time PCR Assays for Races of the Spinach Fusarium Wilt Pathogen, Fusarium oxysporum f. sp. spinaciae.

Fusarium wilt of spinach, caused by Fusarium oxysporum f. sp. spinaciae, is a significant limitation for producers of vegetative spinach and spinach seed crops during warm temperatures and/or on acid soils. Identification of isolates of F. oxysporum f. sp. spinaciae, and distinction of isolates of the two known races, entails time-intensive pathogenicity tests. In this study, two real-time PCR assays were developed: one for a candidate effector gene common to both races of F. oxysporum f. sp. spinaciae, and another for a candidate effector gene unique to isolates of race 2. The assays were specific to isolates of F. oxysporum f. sp. spinaciae (n = 44) and isolates of race 2 (n = 23), respectively. Neither assay amplified DNA from 10 avirulent isolates of F. oxysporum associated with spinach, 57 isolates of other formae speciales and Fusarium spp., or 7 isolates of other spinach pathogens. When the assays were used to detect DNA extracted from spinach plants infected with an isolate of race 1, race 2, or a 1:1 mixture of both races, the amount of target DNA detected increased with increasing severity of wilt. Plants infected with one or both isolates could be distinguished based on the ratio in copy number for each target locus. The real-time PCR assays enable rapid diagnosis of Fusarium wilt of spinach and will facilitate research on the epidemiology and management of this disease, as well as surveys on the prevalence of this understudied pathogen in regions of spinach and/or spinach seed production.

Shedding Light on Races of the Spinach Fusarium Wilt Pathogen, Fusarium oxysporum f. sp. spinaciae.

Two pathogenicity groups of Fusarium oxysporum f. sp. spinaciae, the causal agent of Fusarium wilt of spinach (Spinacia oleracea), were described recently based on virulence of isolates on proprietary spinach inbreds. In this study, a wide range in severity of wilt was observed for 68 spinach cultivars inoculated with an isolate of each pathogenicity group, with 22 (32.4%) cultivars displaying differential responses to the isolates. In a second set of trials, seven spinach cultivars were inoculated with five isolates of each pathogenicity group. The cultivars had similar wilt responses to isolates within each group. In both sets of trials, the most severe wilt developed on cultivars inoculated with pathogenicity group 2 isolates when daylength was shorter and light intensity lower. To test whether light intensity exacerbates severity of Fusarium wilt, three spinach cultivars were inoculated with two isolates of each pathogenicity group and grown with or without shading. Shaded plants developed more severe wilt than nonshaded plants. This difference in wilt severity was greatest for plants inoculated with pathogenicity group 2 isolates. We propose naming isolates of pathogenicity groups 1 and 2 as races 1 and 2 of F. oxysporum f. sp. spinaciae, respectively, and recommend the cultivars Kiowa (susceptible to both races) and Magnetic (susceptible to race 2 and highly resistant to race 1) as differentials. Results of this study should help breeders screen spinach germplasm for resistance to both races of F. oxysporum f. sp. spinaciae.

Pre-stress salicylic-acid treatment as an intervention strategy for freeze-protection in spinach: Foliar versus sub-irrigation application and duration of efficacy.

Exogenous application of salicylic acid (SA) to plant tissues has been shown to confer tolerance against various abiotic stresses. Recently, SA application through sub-irrigation was shown to improve plant freezing tolerance (FT). For SA treatment to be employable as an effective intervention strategy for frost protection under field conditions, it is important to study its effect on FT when applied as a foliar spray to whole plants. It is also important to determine for how long the FT-improvement by SA lasts. Present study was conducted to compare SA-induced FT of spinach (Spinacia oleracea L. 'Reflect') seedlings following SA-application by foliar spray vs. sub-irrigation. Durability of FT-promotive effect of SA was evaluated using three freeze-tests over a 4-d period, i.e., at 10-d, 12-d, and 14-d after the SA application. Freezing stress was applied using a temperature-controlled freeze-thaw protocol, and FT was assessed by visual observations (leaf flaccidness vs. turgidity) as well as ion-leakage assay. Data indicated that both foliar spray and sub-irrigation methods improved FT of the seedlings against a relatively moderate (-5.5 °C) as well as severe stress (-6.5 °C). Moreover, improved FT against moderate stress was sustained over a 4-d period, whereas such benefit waned somewhat against the severe stress. SA-treated leaves' growth performance was similar to the non-treated control based on dry weight, fresh weight, leaf area, and dry weight/leaf area parameters. Our results suggest that SA application as a foliar spray can potentially be used to protect field-grown transplants against episodic frosts.

Lemon Juice Formulations Modulate In Vitro Digestive Recovery of Spinach Phytochemicals.

Research background: Citrus limon (L.) Burm lemon juice is rich in many important natural chemical components (flavonoids, citric acid and vitamin C) and its use in traditional medicine is well known. Formulations of lemon juice with fruit polyphenols in beverages have been investigated, but there is very little information about their ability to modulate the digestive behaviour of polyphenols. The goal of this study is to determine the stability and digestive availability of spinach (Spinacia oleracea L.) polyphenols by adding different volume fractions of lemon juice (0, 2, 5, 10 and 20%) during in vitro digestion. Experimental approach: The content of polyphenols and other abundant compounds including nitrates, oxalic acid and l-ascorbic acid in spinach formulation with various volume fractions of lemon juice were measured in predigested and digested samples using in vitro human digestion model. Antioxidant and α-amylase inhibitory activities of spinach lemon juice formulation were also measured. Results and conclusions: The highest increases in total polyphenols, total flavonoids, total phenolic acids, oxalic acid and nitrate content were noted in predigested and almost all digested spinach samples formulated with the highest volume fraction of lemon juice. In the same sample, the content of individual compounds significantly increased after salivary (l-ascorbic acid), initial (p-coumaric acid) and intestinal (quercetin) phase of digestion. High bioaccessibility of polyphenols and l-ascorbic acid in all phases of digestion was observed in almost all spinach lemon juice formulations, with the exception of nitrates in gastric and intestinal phases and oxalic acid in the intestinal phase, which had moderate bioaccessibility. Novelty and scientific contribution: For the first time the stability and digestive availability of spinach polyphenols, oxalic acid, nitrates and l-ascorbic acid were tested with the addition of different volume fractions of lemon juice. The pH of lemon juice and its l-ascorbic acid content increase the stability and availability of polyphenols in spinach lemon juice formulation during in vitro digestion. Antioxidant and α-amylase inhibitory activities increase in dose-dependent manner after lemon juice addition. Accordingly, spinach formulated with 20% of lemon juice appears as the best source of dietary polyphenols with antioxidant and antidiabetic activities and nitrates that may be used as a functional drink.

Unrevealing arsenic and lead toxicity and antioxidant response in spinach: a human health perspective.

Arsenic (As) and lead (Pb) are highly toxic and carcinogenic metal(loid)s. The present study evaluated the human exposure risk via estimating As and Pb uptake and physiological/biochemical modifications inside spinach plant grown under metal(loid)-contaminated growth medium. Plants were treated with three levels of each metal(loid) (0, 25 and 125 µM) for four weeks. The spinach plants accumulated high concentration of metal(loid)s in roots (0-18.9 ug g-1 Pb and 0.2-22.7 ug g-1 As) and less were translocated towards shoot (0-0.3 ug g-1 Pb and 0.2-8.8 ug g-1 As). Metal(loid) accumulation in plants decreased plant biomass and pigment contents and provoked oxidative stress by increased hydrogen peroxide (H2O2) production in roots up to 65% and 22%, respectively, for As and Pb. The production of H2O2 in leaves was decreased up to 59% and 45%, respectively, for As and Pb than control. Moreover, the antioxidant system (superoxide, catalase, guaiacol peroxidase, ascorbate peroxidase) gets activated under metal(loid) stress. The exposure assessment indices revealed high carcinogenic (CR > 10-4) and non-carcinogenic (HQ > 1) risks owing to the consumption of As- and Pb-contaminated spinach leaves. Results revealed As is being more toxic to plants and humans than Pb. These findings suggest possible alarming consequences of As and Pb to spinach and their assimilation within the edible tissues.

Putative Effector Genes Distinguish Two Pathogenicity Groups of Fusarium oxysporum f. sp. spinaciae.

Fusarium wilt of spinach, caused by Fusarium oxysporum f. sp. spinaciae, is an important disease during warm conditions in production regions with acid soils, yet little is known about what confers pathogenicity to spinach in F. oxysporum f. sp. spinaciae genetically. To identify candidate fungal genes that contribute to spinach Fusarium wilt, each of 69 geographically diverse F. oxysporum isolates was tested for pathogenicity on each of three spinach inbreds. Thirty-nine isolates identified as F. oxysporum f. sp. spinaciae caused quantitative differences in disease severity among the inbreds that revealed two distinct pathogenicity groups of F. oxysporum f. sp. spinaciae. Putative effector gene profiles, predicted from whole-genome sequences generated for nine F. oxysporum f. sp. spinaciae isolates and five nonpathogenic, spinach-associated F. oxysporum (NPS) isolates, distinguished the F. oxysporum f. sp. spinaciae isolates from the NPS isolates, and separated the F. oxysporum f. sp. spinaciae isolates into two groups. Five of the putative effector genes appeared to be unique to F. oxysporum f. sp. spinaciae, as they were not found in 222 other publicly available genome assemblies of F. oxysporum, implicating potential involvement of these genes in pathogenicity to spinach. In addition, two combinations of the 14 known Secreted in Xylem (SIX) genes that have been affiliated with host pathogenicity in other formae speciales of F. oxysporum were identified in genome assemblies of the nine F. oxysporum f. sp. spinaciae isolates, either SIX8 and SIX9 or SIX4, SIX8, and SIX14. Characterization of these putative effector genes should aid in understanding mechanisms of pathogenicity in F. oxysporum f. sp. spinaciae, developing molecular tools for rapid detection and quantification of F. oxysporum f. sp. spinaciae, and breeding for resistance to Fusarium wilt in spinach.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Commercial wash of leafy vegetables do not significantly decrease bacterial load but leads to shifts in bacterial species composition.

Production of leafy vegetables for the "Ready-to-eat"-market has vastly increased the last 20 years, and consumption of these minimally processed vegetables has led to outbreaks of food-borne diseases. Contamination of leafy vegetables can occur throughout the production chain, and therefore washing of the produce has become a standard in commercial processing. This study explores the bacterial communities of spinach (Spinacia oleracea) and rocket (Diplotaxis tenuifolia) in a commercial setting in order to identify potential contamination events, and to investigate effects on bacterial load by commercial processing. Samples were taken in field, after washing of the produce and at the end of shelf-life. This study found that the bacterial community composition and diversity changed significantly from the first harvest to the end of shelf-life, where the core microbiome from the first to the last sampling constituted <2% of all OTUs. While washing of the produce had no reducing effect on bacterial load compared to unwashed, washing led to a change in species composition. As the leaves entered the cold chain after harvest, a rise was seen in the relative abundance of spoilage bacteria. E. coli was detected after the washing indicating issues of cross-contamination in the wash water.

Ca2SiO4 chemigation reduces cadmium localization in the subcellular leaf fractions of spinach (Spinacia oleracea L.) under cadmium stress.

Heavy metal like cadmium (Cd) is inessential and highly toxic and is posing serious environmental problems for agriculture worldwide. Presence of Cd gives rise to several physiological and structural disorders that leads to reduction in growth and performance of agricultural plants. Evidence related to subcellular distribution and accumulation of Cd is still enigmatic. Experiment was conducted using hydroponic culture to examine the subcellular accumulation of Cd in Spinacia oleracea L. leaves under Cd stress (50 µM and 100 µM); moreover, the Cd toxicity alleviation using 5 mM silicon (Si) was investigated. Our findings suggest that fresh and dry biomass, shoot and root length, leaf area and length of leaf declined when exposed to Cd stress (50 µM and 100 µM); however, an increase was noticed when Cd treated plants were supplied with Si (5 mM). The content of Ca2+, Mg2+ and Fe2+ in apoplastic washing fluid and symplasm were found to be lower in plants treated with alone Cd, when compared to control. Higher Cd2+:Ca2+, Cd2+:Fe2+ and Cd2+:Mg2+ ratios were detected under cadmium stress in both apoplast and symplast of leaves which were lowered by the addition of 5 mM Si. The novelty of the current study is the detection of increased apoplastic and symplastic Cd concentration in aerial part (i.e., spinach leaves) under alone Cd treatment which was considerably reduced when supplied with Si. Moreover, a noticeable increase in spinach growth and beneficial ionic concentrations suggest that Si can ameliorate the Cd stress in crop plants.

Comparative study of the insecticidal activity of a high green plant (Spinacia oleracea) and a chlorophytae algae (Ulva lactuca) extracts against Drosophila melanogaster fruit fly.

OBJECTIVES: Currently, the global interests tend to take advantage of the plant world as a renewable source of a natural and effective molecule, to find an eco-friendly, cost-effective, and less toxic alternative to the current synthetic pesticide. In this context, the present research was carried out in an attempt to study the insecticidal activity of extracts and pigments derived from the green plant Spinacia oleracea and the green alga Ulva lactuca against the fruit fly Drosophila melanogaster as an alternative to chemical insecticide. METHODS: The toxicity of the aqueous, acetonic and ethanolic extracts as well as of the purified pigments (Chlorophylls and carotenoids) was determined by complementary in vivo tests (application by spraying oranges, toxicity by ingestion and repellent activity). Interestingly, each one of these methods corresponds to a specific mode of exposure. RESULTS: Results showed that acetone extracts, which are rich in green pigments, present the best insecticidal activities. On the other hand, the purified chlorophyllian pigments exhibited an interesting activity only by spraying method. Regarding the repellent activity, the aqueous extract of spinach displayed higher effectiveness. CONCLUSION: Our study suggests the potential of tested plant and algal extracts, as well as of chlorophyllian pigments, to provide a safer alternative way to the use of synthetic pesticides.

Predicting heavy metals uptake by spinach (Spinacia oleracea) grown in integrated industrial wastewater irrigated soils of Haridwar, India.

This investigation aimed to assess the impacts of integrated industrial wastewater (IIW) irrigation on soil properties in the rural area of Haridwar, India, under cultivation of a leafy vegetable, i.e., spinach (Spinacia oleracea). Based on the field data of two cropping years (2016-2017 and 2017-2018), soil characteristics-based prediction models were developed to evaluate heavy metals (HM) uptake by spinach tissues (roots and leaves) using the multivariate regression method. The results showed a significant increase (P < 0.05) in the growth and productivity of spinach plants in IIW irrigated soils as compared to normal borewell water irrigation. For the prediction models, soil parameters including pH, organic matter (%), and HM (mg/Kg) availability showed a significant effect on the HM absorption process by spinach tissues. Besides this, the models were tested using ANOVA (P < 0.001), Student's t test, model efficiency (> 0.50), and coefficient of determination (R2 > 0.81) tools. Furthermore, the prediction models were also verified for their applicability in the 2018-2019 cropping year which gave satisfactory outcomes. The findings of this investigation are important in terms of predicting hazardous HM accumulation in the vegetable crops being grown in wastewater irrigated soils.

Spinacia oleracea L. extract attenuates hippocampal expression of TNF-α and IL-1ß in rats exposed to chronic restraint stress.

Background: Restraint stress causes inflammation in nervous system that leads to emersion of neurodegenerative diseases. Spinach (Spinacia oleracea L.) contains different agents with antioxidant, antiapoptosis, and hepatoprotective properties. This study examined the effect of spinach hydroalcoholic extract (SHE) on TNF-α and IL-1ß expression in hippocampus of male Wistar rats exposed to chronic restraint stress. Methods: Rats were divided into 6 groups of 5: (1) control (intact); (2) nS-S200; (3) nS-S400; (4) stress; (5) stress-S200; (6) stressS400. Groups 2 and 3 and groups 5 and 6 received S. oleracea leaf hydroalcoholic extract in 200 and 400 mg/kg doses for 21 consecutive days by gavage. Groups 4, 5 and 6 were put in a restrainer 6 hours per day for 21 consecutive days. Then, the expression of IL-1ß and TNF-α mRNAs and neuronal death in the hippocampus of rats were assessed by real time PCR and Nissl staining, respectively. Oneway analysis of variance was used for data analysis, and p<0.05 was considered statistically significant. Results: The results showed that the expression of IL-1ß and TNF-α was increased in hippocampus of rats exposed to stress compared to control groups (p<0.001). Furthermore, the expression of these proinflammatory cytokines was decreased in the stress-S200 and stress-S400 groups when compared to stress group (p<0.001). Immobility also caused neuronal death in CA1 region of hippocampus, and SHE reduced damage in CA1 pyramidal neurons layer in stressed rats. Conclusion: Spinach decreases neuroinflammation in hippocampus of stressed rats, which may be due to its abundant antiinflammatory and antioxidant phytochemicals. The results of this study suggest that spinach may be effective in the prevention and treatment of neurodegenerative diseases.

The Effects of Biostimulants, Biofertilizers and Water-Stress on Nutritional Value and Chemical Composition of Two Spinach Genotypes (Spinacia oleracea L.).

In the present study, the effect of biostimulants application on the nutritional quality and bioactive properties of spinach cultivated in protected environment under water stress conditions was evaluated. For this purpose, four commercially available biostimulant products (Megafol (MEG), Aminovert (AM), Veramin Ca (V), Twin Antistress (TA), and two spinach genotypes (Fuji F1 and Viroflay) were tested under two irrigation regimes (normal irrigation (W+), and water-holding (W-). Fat and carbohydrates content was favored by water stress when Megafol (MEGW+) and Veramin (VW+) were applied on Fuji plants, while calorific value was also increased by MEGW+ treatment. In contrast, protein and ash content increased when AMW- and TAW+ were applied on Viroflay plants. Raffinose and glucose were the most abundant sugars, followed by sucrose and fructose, with the highest contents recorded for Fuji plants when AMW+ (fructose, glucose and total carbohydrates), CW- (sucrose), and TAW- (raffinose) treatments were applied. Regarding organic acids, oxalic and malic acid which had the highest contents for the TAW- (Viroflay plants) and AMW- (Fuji plants) treatments, respectively. α- and γ-tocopherol were the only isoforms detected with MEGW- and VW- inducing the biosynthesis of α-tocopherol, while AMW+ increased γ-tocopherol content in Fuji plants. The main fatty acids were α-linolenic and linoleic acids which were detected in the highest amounts in AMW-, AMW+, and TAW+ the former and in AMW-, VW-, and CW+ the latter. Regarding phenolic compounds content, peak 12 (5,3',4'-Trihydroxy-3-methoxy-6:7-methylenedioxyflavone-4'-glucuronide) was the most abundant compound, especially in Viroflay plants under normal irrigation and no biostimulants added (CW-). The antioxidant and cytotoxic activity of the tested samples did not show promising results when compared with the positive controls, while a variable antibacterial activity was recorded depending on the tested biostimulant, irrigation regime and genotype. In conclusion, a variable effect of the tested biostimulants and irrigation regimes was observed on bioactive properties and chemical composition of both spinach genotypes which highlights the need for further research in order to make profound conclusions regarding the positive effects of biostimulants under water stress conditions.

Iodine uptake, storage and translocation mechanisms in spinach (Spinacia oleracea L.).

Iodine is an essential micronutrient for human health; phytofortification is a means of improving humans' nutritional iodine status. However, knowledge of iodine uptake and translocation in plants remains limited. In this paper, plant uptake mechanisms were assessed in short-term experiments (24 h) using labelled radioisotopes; the speciation of iodine present in apoplastic and symplastic root solutions was determined by (HPLC)-ICP-QQQ-MS. Iodine storage was investigated in spinach (Spinacia oleracea L.) treated with I- and IO3-. Finally, translocation through the phloem to younger leaves was also investigated using a radioiodine (129I-) label. During uptake, spinach roots demonstrated the ability to reduce IO3- to I-. Once absorbed, iodine was present as org-I or I- with significantly greater concentrations in the apoplast than the symplast. Plants were shown to absorb similar concentrations of iodine applied as I- or IO3-, via the roots, grown in an inert growth substrate. We found that whilst leaves were capable of absorbing radioactively labelled iodine applied to a single leaf, less than 2% was transferred through the phloem to younger leaves. In this paper, we show that iodine uptake is predominantly passive (approximately two-thirds of total uptake); however, I- can be absorbed actively through the symplast. Spinach leaves can absorb iodine via foliar fertilisation, but translocation is severely limited. As such, foliar application is unlikely to significantly increase the iodine content, via phloem translocation, of fruits, grains or tubers.

Gender-specific expression of GIBBERELLIC ACID INSENSITIVE is critical for unisexual organ initiation in dioecious Spinacia oleracea.

While unisexual flowers have evolved repeatedly throughout angiosperm families, the actual identification of sex-determining genes has been elusive, and their regulation within populations remains largely undefined. Here, we tested the mechanism of the feminization pathway in cultivated spinach (Spinacia oleracea), and investigated how this pathway may regulate alternative sexual development. We tested the effect of gibberellic acid (GA) on sex determination through exogenous applications of GA and inhibitors of GA synthesis and proteasome activity. GA concentrations in multiple tissues were estimated by enzyme-linked immunosorbent assay analysis. Gene function was investigated and pathway analysis was performed through virus-induced gene silencing. Relative gene expression levels were estimated by quantitative reverse transcription-polymerase chain reaction. Inhibition of GA production and proteasome activity feminized male flowers. However, there was no difference in GA content in tissues between males and females. We characterized a single DELLA family transcription factor gene (GIBBERELLIC ACID INSENSITIVE (SpGAI)) and observed inflorescence expression in females two-fold higher than in males. Reduction of SpGAI expression in females to male levels phenocopied exogenous GA application with respect to flower development. These results implicate SpGAI as the feminizing factor in spinach, and suggest that the feminizing pathway is epistatic to the masculinizing pathway. We present a unified model for alternative sexual development and discuss the implications for established theory.

Higher anthocyanin accumulation associated with higher transcription levels of anthocyanin biosynthesis genes in spinach.

Spinach (Spinacia oleracea L.) is widely cultivated as an economically important green leafy vegetable crop for fresh and processing consumption. The red-purple spinach shows abundant anthocyanin accumulation in the leaf and leaf petiole. However, the molecular mechanisms of anthocyanin synthesis in this species are still undetermined. In the present study, we investigated pigment formation and identified anthocyanin biosynthetic genes in spinach. We also analyzed the expression of these genes in purple and green cultivars by quantitative PCR. The accumulation of anthocyanin showed that it was the dominant pigment resulting in the red coloration in spinach. In total, 22 biosynthesis genes and 25 regulatory genes were identified in spinach, based on the spinach genomic and transcriptomic database. Furthermore, the expression patterns of genes encoding enzymes indicated that SoPAL, SoUFGT3, and SoUFGT4 were possible candidate genes for anthocyanin biosynthesis in red-purple spinach. The expression patterns of transcription factors indicated that two SoMYB genes, three SobHLH genes, and one SoWD40 gene were drastically up-regulated and co-expression in red-purple spinach, suggesting an essential role of regulatory genes in the anthocyanin biosynthesis of spinach. These results will enhance our understanding of the molecular mechanisms of anthocyanin biosynthesis in purple spinach.