Ecotoxicological effects of paracetamol on the biochemical and molecular responses of spinach (Spinacia oleracea L.).

The widespread use of pharmaceuticals, including paracetamol, has raised concerns about their impact on the environment and non-target species. The aim of this study was to investigate the biochemical and molecular responses of Spinacia oleracea (spinach) to high paracetamol concentrations in order to understand the plant's stress responses and underlying mechanisms. Under controlled conditions, spinach plants were exposed to different paracetamol concentrations (0, 50, 100, and 200 mg/L). The study evaluated the impact of paracetamol exposure on biochemical parameters such as oxidative stress markers (H2O2, MDA), activities of antioxidant enzymes (APX, CAT, GPOD, SOD), levels of non-enzymatic components (phenolics and flavonoids), and phytohormones (ABA, SA, and IAA). Furthermore, the study assessed molecular impacts by analyzing stress-related genetic variation and alterations in the gene expression of the antioxidant enzymes. Results showed that paracetamol exposure significantly increased oxidative stress in spinach, which was evident through the elevated H2O2 and MDA levels. However, the antioxidant defense mechanisms were activated to counteract this effect, as evidenced by increased activity of antioxidant enzymes and higher phenolics and flavonoid levels. Moreover, induction in the phytohormone levels indicated a stress response in paracetamol-treated plants compared to control plants. RAPD analysis revealed polymorphism indicating the DNA damage, and the Real-time qRT-PCR method showed significant upregulation of stress-responsive genes, highlighting the severe impact of paracetamol at the molecular level. The study concludes that high paracetamol concentrations pose a significant threat to spinach growth by affecting both biochemical and molecular processes. These findings underscore the need for strict environmental management practices to mitigate the possible impact of continuous release, accumulation, and long-term exposure of pharmaceutical contaminants to the environment and implement policies to reduce pharmaceutical pollutants to preserve ecological health and biodiversity.

Dual transcriptional characterization of spinach and Peronospora effusa during resistant and susceptible race-cultivar interactions.

BACKGROUND: Spinach downy mildew, caused by the obligate oomycete pathogen, Peronospora effusa remains a major concern for spinach production. Disease control is predominantly based on development of resistant spinach cultivars. However, new races and novel isolates of the pathogen continue to emerge and overcome cultivar resistance. Currently there are 20 known races of P. effusa. Here we characterized the transcriptomes of spinach, Spinacia oleracea, and P. effusa during disease progression using the spinach cultivar Viroflay, the near isogenic lines NIL1 and NIL3, and P. effusa races, R13 and R19, at 24 h post inoculation and 6 days post inoculation. A total of 54 samples were collected and subjected to sequencing and transcriptomic analysis. RESULTS: Differentially expressed gene (DEG) analysis in resistant spinach interactions of R13-NIL1 and R19-NIL3 revealed spinach DEGs from protein kinase-like and P-loop containing families, which have roles in plant defense. The homologous plant defense genes included but were not limited to, receptor-like protein kinases (Spiol0281C06495, Spiol06Chr21559 and Spiol06Chr24027), a BAK1 homolog (Spiol0223C05961), genes with leucine rich repeat motifs (Spiol04Chr08771, Spiol04Chr01972, Spiol05Chr26812, Spiol04Chr11049, Spiol0084S08137, Spiol03Chr20299) and ABC-transporters (Spiol02Chr28975, Spiol06Chr22112, Spiol06Chr03998 and Spiol04Chr09723). Additionally, analysis of the expression of eight homologous to previously reported downy mildew resistance genes revealed that some are differentially expressed during resistant reactions but not during susceptible reactions. Examination of P. effusa gene expression during infection of susceptible cultivars identified expressed genes present in R19 or R13 including predicted RxLR and Crinkler effector genes that may be responsible for race-specific virulence on NIL1 or NIL3 spinach hosts, respectively. CONCLUSIONS: These findings deliver foundational insight to gene expression in both spinach and P. effusa during susceptible and resistant interactions and provide a library of candidate genes for further exploration and functional analysis. Such resources will be beneficial to spinach breeding efforts for disease resistance in addition to better understanding the virulence mechanisms of this obligate pathogen.

Potential to Ensure Safe Production of Water Spinach in Heavy Metals-Contaminated Soil by Substituting Chemical Fertilizer with Organic Fertilizer.

Organic fertilizers are widely used to improve soil quality. However, their potential for ensuring the safe production of vegetables in soils with varying levels of heavy metals pollution remains inadequately explored. Here, we conducted a pot experiment to investigate the effects of substituting chemical fertilizers with organic fertilizer on the HMs accumulation in water spinach by simulating soils with different levels of HMs pollution. The results showed that the organic fertilizer significantly increased the soil pH, cation exchange capacity (CEC), and organic matter (OM). Furthermore, it led to a reduction in the soil DTPA-Cd and DTPA-Pb levels by 3.3-20.6% and 22.4-47.3%, respectively, whereas the DTPA-As levels increased by 0.07-7.7 times. The organic fertilizer effectively reduced the Cd and Pb content in water spinach below the safety limits when the added Cd content in the soil was less than 2 mg/kg and the Pb content was equal to or less than 90 mg/kg. However, its efficacy in reducing As accumulation in water spinach was limited, emphasizing the need for caution when using organic fertilizers in As-contaminated soils. Our results provide valuable insights for the scientific and precise utilization of organic fertilizers, thereby contributing to the safe production of vegetables.

Circular Utilization of Coffee Grounds as a Bio-Nutrient through Microbial Transformation for Leafy Vegetables.

This study explores the production of bio-nutrients from bioactive compound-rich spent coffee grounds (SCG) and biochar (BC) through composting after inoculation with a biological agent and its impact on the growth performance of garden cress and spinach. The SCG was composted with six doses of BC (0, 5, 10, 15, 20, and 25%). The compost with 10% BC exhibited the best maturity, humification, and phytotoxicity index values of dissolved organic carbon (DOC), humification index (E4/E6), and germination index (GI). A metagenome analysis showed that compost starter enhanced the bacterial community's relative abundance, richness, and diversity in SCG and BC treatments. This improvement included increased Patescibacteria, which can break down noxious phenolic compounds found in SCG and BC. The BC enriched the compost with phosphorus and potassium while preserving the nitrogen. In plant growth experiments, the total chlorophyll content in compost-treated garden cress and spinach was 2.47 and 4.88 mg g-1, respectively, which was significantly greater (p ≤ 0.05) than in unfertilized plants and similar to the plants treated with traditional fertilizer. Overall, the results show that the compost of SCG + BC was well-suited for promoting the growth of garden cress and spinach, providing adequate nutrients as a fertilizer for these leafy vegetables.

Biocompatible and Safe Decellularized Spinach With Antibacterial and Wound Healing Activity.

Creating acellular vascularized constructs from animal and plant tissue is one of the well-known strategies for scaffold assembly. Decellularization takes an important position among these strategies. The most common method is chemical decellularization. This approach employs high concentrations of detergents, primarily Triton X-100, sodium dodecyl sulfate (SDS), and sodium hypochlorite (SH). In this work, novel techniques for decellularizing spinach were developed using detergents frequently utilized in laboratories. Spinach leaves were decellularized using Tween-20, SDS, and SH at low concentrations to generate an acellular plant matrix for tissue engineering. We measured the quantities of DNA and protein, as well as the decellularization using hematoxylin and eosin (H&E) staining. The biocompatibility and capacity of the biostructures to stimulate fibroblast wound healing were assessed using MTT and the Scratch assay. The antibacterial activity of the scaffolds was also tested against a gram-positive bacterium, Staphilococcus aureus, which is a common pathogen associated with wound healing. The best shape, evident vascularization, and good biocompatibility were seen in the Tween-20 decellularized samples at 1% concentration at 21°C and 37°C through the enhancement of cell proliferation and wound healing. In terms of antibacterial activity, all scaffold samples had a significant effect on Staphilococcus aureus, where the number of bacterial colonies in all six scaffold groups became zero after 4 h of treatment. The scaffolds also showed a 100% kill rate on Staphilococcus aureus, which could avoid wound infection during the repair process, and that can be suggested as a scaffold for tissue engineering applications and an important constituent for pharmacological activities.

Corrigendum: Reduction in residual cyantraniliprole levels in spinach after various washing and blanching methods.

[This corrects the article DOI: 10.3389/fnut.2022.948671.].

Endophytic Colletotrichum fructicola KL19 and Its Derived SeNPs Mitigate Cd-Stress-Associated Damages in Spinacia oleracea L.

The application of nanotechnology in agriculture has received much attention in order to improve crop yield, quality and food safety. In the present study, a Cd-tolerant endophytic fungus Colletotrichum fructicola KL19 was first ever reported to produce SeNPs, and the production conditions were optimized using the Box-Behnken design in the Response Surface Methodology (RSM-BBD), achieving a peak yield of 1.06 mM under optimal conditions of 2.62 g/20 mL biomass, 4.56 mM Na2SeO3, and pH 6.25. Following this, the properties of the biogenic SeNPs were elucidated by using TEM, DLS, and FTIR, in which the 144.8 nm spherical-shaped SeNPs were stabilized by different functional groups with a negative zeta potential of -18.3 mV. Furthermore, strain KL19 and SeNPs (0, 5, 10, 20 and 50 mg/L) were inoculated in the root zone of small-leaf spinach (Spinacia oleracea L.) seedlings grown in the soil with 33.74 mg/kg Cd under controlled conditions for seven weeks. Impressively, compared with Cd stress alone, the strain KL19 and 5 mg/L SeNPs treatments significantly (p < 0.05) exhibited a reduction in Cd contents (0.62 and 0.50 folds) within the aboveground parts of spinach plants and promoted plants' growth by improving the leaf count (0.92 and 1.36 folds), fresh weight (2.94 and 3.46 folds), root dry weight (4.00 and 5.60 folds) and root length (0.14 and 0.51 folds), boosting total chlorophyll synthesis (0.38 and 0.45 folds), enhancing antioxidant enzymes (SOD, POD) activities, and reducing the contents of reactive oxygen species (MDA, H2O2) in small-leaf spinach under Cd stress. Overall, this study revealed that utilizing endophytic fungus C. fructicola or its derived SeNPs could mitigate reactive oxygen species generation by enhancing antioxidant enzyme activity as well as diminish the absorption and accumulation of Cd in small-leaf spinach, promoting plant growth under Cd stress.

First report of Cotton leaf curl Multan virus of Spinach in China.

Cotton leaf curl Multan virus(CLCuMuV; Begomovirus gossypimultanese, family Geminiviridea) is a single-stranded circular DNA virus, with a genome size of about 2.7 kb, CLCuMuV, which is commonly associated with its satellite DNA, Cotton leaf curl Multan betasatellite (CLCuMuB) (Mansoor et al., 2003), is a serious threat in cotton production causing cotton leaf curl disease (CLCuD) (Briddon et al., 2000). The spread of CLCuMuV is closely linked to its insect vector, whitefly (Bemisia tabaci), which is the exclusive vector species for CLCuMuV transmission (Pan et al., 2018). In May 2019, two spinach (Spinacia oleracea L) samples (XJBC01, XJBC02) showing upward curling of the leaf margins, vein thickening, and enation, symptoms were collected in Shihezi City, Xinjiang, China (Fig. 1B). A 570 bp fragment was amplified from the two symptomatic spinach samples using Begomovirus universal primer pair AV494 (5'-GCCYATRTAYAGRAAGCCMAG-3') and COPR (5'-GANGSATGHTRCADGCCAT ATA-3'), Sequences generated from these amplicons shared 99% nucleotide sequence identities with CLCuMuV DNA-A sequences, suggesting CLCuMuV infection in spinach. To our knowledge CLCuMuV has not been reported in spinach previously. The complete sequences of CLCuMuV and CLCuMuB were then sequenced using CLCuMuV-specific primers GD37-F (5'-GGATCCATTGTTAAACGAATTTCC-3') and GD37-R (5'-GGATCCCACATGTTTGAATTTGA-3') (Gu et al., 2015), as well as betasatellite universal primers ß01 (5'-GGTACCACTACGCTACGCAGCAGCC-3') and ß02 (5'-GGTACCTACCCTCCCAGGGGTACAC-3') (Zhou et al.,2003). The full length CLCuMuV DNA-A in spinach spans 2737 nt (GenBank accession number: MW561346), while CLCuMuB in spinach covers 1343 nt (GenBank accession number: MW561347). The 2737 nt full length CLCuMuV DNA-A and the associated 1343 nt CLCuMuB genome sequences generated from spinach samples were deposited in the GenBank with accession numbers MW561346 and MW561347. The MW561346 shared 99.5% sequence identity with CLCuMV GD37 from Hibiscus rosasinensis. Whereas the MW561347 shared 98.4% sequence identity with CLCuMuB GD37ß. Therefore, we used infectious clones of CLCuMuV (GD37) and CLCuMuB (GD37ß), provided by Xueping Zhou (Gu et al., 2015), to inoculate healthy spinach via Agrobacterium. Infected plants showed typical symptoms 14 days post-inoculation, including leaf edge curling, shrinkage, and vein enlargement, which is consistent with symptoms observed in infected spinach plants in the field (Fig. 1C). The expected 570 bp fragments were amplified in the uninoculated upper leaves of spinach showing symptoms, while not detected in the control spinach, indicating that the symptoms on spinach plants were caused by CLCuMuV associated with CLCuMuB. The transmission efficiency of CLCuMuV to spinach was assessed using two whitefly species, MEAM1 and MED, which were fed on h. rosasinensis infected with CLCuMuV. To compare the transmission efficiency between the two species, 14 spinach plants were inoculated with MEAM1, and 11 spinach plants were inoculated with MED. Each spinach plant was inoculated by releasing 10 whiteflies. After 30 days, MEAM1 transmitted CLCuMuV to spinach inducing typical symptoms (Fig. 1D), with a 78.57% (11/14) transmission efficiency. Similarly, MED also transmitted CLCuMuV to spinach but with a lower efficiency of 54.54% (6/11). These results suggested both MEAM1 and MED could transmit CLCuMuV to spinach, with MEAM1 demonstrating higher efficiency than MED. To the best of our knowledge, this study marks the first report of CLCuMuV infecting spinach, indicating an expanded host range for the virus.

Transcriptome-guided selection of stable reference genes for expression analysis in spinach.

Accurate measurement of gene expression levels is vital for advancing plant biology research. This study explores the identification and validation of stable reference genes (RGs) for gene expression analysis in Spinacia oleracea. Leveraging transcriptome data from various developmental stages, we employed rigorous statistical analyses to identify potential RGs. A total of 1196 candidate genes were initially screened based on expression variability, with subsequent refinement using criteria such as low variance and stability. Among 12 commonly used candidate RGs, EF1α and H3 emerged as the most stable across diverse experimental conditions, while GRP and PPR exhibited lower stability. These findings were further validated through qRT-PCR assays and comprehensive statistical analyses, including geNorm, NormFinder, BestKeeper, and RefFinder. Our study underscores the importance of systematic RG selection to ensure accurate normalization in gene expression studies, particularly in the context of S. oleracea developmental stages and physiological processes like flowering. These validated RGs provide a robust foundation for future gene expression analysis in S. oleracea and contribute to the advancement of molecular research in plant biology.

High Bioavailability of Spinach Folate Evaluated by Functional Biomarkers in a Folate Depletion-Repletion Mouse Model.

The bioavailability of natural folates is 50% lower than that of synthetic folic acid (FA); however, it remains unclear whether this value is universally applicable to all foods. Therefore, the present study investigated the bioavailability of folate from spinach using multiple biomarkers in a folate depletion-repletion mouse model. Mice were fed a folate-deficient diet for 4 wk and subsequently divided into three groups: folate-deficient, FA, and spinach folate. The folate repletion group received either FA or spinach folate at 2 mg/kg diet for 9 d. On the 7th day of repletion, half of each group underwent low-dose total body X-ray irradiation to induce chromosomal damage in bone marrow. Folate bioavailability biomarkers included measurements of folate levels in plasma, liver, and bone marrow along with an analysis of plasma homocysteine levels and chromosome damage, both of which are functional biomarkers of body folate. The consumption of a folate-deficient diet led to decreased tissue folate levels, increased plasma homocysteine levels, and chromosomal damage. Repletion with spinach folate restored folate levels in plasma, liver, and bone marrow to 69, 13, and 68%, respectively, of FA levels. Additionally, spinach folate repletion reduced plasma homocysteine levels and chromosome damage to 83% and 93-117%, respectively, of FA levels. Collectively, the present results demonstrated that the bioavailability of spinach folate exceeded 83% of FA, particularly when assessed using functional biomarkers.

Optimizing the green synthesis of antibacterial TiO2 - anatase phase nanoparticles derived from spinach leaf extract.

Titanium dioxide nanoparticles, renowned for their abundance, non-toxicity, and stability, have emerged as indispensable components in various fields such as air purification, healthcare, and industrial processes. Their applications as photocatalysts and antibacterial agents are particularly prominent. The synthesis methods significantly influence the properties and subsequent applications of these nanoparticles. While several techniques exist, the biological approach using plant extracts offers advantages such as simplicity, biocompatibility, and cost-effectiveness. This study focused on the green synthesis of titanium dioxide nanoparticles utilizing spinach leaf extract. Within the scope of this investigation, the green synthesis of titanium dioxide nanoparticles through spinach leaf extract were synthesized and optimized, followed by a comprehensive examination of their morphological, structural, and chemical attributes with UV-visible spectroscopy, FTIR spectroscopy, XRD, FESEM, and EDX. The minimum inhibitory concentration (MIC) against E. coli and S. aureus was determined to evaluate their antibacterial potential. Optimal synthesis conditions were identified at 50 °C, using a 1/30 concentration and 20 ml of spinach leaf extract. Spherical anatase nanoparticles, ranging from 10 to 40 nm, were produced under these conditions. The change in the color of the extract, absorption at 247 nm, change and increase of the peak at 800 - 400 wavelengths, and the maximum intensity of X-ray diffraction at the angle of 25.367 with the crystal plane 101 were indications of the synthesis of these nanoparticles. Notably, the synthesized nanoparticles exhibited antibacterial activity with MIC values of 0.5 mg/ml against E. coli and 2 mg/ml against S. aureus. This research presents a novel, eco-friendly approach to synthesizing titanium dioxide nanoparticles with promising antibacterial properties.

Developing a plant microbial fuel cell by planting water spinach in a hanging-submerged plant pot system.

To plant crops (especially dry crops such as water spinach) with concomitant electricity recovery, a hanging-submerged-plant-pot system (HSPP) is developed. The HSPP consists of a soil pot (anodic) partially submerged under the water surface of a cathode tank. The microbial communities changed with conditions were also investigated. It was found that with chemical fertilizers the closed-circuit voltage (CCV, with 1 kΩ) was stable (approximately 250 mV) within 28 d; however, without fertilizer, the water spinach could adjust to the environment to obtain a better power output (approximately 3 mW m-2) at day 28. The microbial-community analyses revealed that the Pseudomonas sp. was the only exoeletrogens found in the anode pots. Using a secondary design of HSPP, for a better water-level adjustment, the maximum power output of each plant was found to be approximately 27.1 mW m-2. During operation, high temperature resulted in low oxygen solubility, and low CCV as well. At this time, it is yet to be concluded whether the submerged water level significantly affects electricity generation.

Enhanced aqueous phosphorus removal and mechanism by water spinach (Ipomoea aquatica Forsk) pretreated with lanthanum nitrate.

Excess nutrients such as phosphate (PO43-) entering surface waters promote eutrophication, and phosphorous (P) removal is important to clear the water. Phytoremediation efforts have been used to improve water quality by varieties of P removal plants, such as water spinach (Ipomoea aquatica Forsk). Water spinach can reduce both internal and external resources of phosphorus from waterbody. The ion of lanthanum (La), one rare earth element (REE), is an immobilization substance for aqueous phosphate and also a fertilizer for plants. Therefore, lanthanum nitrate La (NO3)3 was used further to improve the phytoextraction of P from the polluted water. This study investigated the effects of La on the aqueous P removal by two genotypes of water spinach, green stem large leaves (GSLL) and green stem willow leaves (GSWL). The low concentration La (NO3)3 helped the plant to remove more phosphorous from eutrophic water, but La at high concentration lowered the removal of P. Under La (NO3)3 treatments, the optimum concentration for maximum P removal in GSLL is 3 mg/L, and for GSWL, it is 10 mg/L and P removal rates were enhanced to 95% and 96%, respectively. When the concentration of La (NO3)3 is 100 mg/L, the removal percentage of P was only 10% for both genotypes. The very high concentration of La will impose toxicity and even cause the death of the water spinach and produce secondary pollution; for example, under some specific circumstances, the bond between lanthanum and nitrates dissociates into lanthanum ions (La3⁺) and nitrate ions (NO3⁻). If the concentration is high, then it accumulates in the aquatic water organisms and plants and causes toxicity in their bodies. If humans eat up these plants and fish, it causes toxic effects in humans. The La (NO3)3 positively affects different parameters of plants. La (NO3)3 increases the growth, pigments, enzyme activity, and malondialdehyde (MDA) of plants which were also discussed in this study. The biological mechanism should be responsible for the enhanced aqueous phosphorus removal by water spinach using lanthanum nitrate.

Application of EFSA EU menu database and R computing language to calculate the green chlorophyll intake in the European population.

The growing evidence of the health benefits of chlorophyll pigments and the claims that could arise from industry and academia require data on their common dietary intakes. This study presents data on the chronic intake of green chlorophyll in 23 European countries using standardised methodologies to manage food consumption data within the EU Menu methodology. A mean intake of 207.12 mg of green chlorophylls/(d × person) for the adult population was calculated, considering significant covariates. The hierarchical cluster and partial least squares discriminant analysis (PLS-DA) techniques were applied to analyse intake disparities by region and age groups, identifying common food sources of green chlorophylls, such as olive oil, kale, and spinach. This paper presents a modern mathematical approach for obtaining novel information from existing databases of food composition data. Future challenges include building a comprehensive chlorophyll composition database for foods and extending the estimation to non-green chlorophyll pigments and metallo-chlorophyll food colourants.

Characterization of Stemphylium species associated with Stemphylium leaf spot of spinach (Spinacia oleracea).

Stemphylium leaf spot can result in significant losses to spinach seed, processing, and fresh market crops. Stemphylium isolates (n = 1,775) collected from 2000 to 2022 from spinach seed, leaves, and seed crop stem residues were used to assess the diversity of species associated with spinach. Eleven Stemphylium species were identified based on cmdA sequences: S. vesicarium (63.6% of isolates), S. beticola (48.9%), S. amaranthi (5.1%), S. eturmiunum (4.5%), S. astragali (4.0%), S. simmonsii (3.4%), and S. lucomagnoense, S. drummondii, S. gracilariae, S. lycopersici, and S. chrysanthemicola (each 0.6 to 1.7%). Only isolates of S. beticola, S. drummondii, and S. vesicarium were pathogenic to spinach. The incidence of spinach seed on which Stemphylium was observed ranged from 2.5 to 73.5% per seed lot, with S. vesicarium and S. beticola predominant. However, only 60.7 and 62.3% of isolates tested for these two species were pathogenic to spinach, respectively. Therefore, the incidence of Stemphylium species on spinach seed may not reflect accurately the risk of a seed lot carrying pathogenic isolates. Fused MAT1-1 and MAT1-2 genes were detected in isolates of S. vesicarium, but only MAT1-1 was detected in S. beticola isolates, which corroborates previous studies that have proposed the two species to be self-fertile. The duration of ascospore dispersal of S. beticola and S. vesicarium from spinach seed crop stem residues in western Washington, the primary region of spinach seed production in the USA, occurred from mid-winter to late spring or early fall, potentially serving as inoculum for the next season's spinach seed crops. Growers should incorporate residues into the soil after harvest to reduce inoculum production of these pathogens on spinach seed crop residues.

Isolation, characterization, and application of a lytic bacteriophage SSP49 to control Staphylococcus aureus contamination on baby spinach leaves.

Staphylococcus aureus, a major foodborne pathogen, is frequently detected in fresh produce. It often causes food poisoning accompanied by abdominal pain, diarrhea, and vomiting. Additionally, the abuse of antibiotics to control S. aureus has resulted in the emergence of antibiotics-resistant bacteria, such as methicillin resistant S. aureus. Therefore, bacteriophage, a natural antimicrobial agent, has been suggested as an alternative to antibiotics. In this study, a lytic phage SSP49 that specifically infects S. aureus was isolated from a sewage sample, and its morphological, biological, and genetic characteristics were determined. We found that phage SSP49 belongs to the Straboviridae family (Caudoviricetes class) and maintained host growth inhibition for 30 h in vitro. In addition, it showed high host specificity and a broad host range against various S. aureus strains. Receptor analysis revealed that phage SSP49 utilized cell wall teichoic acid as a host receptor. Whole genome sequencing revealed that the genome size of SSP49 was 137,283 bp and it contained 191 open reading frames. The genome of phage SSP49 did not contain genes related to lysogen formation, bacterial toxicity, and antibiotic resistance, suggesting its safety in food application. The activity of phage SSP49 was considerably stable under various high temperature and pH conditions. Furthermore, phage SSP49 effectively inhibited S. aureus growth on baby spinach leaves both at 4 °C and 25 °C while maintaining the numbers of active phage during treatments (reductions of 1.2 and 2.1 log CFU/cm2, respectively). Thus, this study demonstrated the potential of phage SSP49 as an alternative natural biocontrol agent against S. aureus contamination in fresh produce.

Genome-wide analysis of MADS-box genes and their expression patterns in unisexual flower development in dioecious spinach.

Evolution of unisexual flowers involves extreme changes in floral development. Spinach is one of the species to discern the formation and evolution of dioecy. MADS-box gene family is involved in regulation of floral organ identity and development and in many other plant developmental processes. However, there is no systematic analysis of MADS-box family genes in spinach. A comprehensive genome-wide analysis and transcriptome profiling of MADS-box genes were undertaken to understand their involvement in unisexual flower development at different stages in spinach. In total, 54 MADS-box genes found to be unevenly located across 6 chromosomes and can be divided into type I and type II genes. Twenty type I MADS-box genes are subdivided into Mα, Mß and Mγ subgroups. While thirty-four type II SoMADSs consist of 3 MIKC*, and 31 MIKCC -type genes including sixteen floral homeotic MADS-box genes that are orthologous to the proposed Arabidopsis ABCDE model of floral organ identity determination, were identified in spinach. Gene structure, motif distribution, physiochemical properties, gene duplication and collinearity analyses for these genes are performed in detail. Promoters of both types of SoMADS genes contain mainly MeJA and ABA response elements. Expression profiling indicated that MIKCc genes exhibited more dynamic and intricate expression patterns compared to M-type genes and the majority of type-II genes AP1, SVP, and SOC1 sub-groups showed female flower-biased expression profiles, suggesting their role in carpel development, while PI showed male-biased expression throughout flower developmental stages, suggesting their role in stamen development. These results provide genomic resources and insights into spinach dioecious flower development and expedite spinach improvement.

Investigating the carcinogenic and non-carcinogenic health hazards of heavy metal ions in Spinacia oleracea grown in agricultural soil treated with biochar and humic acid.

This study addressed the bioaccumulation and human health risk among the consumption of Spinacia oleracea grown in agricultural soil treated with humic acid (189-2310 ppm) and biochars (0.00-5.10%.wt). The biochars came from two local feedstocks of rice-husk (RH) and sugar-beet-pulp (SBP) pyrolyzed at temperatures 300 and 600 °C. Total concentrations of Cu, Cd, and Ni found in both the soil and biomass/biochar exceeded global safety thresholds. The bioaccumulation levels of HMs in spinach leaves varied, with Fe reaching the highest concentration at 765.27 mg kg-1 and Cd having the lowest concentration at 3.31 mg kg-1. Overall, the concentrations of Zn, Cd, Pb, and Ni in spinach leaves exceeded the safety threshold limits, so that its consumption is not recommended. The assessment of hazard quotient (HI) for the HMs indicated potential health hazards for humans (HI > 1) from consuming the edible parts of spinach. The biochar application rates of 4.35%wt and 0.00%.wt resulted in the highest (3.69) and lowest (3.15) HI values, respectively. The cumulative carcinogenic risk (TCR) ranged from 0.0085 to 0.0119, exceeding the cancer risk threshold. Introducing 5.10%wt biomass/biochar resulted in a 36% rise in TCR compared to the control. The utilization of humic acid alongside HMs-polluted biochars results in elevated levels of HMs bioaccumulation exceeding the allowable thresholds in crops (with a maximum increase of 49% at 2000 ppm humic acid in comparison to 189 ppm). Consequently, this raised the HI by 46% and the TCR by 22%. This study demonstrated that the utilization of HMs-polluted biochars could potentially pose supplementary health hazards. Moreover, it is evident that the utilization of HMs-polluted biochars in treating metal-contaminated soil does not effectively stabilize or reduce pollution.

A longitudinal study on the bacterial quality of baby spinach cultivated in Arizona and California.

In the U.S., baby spinach is mostly produced in Arizona (AZ) and California (CA). Characterizing the impact of growing region on the bacterial quality of baby spinach can inform quality management practices in industry. Between December 2021 and December 2022, baby spinach was sampled after harvest and packaging for microbiological testing, including shelf-life testing of packaged samples that were stored at 4°C. Samples were tested to (i) determine bacterial concentration, and (ii) obtain and identify bacterial isolates. Packaged samples from the Salinas, CA, area (n = 13), compared to those from the Yuma, AZ, area (n = 9), had a significantly higher bacterial concentration, on average, by 0.78 log10 CFU/g (P < 0.01, based on aerobic, mesophilic plate count data) or 0.67 log10 CFU/g (P < 0.01, based on psychrotolerant plate count data); the bacterial concentrations of harvest samples from the Yuma and Salinas areas were not significantly different. Our data also support that an increase in preharvest temperature is significantly associated with an increase in the bacterial concentration on harvested and packaged spinach. A Fisher's exact test and linear discriminant analysis (effect size), respectively, demonstrated that (i) the genera of 2,186 bacterial isolates were associated (P < 0.01) with growing region and (ii) Pseudomonas spp. and Exiguobacterium spp. were enriched in spinach from the Yuma and Salinas areas, respectively. Our findings provide preliminary evidence that growing region and preharvest temperature may impact the bacterial quality of spinach and thus could inform more targeted strategies to manage produce quality. IMPORTANCE: In the U.S., most spinach is produced in Arizona (AZ) and California (CA) seasonally; typically, spinach is cultivated in the Yuma, AZ, area during the winter and in the Salinas, CA, area during the summer. As the bacterial quality of baby spinach can influence consumer acceptance of the product, it is important to assess whether the bacterial quality of baby spinach can vary between spinach-growing regions. The findings of this study provide insights that could be used to support region-specific quality management strategies for baby spinach. Our results also highlight the value of further evaluating the impact of growing region and preharvest temperature on the bacterial quality of different produce commodities.

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.