Ketoprofen exposure perturbs nitrogen assimilation and ATP synthesis in rice roots: An integrated metabolome and microbiome analysis.

Ketoprofen, a commonly used non-steroidal anti-inflammatory drug (NSAID), can enter farmland environments via sewage irrigation and manure application and is toxic to plants. However, there have been relatively few studies on the association of ketoprofen with nitrogen (N) assimilation and metabolic responses in plants. Accordingly, the goal of this study was to investigate the effects of ketoprofen on ATP synthesis and N assimilation in rice roots. The results showed that with increasing ketoprofen concentration, root vitality, respiration rate, ATP content, and H+-ATPase activity decreased and plasma membrane permeability increased. The expressions of OSA9, a family III H+-ATPase gene, and OSA6 and OSA10, family IV genes, were upregulated, indicating a response of the roots to ketoprofen. Nitrate, ammonium, and free amino acids content decreased with increased ketoprofen. The levels of enzymes involved in N metabolism, namely nitrate reductase, nitrite reductase, glutamine synthetase, glutamate synthetase, and glutamate dehydrogenase, also decreased under ketoprofen treatment. Principal component analysis revealed that ketoprofen treatment can significantly affect energy synthesis and nitrogen assimilation in rice roots, while these effects can be alleviated by the antioxidant response. Most of the metabolite contents increased, including amino acids, carbohydrates, and secondary metabolites. Key metabolic pathways, namely substance synthesis and energy metabolism, were found to be disrupted. Microbiome analysis showed that community diversity and richness of rice root microorganisms in solution increased with increasing levels of ketoprofen treatment, and the microbial community structure and metabolic pathways significantly changed. The results of this study provides new insights into the response of rice roots to ketoprofen.

Polystyrene microplastic attenuated the toxic effects of florfenicol on rice (Oryza sativa L.) seedlings in hydroponics: From the perspective of oxidative response, phototoxicity and molecular metabolism.

Antibiotics and microplastics (MPs) are two emerging pollutants in agroecosystems, however the effects of co-exposure to antibiotics and MPs remain unclear. The toxicity of florfenicol (FF) and polystyrene microplastics (PS-MPs) on rice seedlings was investigated. FF and PS-MPs caused colloidal agglomeration, which changed the environmental behavior of FF. FF inhibited rice growth and altered antioxidant enzyme (superoxide dismutase, peroxidase, and catalase) activities, leading to membrane lipid peroxidation; impaired photosynthetic systems, decreased photosynthetic pigments (Chlorophyll a, Chlorophyll b, and carotene), chlorophyll precursors (Proto IX, Mg-Proto IX, and Pchlide), photosynthetic and respiratory rates. The key photosynthesis related genes (PsaA, PsaB, PsbA, PsbB, PsbC, and PsbD) were significantly down-regulated. The ultrastructure of mesophyll cells was destroyed with chloroplast swelling, membrane surface blurring, irregular thylakoid lamellar structure, and number of peroxisomes increased. PS-MPs mitigated FF toxicity, and the IBR index values showed that 10 mg∙L-1 PS-MPs were more effective. Metabolomic analysis revealed that the abundance of metabolites and metabolic pathways were altered by FF, was greater than the combined "MPs-FF" contamination. The metabolism of amino acids, sugars, and organic acids were severely interfered. Among these, 15 metabolic pathways were significantly altered, with the most significant effects on phenylalanine metabolism and the citric acid cycle (p < 0.05).

Comparative study of the toxicity mechanisms of quinolone antibiotics on soybean seedlings: Insights from molecular docking and transcriptomic analysis.

The ecological effects of quinolone antibiotics (QNs) on non-target organisms have received widespread attention. The toxicological mechanisms of three common QNs, that is, enrofloxacin, levofloxacin, and ciprofloxacin, on soybean seedlings were investigated in this study. Enrofloxacin and levofloxacin caused significant growth inhibition, ultrastructural alterations, photosynthetic suppression, and stimulation of the antioxidant system, with levofloxacin exhibiting the strongest toxic effects. Ciprofloxacin (<1 mg·L-1) did not have a significant effect on the soybean seedlings. As the concentrations of enrofloxacin and levofloxacin increased, antioxidant enzyme activities, malondialdehyde content, and hydrogen peroxide levels also increased. Meanwhile, the chlorophyll content and chlorophyll fluorescence parameters decreased, indicating that the plants underwent oxidative stress and photosynthesis was suppressed. The cellular ultrastructure was also disrupted, which was manifested by swollen chloroplasts, increased starch granules, disintegration of plastoglobules, and mitochondrial degradation. The molecular docking results suggested that the QNs have an affinity for soybean target protein receptors (4TOP, 2IUJ, and 1FHF), with levofloxacin having the highest binding energy (-4.97, -3.08, -3.8, respectively). Transcriptomic analysis has shown that genes were upregulated under the enrofloxacin and levofloxacin treatments were mainly involved in ribosome metabolism and processes to synthesize oxidative stress-related proteins. Downregulated genes in the levofloxacin treatment were primarily enriched in photosynthesis-related pathways, indicating that levofloxacin significantly inhibited gene expression for photosynthesis. Genes expression level by quantitative real-time PCR analysis was consistent with the transcriptomic results. This study confirmed the toxic effect of QNs on soybean seedlings, and provided new insights into the environmental risks of antibiotics.

Oxidative response of rice (Oryza sativa L.) seedlings to quinolone antibiotics and its correlation with phyllosphere microbes and antibiotic resistance genes.

With the increasing use of veterinary antibiotics, quinolone antibiotics may enter farmland systems after livestock manure has been composted. However, the phytotoxicity mechanism of antibiotics in crops is still unclear. In this study, the oxidative responses of rice (Oryza sativa L.) seedlings to three typical quinolone antibiotics and their underlying mechanisms were investigated. The bioconcentration factor values were 1.47, 0.55, and 0.23 in the levofloxacin, enrofloxacin and norfloxacin treatment, respectively. The inhibitory effects on rice seedlings were in the order of levofloxacin > enrofloxacin > norfloxacin, which may be due to the high uptake of levofloxacin. The H2O2 level, MDA content, and ion leakage rate increased significantly (p < 0.05), and cell plasmolysis was observed, showing that antibiotics can cause membrane lipid peroxidation and damage the cell membrane structure. Antioxidant enzyme activities (superoxide dismutase, catalase, and peroxidase) changed with the antibiotic concentration. Integrated biomarker response analysis showed that levofloxacin caused the greatest oxidative stress in rice seedlings. Transcriptomic analysis identified 5880 differentially expressed genes, and these were annotated as 20 biological functions; the greatest abundances were cellular and metabolic processes, cell part, and membrane part and organelle; SOD and CAT related genes were up-regulated. The richness and diversity of the phyllosphere microbial community decreased significantly (p < 0.05) and the microbiome changed at the phylum and genus levels. The H2O2 level was correlated with changes in phyllosphere microbial communities. The number of antibiotic resistance genes (ARGs) and mobile genetic elements decreased, while their abundance increased. In conclusion, enrofloxacin exposure not only affects the microbial community but may also affect the ARGs carried by microbes. The relative abundance of MGEs and ARGs was significantly positively correlated (R2 = 0.760, p = 0.0148), indicating that MGEs can significantly promote the spread of ARGs.

Metabolomic analysis reveals the impact of ketoprofen on carbon and nitrogen metabolism in rice (Oryza sativa L.) seedling leaves.

Pharmacologically active compounds (PACs) are becoming common pollutants in the natural environment, posing potential risks to crop quality; however, the toxic effects and metabolic changes that they cause in agricultural plants remain unclear. Here, we investigated the effects of ketoprofen on respiration rate, ATP synthesis, carbon and nitrogen metabolism, and metabolomics in rice seedling leaves. The results showed that ketoprofen treatment adversely affected the respiration rate, ATP content, H+-ATPase activity and induced changes in the contents of carbon assimilation products (soluble sugar, reducing sugar, sucrose, and starch) and the activities of key enzymes in carbon metabolism (sucrose synthase (SS), sucrose phosphate synthase (SPS), and sucrose invertase (InV)). The contents of nitrate, ammonium, and free amino acids, and the activities of key enzymes involved in nitrogen metabolism (nitrate reductase (NR), nitrite reductase (NiR), glutamine synthetase (GS), glutamate synthase (GOGAT), and glutamate dehydrogenase (GDH)) were also affected in a concentration-dependent manner. Metabolomics analysis showed that ketoprofen disturbed the type and content of metabolites (amino acids, carbohydrates, and secondary metabolites) to varying degrees and perturbed key metabolic pathways (substance synthesis and energy metabolism), ultimately resulting in the reduction of rice seedling biomass. This study provides important information and a useful reference for the accurate assessment of the environmental risks of PACs.

Metabolomics and proteomics reveal the toxicological mechanisms of florfenicol stress on wheat (Triticum aestivum L.) seedlings.

Although the ecological impacts of antibiotics have received attention worldwide, research on the toxicity of florfenicol is still limited. We conducted a metabolomic and proteomic study on wheat (Triticum aestivum L.) seedlings to reveal the toxicological mechanism of florfenicol. The growth of the wheat seedlings was found to be inhibited by florfenicol. Antioxidant enzyme activities (superoxide dismutase, peroxidase and catalase), malondialdehyde content and membrane permeability increased with increasing florfenicol concentration. The contents of chlorophyll and chlorophyll synthesis precursor substances (Proto IX, Mg-proto IX and Pchlide), photosynthetic and respiration rates, and chlorophyll fluorescence parameters decreased, indicating that photosynthesis was inhibited. The ultrastructure of chloroplasts was destroyed, as evidenced by the blurred membrane surface, irregular grana arrangement, irregular thylakoid lamella structure, and increased plastoglobuli number. Proteome analysis revealed that up-regulated proteins were highly involved in protein refolding, translation, oxidation-reduction, tricarboxylic acid cycle (TCA cycle), reactive oxygen species metabolic process, cellular oxidant detoxification, and response to oxidative stress. The down-regulated proteins were mainly enriched in photosynthesis-related pathways. In the metabolome analysis, the content of most of the metabolites in wheat leaves, such as carbohydrates and amino acids increased significantly (p < 0.05). Combined pathway analysis showed that florfenicol stress stimulated the TCA cycle pathway and downregulated the photosynthesis pathway.