Ecotoxicology Mechanism And Resistance Omics Of Quinolone Antibiotics On Arabidopsis Thaliana And Enchytraeus Crypticus

Antibiotics are widely used in modern disease treatment as a drug that kills microorganisms or inhibits their growth through biochemical action and as a feed additive for disease prevention or growth promotion in livestock industry.However,due to the low metabolic rate in organism,antibiotics are excreted to environment by excrement,causing harm to the ecological environment.In this paper,three typical quinolone antibiotics（enrofloxacin,norfloxacin,levofloxacin）were selected.And the effect on basic physiology,photosystem,oxidative stress system,phyllospheric microbes to model plant Arabidopsis thaliana;the effect on basic physiology,defend response system,gut microbes,antibiotics resistance genes to soil non-target biolumella Enchytraeus crypticus were studied to to reveal the toxicological effects on plant-soil system.Under the pressure of three quinolone antibiotics,the fresh weight and root length of A.thaliana decreased with higher concentration,showing a significant dose effect and the the toxicity is:enrofloxacin>norfloxacin>levofloxacin.Under the pressure of antibiotics,the growing leaves with the increasing of antibiotic concentration,growing leaves of A.thaliana appeared albino and increased in higher antibiotic concentration.The cell permeability increased with antibiotic concentration.In 50 mg/L enrofloxacin treatment,the leaf of A.thaliana was fuzzy,organelles were swollen and starch granules were increased.Under the pressure of three quinolone antibiotics,fluorescence parametersthe（F₀,F_m,F_v/F_m）and light curve（Y（II）and NPQ）were detected.F₀ and F_m values increased with higher concentration indicating the damage to PSII reaction centers.F_v/F_m,Y（II）values decreased with higher antibiotics concentration indicating the inhibition of photosynthetic efficiency.NPQ values decreased with higher antibiotics indicating an inhibition of dissipation capacity of excess light energy,whereas,higher F_v/F_m,Y（II）and NPQ values in growing leaves indicating an adaptation in A.thaliana to ensure better growth.The inhibitory effect of the three antibiotics on the photosystem is:enrofloxacin>norfloxacin>levofloxacin.Under the pressure of three quinolone antibiotics,reactive oxygen species（ROS）level got higher and caused intracellular lipid peroxidation leading a higher content of malondialdehyde in A.thaliana.In high concentration treatment groups,SOD and GPX activity were decreassed might due to the external pressure was too high.The POD activity continues to rise,might due to hydrogen peroxide（H₂O₂）concentration in A.thaliana.The superoxide response genes（At COBL5,At ERD7,At GRP,At UCP2 and At OGO）were up-regulated,corresponding to the increased tolerance to oxidative stress in A.thaliana.Under the pressure of enrofloxacin,Proteobacteria,Chlamydiae,Cyanobacteria,Bacteroidetes,Firmicutes and Actinobacteria were detected in phyllospheric microbes of A.thaliana.The decrease of OUT numbers,Chao and Shannon index indicating the decrease of abundance and diversity in phyllospheric microbes.Peroxisome,glutathione metabolism were up-regulated indicating that the host’s antioxidant capacity was enhanced.The drug metabolism-cytochrome P450,metabolism of xenobiotics by cytochrome P450 were up-regulated indicating that the metabolism and detoxification of antibiotics was enhanced.Plant-pathogen interaction,starch and sucrose metabolism and carbon fixation pathways were down-regulated might leading the inhibition of growth and the incease of disease risk.Under the pressure of three quinolone antibiotics,the weight and larvae number of E.crypticus decreased with higher antibiotics concentration,showing a significant dose effect.The significant changes of larvae number just appeared in high concentration treatment groups.The toxic effect shows that the toxicity is:enrofloxacin>norfloxacin>levofloxacin.The ROS concentration increased in higher treatment group.SOD and glutathione S transferase（GST）activity changed indicating the antioxidant stress mechanism in E.crypticus.was stimulated.As a exogenous pollutant detoxification enzyme,cytochrome P450（CYP450）activity increased with higher antibiotic concentration to detoxify external pollutants.Under the pressure of three quinolone antibiotics,as the recognition factor of pathogens in the immune system,the relative gene expression abundance of coelomic cytolytic factor increased showing a significant difference in enrofloxacin treatment.The relative gene expression abundance of antibacterial protein with hemolytic fuction in cells increased showing a significant difference in norfloxacin and enrofloxacin treatment.The relative gene expression abundance of Lysozyme was increased.The response of immune system in E.crypticus to three quinolone antibiotics is:enrofloxacin>norfloxacin>levofloxacin.Under the pressure of enrofloxacin,the gut microbes in E.crypticus at phylum level had not changed and Proreobacteria,Actinobacteria,Bacteroidetes,Firmicutes,Planctomycetota were detected.The decrease of OUT numbers,Chao and Shannon index indicating the decrease of abundance and diversity in phyllospheric microbes.Peroxisome and?-alanine metabolism were up-regulated indicating that antioxidant capacity of E.crypticus was enhanced.Butyrate metabolism,tryptophan metabolism were up-regulated indicating the immune protection was enhanced.Oxidative phosphorylation was down-regulated indicating that chondrial energy metabolism was inhibited.Pyrimidine metabolism,DNA replication,ribosome and homologous recombination were down-regulated indicating that the genome health of E.crypticus was threatened.Lipopolysaccharide biosynthesis was down-regulated indicating that the cell membrane integrity of gut microbial was threatened.The relative abundance of Bacillus,Acinetobacter,Burkholderia,Rhodococcus were increased indicating that the infection risk of E.crypticus was increased.The relative abundance of Sphingomonas was decreased whereas increased in Streptomyces,Nocardioides indicating that antibiotics production in E.crypticus was increased to eliminate pathogenic bacteria.Under the pressure of enrofloxacin,ARGs and MGEs were significantly enriched in the intestinal microbes of E.crypticus.The relative abundances of qnr D ARG subtypes increased by up to 11.72 times indicating enrofloxacin exerted exogenous pressure on the evolution of E.crypticus.The analysis of AGRs/MGEs and gut microbial correlation showed that multiple ARGs are positively correlated,which might be the cause of co-resistance in E.crypticus;three pathogenic bacteria（Bacillus,Rhodococcus,Escherichia）were positively correlated with ARGs/MGEs,might be the reason for the increase of relative abundance;Sphingomonas were negatively correlated with ARGs/MGEs,indicating that the ability of E.crypticus to degrade antibiotics was inhibited to ensure kill the increasing of pathogenic bacteria.