A new parameter for quantitatively characterizing antibiotic hormesis: QSAR construction and joint toxic action judgment.

Antibiotics usually induce the hormetic effects on bacteria, featured by low-dose stimulation and high-dose inhibition, which challenges the central belief in toxicity assessment and environmental risk assessment of antibiotics. However, there are currently no ideal parameters to quantitatively characterize hormesis. In this study, an effective area in hormesis (AH) was developed to quantify the biphasic dose-responses of single antibiotics (sulfonamides (SAs), sulfonamides potentiators (SAPs), and tetracyclines (TCs)) and binary mixtures (SAs-SAPs, SAs-TCs, and SAs-SAs) to the bioluminescence of Aliivibrio fischeri. Using Ebind (the lowest interaction energy between antibiotic and target protein) and Kow (octanol-water partition coefficient) as the structural descriptors, the reliable quantitative structure-activity relationship (QSAR) models were constructed for the AH values of test antibiotics and mixtures. Furthermore, a novel method based on AH was established to judge the joint toxic actions of binary antibiotics, which mainly exhibited synergism. The results also indicated that SAPs (or TCs) contributed more than SAs in the hormetic effects of antibiotic mixtures. This study proposes a new quantitative parameter for characterizing and predicting antibiotic hormesis, and considers hormesis as an integrated whole to reveal the combined effects of antibiotics, which will promote the development of risk evaluation for antibiotics and their mixtures.

Tetracyclines uptake from irrigation water by vegetables: Accumulation and antimicrobial resistance risks.

Wastewater irrigation may introduce antibiotic residues in the soil-plant systems. This study aimed to investigate the uptake of tetracyclines by spinach and collard greens and assess associated ecological and human health risks. Synthetic wastewater spiked with 1 ppm and 10 ppm of oxytetracycline, doxycycline, and tetracycline was used to grow vegetables in a greenhouse pot experiment. The uptake and accumulation of the tetracyclines were low and residual concentrations in the soil were negligible. All the tetracyclines were detected at concentrations ranging from 1.68 to 51.41 µg/g (spinach) and 1.94-30.95 µg/g (collard greens). The accumulation rate was in a dose-response scenario with a bioconcentration factor of 6.34 mL/kg (spinach) and 2.64 mL/kg (collard greens). Oxytetracycline had the highest accumulation in leaves, followed by doxycycline and tetracycline, and the residual concentrations followed the same order. The highest residual concentration was in soils receiving 10 ppm oxytetracycline. Residual concentrations in the soil were lower than accumulated levels and exerted negligible ecological risks. Tetracyclines accumulation in spinach significantly differed between the vegetables demonstrating a subspecies difference in uptake and accumulation. Ecological risk quotient (RQ) and human health risk quotient (HQ) were below thresholds that would exert toxicity and resistance selection impacts. Although RQs and HQs are low (<0.1), this study shows that the vegetables accumulate tetracyclines from irrigation water, posing plausible human health risks to allergic individuals. Similarly, the ecological risks cannot be ignored because the synergistic and antagonistic effects of sublethal concentrations can perturb ecosystem processes.

Tetracyclines in the environment: An overview on the occurrence, fate, toxicity, detection, removal methods, and sludge management.

Tetracyclines (TCs) are a group of broad-spectrum antibiotics having vast human, veterinary, and aquaculture applications. The continuous release of TCs residues into the environment and the inadequate removal through the conventional treatment systems result in its prevalent occurrence in soil, surface water, groundwater, and even in drinking water. As aqueous TCs contamination is the tip of the iceberg, and TCs possess good sorption capacity towards soil, sediments, sludge, and manure, it is insufficient to rely on the sorptive removal in the conventional water treatment plants. The severity of the TCs contamination is evident from the emergence of TCs resistance in a wide variety of microorganisms. This paper reviews the recent research on the TCs occurrence in the environmental matrices, fate in natural systems, toxic effects, and the removal methods. The high performance liquid chromatography (HPLC) determination of TCs in environmental samples and the associated technology developments are analyzed. The benefits and limitations of biochemical and physicochemical removal processes are also discussed. This work draws attention to the inevitability of proper TC sludge management. This paper also gives insight into the limitations of TCs related research and the future scope of research in environmental contamination by TCs residues.

Occurrence, fate, and risk assessment of typical tetracycline antibiotics in the aquatic environment: A review.

Tetracyclines (TCs), used as human and veterinary medicines, are the most widely used antibiotics. More than 75% of TCs are excreted in an active form and released into the environment through human and animal urine and feces, causing adverse effects on the ecological system and human health. Few articles review the environmental occurrence and behaviors of TCs, as well as their risks and toxicities. Here, we comprehensively summarized the recent advances on the following important issues: (1) Environmental occurrence of TCs. TCs are used globally and their occurrence in the aquatic environment has been documented, including surface water, groundwater, drinking water, wastewater, sediment, and sludge. (2) Environmental behaviors of TCs, particularly the fate of TCs in wastewater treatment plants (WWTPs). Most WWTPs cannot effectively remove TCs from wastewater, so alternative methods for efficient removal of TCs need to be developed. The latest degradation methods of TCs are summarized, including adsorption, photocatalytic, photochemical and electrochemical, and biological degradations. (3) Toxicities and possible risks of TCs. The toxicological data of TCs indicate that several TCs are more toxic to algae than fish and daphnia. Risk assessments based on individual compound exposure indicate that the risks arising from the current concentrations of TCs in the aquatic environment cannot be ignored.

Hormetic dose-dependent response about typical antibiotics and their mixtures on plasmid conjugative transfer of Escherichia coli and its relationship with toxic effects on growth.

Bacterial resistance caused by the abuse of antibiotics has attracted worldwide attention. However, there are few studies exploring bacterial resistance under the environmental exposure condition of antibiotics that is featured by low-dose and mixture. In this study, sulfonamides (SAs), sulfonamide potentiators (SAPs) and tetracyclines (TCs) were used to determine the effects of antibiotics on plasmid RP4 conjugative transfer of Escherichia coli (E. coli) under single or combined exposure, and the relationship between the effects of antibiotics on conjugative transfer and growth was investigated. The results show that the effects of single or binary antibiotics on plasmid RP4 conjugative transfer all exhibit a hormetic phenomenon. The linear regression reveals that the concentrations of the three antibiotics promoting conjugative transfer are correlated with the concentrations promoting growth and the physicochemical properties of the compounds. The combined effects of SAs-SAPs and SAs-TCs on plasmid conjugative transfer are mainly synergistic and antagonistic. While SAPs provide more effective concentrations for the promotion of conjugative transfer in SAs-SAPs mixtures, SAs play a more important role in promoting conjugative transfer in SAs-TCs mixtures. Mechanism explanation shows that SAs, SAPs and TCs inhibit bacterial growth by acting on their target proteins DHPS, DHFR and 30S ribosomal subunit, respectively. This study indicates that toxic stress stimulates the occurrence of conjugative transfer and promotes the development of bacterial resistance, which will provide a reference for resistance risk assessment of antibiotic exposure.

Microbial communities responded to tetracyclines and Cu(II) in constructed wetlands microcosms with Myriophyllum aquaticum.

Combined antibiotic and heavy metal pollution has generated considerable concern. Constructed wetlands (CWs) have been shown to efficiently remove pollutants; however, the microbial community responses to combined pollutants remain enigmatic. In this study, seven microcosm CWs were planted with Myriophyllum aquaticum, spiked with tetracyclines (TCs) (300-30,000 µg/L), alone or with Cu(II), to investigate the response of plant-associated microbial communities. TCs and the Cu/TC ratio greatly affected the performance of CWs. Tetracyclines led to higher microbial diversity, evenness and richness, while UniFrac distances and principal coordinate (PCO) and redundancy analyses revealed that the co-presence of TCs and Cu(II) led to variations in bacterial communities. Proteobacteria, Cyanobacteria and Bacteroidetes were the dominant microbial phyla and Cloacibacterium, Hydrogenophaga, Rheinheimera and Denitratisoma accounted for 6.2-21.0% of all genera. Therefore, the co-occurrence of heavy metals should be considered when judging the removal potential of TCs in phytoremediation.

Tetracyclines lead to ammonium accumulation during nitrification process.

The effect of tetracyclines used for swine food-production (tetracycline and oxytetracycline) on enriched nitrifying bacteria cultures over time was investigated in this study. Short-term exposure assays were performed in different concentrations of each antibiotic, using ammonia oxidizing bacteria (AOB) culture and nitrifying bacteria. The results pointed out a higher inhibitory effect of tetracycline on both bacterial communities. The AOB was more sensitive to antibiotic exposure when compared to the nitrifying culture. Although high antibiotic concentrations were applied, the half maximal inhibitory concentration (IC50) was achieved only for the AOB culture exposed to tetracycline at a concentration of 273 mg L-1. Nonetheless, the long-term exposure assay demonstrated a reduction of the tetracycline inhibition effect against AOB. The exposure to 100 mg L-1 of tetracycline (TC) did not show relevant influence over ammonium conversion efficiency; however, at 128 mg L-1 of TC, the efficiency decreased from 94% to 72%. Further investigation revealed that TC reduced the final effluent quality due to the development of a resistance mechanism by AOB culture against this antibiotic. This mechanism involves increasing the excretion of extracellular polymeric substances (EPS) and soluble microbial products (SMP), which probably increases BOD, and reduces ammonia consumption by the bacterial culture.

Deciphering the microbial and genetic responses of anammox biogranules to the single and joint stress of zinc and tetracycline.

The feasibility of using anaerobic ammonium oxidation (anammox) process to treat wastewaters containing antibiotics and heavy metals was evaluated in this study. The nitrogen removal performance and characteristic parameters were monitored during the whole experimental period of 258 d. The single and joint effects of zinc and tetracycline on the microbial community were studied in upflow anaerobic sludge blanket (UASB) reactors. The anammox performance remained at levels comparable with the initial state at the lower inhibitor concentrations (zinc, 0-2.26 mg L-1; tetracycline, 0-0.5 mg L-1). When the concentrations of zinc and tetracycline increased to 3.39 mg L-1 in R1 and 1.0 mg L-1 in R2, an obvious deterioration in performance was observed. Dual inhibitors with a total concentration of ≥3 mg L-1 caused dramatic decreases in the nitrogen removal efficiency of R3. The quantification results showed that the abundances of eight antibiotic resistance genes (ARGs), czcA and intI1 in the experimental reactors generally increased under stress from metals or/and antibiotics, with final values higher than in the control, while the functional gene abundances were lower. Moreover, most genes exhibited significant correlations. Microbial community analysis indicated that Planctomycetes (represented by Candidatus Kuenenia) was inhibited by both zinc and tetracycline, but still held the dominant position. Furthermore, Caldilinea (belonging to Chloroflexi) maintained a higher abundance during the inhibitory period, implying its potential resistance to both inhibitors. These findings suggested that anammox could be inhibited by metals and antibiotics, but it has the potential to remove nitrogen from wastewaters containing both of them within the concentration threshold.

Effects of divalent copper on microbial community, enzymatic activity and functional genes associated with nitrification and denitrification at tetracycline stress.

The effects of divalent copper (Cu(II)) on microbial community, enzymatic activity and functional genes in a sequencing batch reactor (SBR) at tetracycline (TC) stress were investigated. The enzymatic activity and functional genes abundance associated with nitrification and denitrification at a 20 mg L-1 TC stress were higher than those at a mixtures stress of 20 mg L-1 TC and 10 mg L-1 Cu(II), while they were lower than those at a mixtures stress of 20 mg L-1 TC and 40 mg L-1 Cu(II). Compared to lactate dehydrogenase (LDH) release and reactive oxygen species (ROS) production at a 20 mg L-1 TC stress, they were lower at the TC stress with 10 mg L-1 Cu(II), while they were higher at the TC stress with 40 mg L-1 Cu(II). The incremental Cu(II) concentration at a 20 mg L-1 TC stress could not change the result that the sensitivity of denitrifying enzymatic activity to TC was higher than nitrifying enzymatic activity. Compared to the relative abundance of nitrifers and denitrifers at a 20 mg L-1 TC stress, the 10 mg L-1 Cu(II) addition resulted in their increase, while they decreased as the 40 mg L-1 Cu(II) addition. The relative abundance of genera Pseudomonas, Rivibacter and Nitrobacter at the stress of Cu(II) and TC were higher than those at TC stress, suggested they had an ability to resist the stress of Cu(II) and TC.

Toxic effects of tetracycline and its degradation products on freshwater green algae.

Tetracycline antibiotics are the most widely used antibiotics in the world and the most common veterinary drugs and feed additives used in livestock, poultry and aquaculture operations. Because antibiotics cannot be completely removed by currently existing sewage treatment facilities, these materials enter the environment directly via sewage treatment plant discharge, where they degrade. Accordingly, the metabolism and the ecological toxicity of tetracycline degradation products are worthy of attention. Herein, we investigated the effects of tetracycline and its degradation products (anhydrotetracycline and epitetracycline hydrochloride) on the growth, cell structure and algal cell oxidative stress of common Chlorella vulgaris. The results showed that the 96h-EC50 values of tetracycline (TC), anhydrotetracycline (ATC) and epitetracycline (ETC) on algal cells were 7.73, 5.96 and 8.42 mg/L, respectively. Moreover, the permeability of algal cells exposed to high concentrations of these three drugs was significantly enhanced. In addition, there were structural changes in the cells such as plasmolysis and starch granule deposition appeared, the thylakoid lamellae in the chloroplasts became blurred and deformed, and the vacuoles became larger. Exposure to higher concentrations (>5 mg/L) of TC and its degradation products ATC and ETC significantly upregulated the activity of ROS, as well as the antioxidants SOD and CAT. The levels of the lipid peroxidation product MDA also showed the same trend. Finally, ATC had the strongest toxicity toward algal cells, followed by TC and then ETC.

Ca2+ protect zebrafish embryos from water acidification.

Ionizable strategies are routinely used to enhance the solubility and dissolution rates of various pharmaceuticals. These chemicals may directly affect aquatic environment once discharged from factories, hospitals or livestock farms. Here, we assessed the potential side effect of tetracyclines (TCs) on the development of zebrafish embryos. Tetracycline hydrochloride decreased water pH from 6.4 to 4.4 at 30 mg/L. Acidified water exceeded the tolerance of zebrafish embryos in pure water during the early ten hours post fertilization (hpf). Interestingly, we found that Ca2+ in the embryo medium could increase the tolerance of embryos to acidified water. Furthermore, we found that the protection of Ca2+ was not due to the formation of TCs-Ca2+ complexes under acidic condition, based on spectral analysis. Meanwhile we showed that exogenous addition of Ca2+ could inhibit the accumulation of Ca2+ from the cytoplasm to the surface of embryos. These results may shed light on the strategies for protecting aquatic animals from acidic environments.

Phosphine production in anaerobic wastewater treatment under tetracycline antibiotic pressure.

The influence of tetracycline (TC) antibiotics on phosphine (PH3) production in the anaerobic wastewater treatment was studied. A lab-scale anaerobic baffled reactor with three compartments was employed to simulate this process. The reactor was operated in a TC-absence wastewater and 250µg/L TC-presence wastewater for three months after a start-up period, respectively. The responses of pH, oxidation-reduction potential (ORP), chemical oxygen demand (COD), total phosphorus (TP), enzymes activity (dehydrogenase and acid phosphatase), and microbial community were investigated to reveal the effect of TC on PH3 production. Results suggested that the dehydrogenase (DH) activity, acid phosphatase (ACP) activity and COD have positive relationship with PH3 production, while pH, ORP level and the TP in liquid phase have negative relationship with PH3 production. With prolonged TC exposure, decrease in pH and increase in DH activity are beneficial to PH3 production, while decrease in COD and ACP activity are not the limiting factors for PH3 production.

A QSAR-based mechanistic study on the combined toxicity of antibiotics and quorum sensing inhibitors against Escherichia coli.

Quorum sensing inhibitors (QSIs) have attracted increasing attention due to their potential roles as the antibiotic alternatives. The combination of QSIs and antibiotics in clinical use and their subsequent release into the environment may result in joint effects on the ecology and environment, which has not received enough concerns yet. In this study, eight potential QSIs and three types of commonly used antibiotics, i.e., sulfonamides (SAs), ß-lactams and tetracyclines (TCs), were investigated for their combined toxicity on Escherichia coli (E. coli). The QSAR models for the combined toxicity were constructed using the interaction energies between the chemicals and their target proteins as calculated by molecular docking. It was revealed that the SAs and QSIs presented either additive or antagonistic joint effects in the mixture toxicity test, while ß-lactams and TCs showed only antagonistic effects with the QSIs. The analysis on the coefficients in the QSAR models suggested that the QSIs in the mixtures were more involved in the interaction with the proteins than the antibiotics. This study will help better understand the risks of joint exposure to the antibiotics and QSIs, and provide a new perspective for the study of the combined toxicity mechanism.

Joint toxicity of fluoroquinolone and tetracycline antibiotics to zebrafish (Danio rerio) based on biochemical biomarkers and histopathological observation.

Herein, we report on the joint toxicity of four fluoroquinolones and two tetracyclines (ß-diketone antibiotics-DKAs) to zebrafish based on a series of toxicological endpoints and histopathological observations. A positive dose-dependence was observed in DKA-exposure groups with a 72-hpf EC50 of 130.3 mg/L for hatching rate, 120-hpf LC50 of 149.8 mg/L, and 120-hpf EC50 of 135.1 mg/L for malformation rate. When zebrafish at 60 dpf were exposed to a series of DKA concentrations (45, 60 and 90 mg/L) for 7, 14 and 21 days, creatine kinase and AChE activities were significantly induced, and intracellular malondialdehyde increased in all treatments except for the 45 mg/L treatment. The transcription levels of AHRRa from livers were significantly (p < 0.05) up-regulated in all treatments after two months of DKA exposure. CKma expression from skeletal muscle was significantly down-regulated in the 90 mg/L treatment. A remarkable down-regulation of CYP3A65 was observed in the 60 mg/L treatment. DKA exposure resulted in severe tissue damage including mitochondria swelling, reduction of mitochondrial cristae, deepening of mitochondrial cristae bands, and decreasing and even disappearance of the rough endoplasmic reticulum. Total sperm motility was decreased by ca. 30% due to DKA exposure. These results provide important information for toxicity and health risks due to mixed DKA exposure in aquatic environments.

Reproductive toxicity of ß-diketone antibiotic mixtures to zebrafish (Danio rerio).

So far, few data are available on the reproductive toxicological assessment of ß-diketone antibiotics (DKAs), a class of ubiquitous pseudo-persistent pollutant, in zebrafish (Danio rerio). Herein, we reported the reproductive effects of DKAs by means of transcriptome analysis (F1-zebrafish), changes in a series of reproductive indices (F0-zebrafish) and histopathological observations. A total of 1170, 983 and 1399 genes were found to be differentially expressed when compared control vs. 6.25mg/L, control vs. 12.5mg/L and 6.25 vs. 12.5mg/L DKA-exposure treatments, respectively. Among three comparison groups, 670, 569 and 821 genes were respectively assigned for GO analyses based on matches with sequences of known functions. In 149 KEGG-noted metabolic pathways, the preferential one was the MAPK (mitogen-activated protein kinase) signaling pathway, followed by oxidative phosphorylation, neuroactive ligand-receptor interaction and so on. By qPCR verification, 6 genes (c6ast4, igfbp1b, mrpl42, tnnc2, emc4 and ddit4) showed consistent gene expression with those identified by transcriptome sequencing. Due to DKA-exposure, the concentrations of plasma estradiol and testosterone, and the gonado-somatic index were significantly dose-dependently declined. Also, DKA-exposure led to declining in zebrafish reproductive capacity, reflecting in fertilization, hatchability and egg production. Histopathological observations demonstrated that zebrafish ovary and testis suffered serious damage after DKA-exposure. The 4-oxo-TEMP signals increased obviously with increasing DKA-exposed concentrations, implying disruption of balance between generation and clearance of 1O2. In summary, DKAs not only produce reproductive toxicological effects on F0-zebrafish, but also result in adverse consequences for growth and development of F1-zebrafish.

Toxin Release of Cyanobacterium Microcystis aeruginosa after Exposure to Typical Tetracycline Antibiotic Contaminants.

The global usage of veterinary antibiotics is significant. Antibiotics can be released into aquatic environments and elicit toxic effects on non-target organisms. In this study, the growth characteristics and toxin release of the cyanobacterium Microcystis aeruginosa (M. aeruginosa) were examined to investigate the physiological effects of tetracycline antibiotics on aquatic life. Results showed that the degree of toxicities of the following target antibiotics was TC (tetracycline hydrochloride) > CTC (chlortetracycline hydrochloride) > OTC (oxytetracycline hydrochloride) in terms of growth parameters, EC10 (0.63, 1.86, and 3.02 mg/L, respectively), and EC20 (1.58, 4.09, and 4.86 mg/L, respectively) values. These antibiotics inhibited the production of microcystin-LR (MC-LR) to varying degrees. CTC interfered M. aeruginosa cells and decreased their ability to release MC-LR, but this antibiotic stimulated the ability of these cells to synthesize MC-LR at 2 and 5 mg/L. OTC elicited a relatively weaker toxicity than CTC did and reduced MC-LR release. TC was the most toxic among the three antibiotics, and this antibiotic simultaneously reduced intracellular and extracellular MC-LR equivalents. Our results helped elucidate the effects of tetracycline antibiotics on M. aeruginosa, which is essential for environmental evaluation and protection. Our results are also helpful for guiding the application of veterinary antibiotics in agricultural settings.

Screening and functional identification of lncRNAs under ß-diketone antibiotic exposure to zebrafish (Danio rerio) using high-throughput sequencing.

Long non-coding RNAs (lncRNAs) have attracted considerable research interest, but so far no data are available on the roles of lncRNAs and their target genes under chronic ß-diketone antibiotic (DKAs) exposure to zebrafish (Danio rerio). Herein, we identified 1.66, 3.07 and 3.36×104 unique lncRNAs from the 0, 6.25 and 12.5mg/L DKA treatment groups, respectively. In comparison with the control group, the 6.25 and 12.5mg/L treatments led to up-regulation of 2064 and 2479 lncRNAs, and down-regulation of 778 and 954 lncRNAs, respectively. Of these, 44 and 39 lncRNAs in the respective 6.25 and 12.5mg/L treatments displayed significant differential expression. Volcano and Venn diagrams of the differentially expressed lncRNAs were constructed on the basis of the differentially expressed lncRNAs. After analyzing 10 lncRNAs and potential target genes, a complex interaction network was constructed between them. The consistency of 7 target genes (tenm3, smarcc1b, myo9ab, ubr4, hoxb3a, mycbp2 and CR388046.3), co-regulated by 3 lncRNAs (TCONS_00129029, TCONS_00027240 and TCONS_00017790), was observed between their qRT-PCR and transcriptomic sequencing. By in situ hybridization (ISH), abnormal expression of 3 lncRNAs was observed in hepatic and spleen tissues, suggesting that they might be target organs for DKAs. A similar abnormal expression of two immune-related target genes (plk3 and syt10), co-regulated by the 3 identified lncRNAs, was observed in liver and spleen by ISH. Histopathological observations demonstrated hepatic parenchyma vacuolar degeneration and clot formation in hepatic tissues, and uneven distribution of brown metachromatic granules and larger nucleus in spleen tissues resulting from DKA exposure. Overall, DKA exposure led to abnormal expression of some lncRNAs and their potential target genes, and these genes might play a role in immune functions of zebrafish.

Both cholestatic and steatotic drugs trigger extensive alterations in the mRNA level of biliary transporters in rat hepatocytes: Application to develop new predictive biomarkers for early drug development.

Disruption of the vectorial bile acid transport in the liver is a key feature of cholestatic drugs, although many causal and mechanistic aspects are still unknown. The aim of the present study was to explore if cholestatic drugs can repress or induce the expression of hepatic transporters. To this end, sandwich-cultured rat hepatocytes were treated with cholestatic and non-cholestatic (steatotic, non-hepatotoxic, etc.) drugs and the mRNA expression of 10 uptake and efflux biliary transporters was measured. Results evidenced that all cholestatic drugs cause extensive alterations in the mRNA expression of most biliary transporters. Surprisingly, nearly all steatotic drugs also affected the expression of these genes. The most frequent alterations triggered by both types of drugs were the repression of OATP1A1, NTCP and BSEP, and the induction of MRP2/3/4, MDR2 and ABCG5/8. The majority of these alterations were also observed in vivo, in the livers of treated rats. The common signature of cholestatic and steatotic drugs was the repression of OATP1A1. Indeed, ROC curve analysis indicated that OATP1A1 mRNA is a very sensitive marker to identify drugs with cholestatic or steatotic potential, with a maximal sensitivity and specificity of 0.917 and 0.941, respectively. We conclude that alteration of expression of hepatobiliary transporters is a hallmark of both cholestatic and steatotic drugs, lending support to a connection between these two mechanisms of hepatotoxicity. Moreover, OATP1A1 mRNA is proposed as a very simple and useful screening biomarker for the prediction of new cholestatic or steatotic drugs in early drug development.

Content of biogenic amines in Lemna minor (common duckweed) growing in medium contaminated with tetracycline.

Aquatic plants are continuously exposed to a variety of stress factors. No data on the impact of antibiotics on the biogenic amines in duckweed (Lemna minor) have been available so far, and such data could be significant, considering the ecological role of this plant in animal food chains. In the tissues of control (non-stressed) nine-day-old duckweed, the following biogenic amines were identified: tyramine, putrescine, cadaverine, spermidine and spermine. Based on the tetracycline contents and the computed EC values, the predicted toxicity units have been calculated. The obtained results demonstrated phytoxicity caused by tetracycline in relation to duckweed growth rate, yield and the contents of chlorophylls a and b. The carotenoid content was not modified by tetracycline. It was found that tetracycline as a water pollutant was a stress factor triggering an increase in the synthesis of amines. Tetracycline at 19, 39 and 78µM concentrations increased biogenic amine synthesis by 3.5 times. Although the content of tyramine increased fourteen times with the highest concentration of the drug (and of spermidine - only three-fold) the increase of spermidine was numerically the highest. Among the biogenic amines the most responsive to tetracycline were spermine and tyramine, while the least affected were putrescine and spermidine. Despite putrescine and spermidine being the least sensitive, their sum of contents increased five-fold compared to the control. These studies suggest that tetracycline in water reservoirs is taken up by L. minor as the antibiotic clearly modifies the metabolism of this plant and it may likely pose a risk.

Effects of ß-diketone antibiotics on F1-zebrafish (Danio rerio) based on high throughput miRNA sequencing under exposure to parents.

The toxicity of ß-diketone antibiotics (DKAs), a class of ''pseudo-persistent'' environmental pollutants, to F0-zebrafish (Danio rerio) was investigated using 7-dpf F1-zebrafish miRNA sequencing and bioinformatics analyses. Based on relative expression, 47, 134 and 118 of 193 mature miRNAs were differentially expressed between control vs 6.25 mg/L, control vs 12.5 mg/L and 6.25 vs 12.5 mg/L treatments, respectively. Utilizing three databases, 2523 potential target genes were predicted, and they were assigned to 19 high-abundance KEGG pathways and 20 functional categories by COG analysis. Among 11 significantly differential expression and high-abundance miRNAs, the expression levels for 7 miRNAs (miR-144, -124, -499, -125b, -430b, -430c and -152) assessed by qRT-PCR were consistent with those determined by sRNA-seq. A potential network was plotted between 11 miRNAs and their target genes based on differential expression and binding effectiveness. The high degree of connectivity between miRNA-gene pairs suggests that these miRNAs play critical roles in zebrafish development. The expression of miR-124 and miR-499 in whole-mount in situ hybridization was in general agreement with those from qRT-PCR and miRNA-seq and were DKA concentration-dependent. DKA exposure induced severe histopathological changes and damage in F0-zebrafish ovary tissue, as reflected by an increased number of early developmental oocytes, irregular cell distribution, decreased yolk granules, cytoplasmic shrinkage, cell lysis in mature oocytes, and dissolution of internal corona radiata. Chronic DKA exposure affected reproduction of F0-zebrafish and development of F1-zebrafish. These observations demonstrate the toxic effect transfer relation across parent and their offspring, and enhance our understanding of drug-induced diseases.