Metagenomic and transcriptomic analysis revealing the impact of oxytetracycline and ciprofloxacin on gut microbiota and gene expression in the Chinese giant salamander (Andrias davidianus).

Excessive antibiotic use has led to the spread of antibiotic resistance genes (ARGs), impacting gut microbiota and host health. However, the effects of antibiotics on amphibian populations remain unclear. We investigated the impact of oxytetracycline (OTC) and ciprofloxacin (CIP) on Chinese giant salamanders (Andrias davidianus), focusing on gut microbiota, ARGs, and gene expression by performing metagenome and transcriptome sequencing. A. davidianus were given OTC (20 or 40 mg/kg) or CIP (50 or 100 mg/kg) orally for 7 days. The results revealed that oral administration of OTC and CIP led to distinct changes in microbial composition and functional potential, with CIP treatment having a greater impact than OTC. Antibiotic treatment also influenced the abundance of ARGs, with an increase in fluoroquinolone and multi-drug resistance genes observed post-treatment. The construction of metagenome-assembled genomes (MAGs) accurately validated that CIP intervention enriched fish-associated potential pathogens Aeromonas hydrophila carrying an increased number of ARGs. Additionally, mobile genetic elements (MGEs), such as phages and plasmids, were implicated in the dissemination of ARGs. Transcriptomic analysis of the gut revealed significant alterations in gene expression, particularly in immune-related pathways, with differential effects observed between OTC and CIP treatments. Integration of metagenomic and transcriptomic data highlighted potential correlations between gut gene expression and microbial composition, suggesting complex interactions between the host gut and its gut microbiota in response to antibiotic exposure. These findings underscore the importance of understanding the impact of antibiotic intervention on the gut microbiome and host health in amphibians, particularly in the context of antibiotic resistance and immune function.

Biofilm enhances the interactive effects of microplastics and oxytetracycline on zebrafish intestine.

The enhanced adsorption of pollutants on biofilm-developed microplastics has been proved in many studies, but the ecotoxicological effects of biofilm-developed microplastics on organisms are still unclear. In this study, adult zebrafish were exposed to original microplastics, biofilm-developed microplastics, original microplastics absorbed with oxytetracycline (OTC), and biofilm-developed microplastics absorbed with OTC for 30 days. The intestinal histological damage, intestinal biomarker response, gut microbiome and antibiotic resistance genes (ARGs) profile of zebrafish were measured to explore the roles of biofilm in the effects of microplastics. The results showed that biofilm-developed microplastics significantly increased the number of goblet cells in intestinal epithelium compared with the control group. The biofilm-developed microplastics also induced the oxidative response in the zebrafish intestines, and biofilm changed the response mode in the combined treatment with OTC. Additionally, the biofilm-developed microplastics caused intestinal microbiome dysbiosis, and induced the abundance of some pathogenic genera increasing by several times compared with the control group and the original microplastics treatments, regardless of OTC adsorption. Furthermore, the abundance of ARGs in biofilm-developed microplastics increased significantly compared with the control and the original microplastic treatments. This study emphasized the significant influence and unique role of biofilm in microplastic studies.

Effect of microplastics on oxytetracycline trophic transfer: Immune, gut microbiota and antibiotic resistance gene responses.

Microplastics and antibiotics are prevalent and emerging pollutants in aquatic ecosystems, but their interactions in aquatic food chains remain largely unexplored. This study investigated the impact of polypropylene microplastics (PP-MPs) on oxytetracycline (OTC) trophic transfer from the shrimp (Neocaridina denticulate) to crucian carp (Carassius auratus) by metagenomic sequencing. The carrier effects of PP-MPs promoted OTC bioaccumulation and trophic transfer, which exacerbated enterocyte vacuolation and hepatocyte eosinophilic necrosis. PP-MPs enhanced the inhibitory effect of OTC on intestinal lysozyme activities and complement C3 levels in shrimp and fish, and hepatic immunoglobulin M levels in fish (p < 0.05). Co-exposure of MPs and OTC markedly increased the abundance of Actinobacteria in shrimp and Firmicutes in fish, which caused disturbances in carbohydrate, amino acid, and energy metabolism. Moreover, OTC exacerbated the enrichment of antibiotic resistance genes (ARGs) in aquatic animals, and PP-MPs significantly increased the diversity and abundance of ARGs and facilitated the trophic transfer of teta and tetm. Our findings disclosed the impacts of PP-MPs on the mechanism of antibiotic toxicity in aquatic food chains and emphasized the importance of gut microbiota for ARGs trophic transfer, which contributed to a deeper understanding of potential risks posed by complex pollutants on aquatic ecosystems.

Combined effects of azoxystrobin and oxytetracycline on rhizosphere microbiota of Arabidopsis thaliana.

The rhizosphere is one of the key determinants of plant health and productivity. Mixtures of pesticides are commonly used in intensified agriculture. However, the combined mechanisms underlying their impacts on soil microbiota remain unknown. The present study revealed that the rhizosphere microbiota was more sensitive to azoxystrobin and oxytetracycline, two commonly used pesticides, than was the microbiota present in bulk soil. Moreover, the rhizosphere microbiota enhanced network complexity and stability and increased carbohydrate metabolism and xenobiotic biodegradation as well as the expression of metabolic genes involved in defence against pesticide stress. Co-exposure to azoxystrobin and oxytetracycline had antagonistic effects on Arabidopsis thaliana growth and soil microbial variation by recruiting organic-degrading bacteria and regulating ABC transporters to reduce pesticide uptake. Our study explored the composition and function of soil microorganisms through amplicon sequencing and metagenomic approaches, providing comprehensive insights into the synergistic effect of plants and rhizosphere microbiota on pesticides and contributing to our understanding of the ecological risks associated with pesticide use.

Differential effects of oxytetracycline on detoxification and antioxidant defense in the hepatopancreas and intestine of Chinese mitten crab under cadmium stress.

This study aims to evaluate the effects of oxytetracycline (OTC) on detoxification and oxidative defense in the hepatopancreas and intestine of Chinese mitten crab (Eriocheir sinensis) under cadmium (Cd) stress. The crab was exposed to 0.6 µM Cd, 0.6 µM OTC, and 0.6 µM Cd plus 0.6 µM OTC for 42 days. Our results showed that in the intestine, OTC alone enhanced protein carboxylation (PC) and malondialdehyde (MDA) contents, which was associated with the increased OTC accumulation. Compared to Cd alone, Cd plus OTC increased Cd and OTC contents, and reduced detoxification (i.e., glutathione (GSH) content, gene expressions of cytochrome P450 (CYP) isoforms, 7-ethoxyresorufin O-deethylase (EROD) activity, mRNA levels and activities of glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione-S-transferase (GST)), and antioxidant defense (i.e., gene expressions and activities of catalase (CAT) and superoxide dismutase (SOD)) in the intestine, leading to the increased in PC and MDA contents, suggesting that OTC had a synergistic effect on Cd-induced oxidative damage. In the hepatopancreas, although OTC alone increased OTC accumulation, it did not affect PC and MDA contents. Compared to Cd alone, Cd plus OTC reduced MDA content, which was closely related to the improvement of detoxification (i.e., GSH content, mRNA levels of CYP isoforms, EROD activity, gene expressions and activities of GPx, GR and GST), and antioxidant defense (gene expressions and activities of CAT and SOD, metallothionein content). Aryl hydrocarbon receptor (AhR) and nuclear factor E2-related factor 2 (Nrf2) transcriptional expressions were positively correlated with most detoxification- and antioxidant-related gene expressions, respectively, indicating that AhR and Nrf2 were involved in the regulation of these gene expressions. Our results unambiguously demonstrated that OTC had tissue-specific effects on Cd-induced toxicological effect in E. sinensis, which contributed to accurately evaluating Cd toxicity modulated by TCs in crab.

Z-scheme heterojunction photocatalyst formed by MOF-derived C-TiO2 and Bi2WO6 for enhancing degradation of oxytetracycline: Mechanistic insights and toxicity evaluation in the presence of a single active species.

In the work, Bi2WO6/C-TiO2 photocatalyst was successfully synthesized for the first time by loading narrow bandgap semiconductor Bi2WO6 on MOF-derived carboxyl modified TiO2. The phase structure, morphology, photoelectric properties, surface chemical states and photocatalytic performance of the prepared photocatalysts were systematically investigated using various characterization tools. The degradation efficiency of oxytetracycline by 6BT Z-scheme heterojunction photocatalyst under visible light could reach 93.6 % within 100 min, which was related to the high light harvesting and effective separation and transfer of photo-generated carriers. Furthermore, the effects of various environmental factors in actual wastewater were further investigated, and the results showed that 6BT exhibited good adaptability, durability and resistance to interference. Unlike most works, the degradation system with a different single active species were designed and constructed based on their formation mechanism. In addition, for the first time, a positive study was conducted on the priority attack sites, intermediate products, and degradation pathways for the photocatalytic degradation of oxytetracycline by a single active species through HPLC-MS and Fukui index calculations. The toxicity changes of intermediate products produced in three different single active species oxidation systems were evaluated using toxicity assessment software tools (T.E.S.T.), Escherichia coli growth experiments, and wheat growth experiments. Among them, the intermediate products formed through O2- oxidation had the lowest toxicity and the main active sites it attacked were the 20C, 38O, 18C, 41O, and 55O atoms with high f+ values in the oxytetracycline molecular structure. This work provided the insight into the role of each active species in the degradation of antibiotics and offered new ideas for the design and synthesis of efficient and eco-friendly photocatalysts.

Ecotoxicity of five veterinary antibiotics on indicator organisms and water and soil communities.

This study explores the environmental effects of five common veterinary antibiotics widely detected in the environment, (chlortetracycline,CTC; oxytetracycline,OTC; florfenicol,FF; neomycin, NMC; and sulfadiazine, SDZ) on four bioindicators: Daphnia magna, Vibrio fischeri, Eisenia fetida, and Allium cepa, representing aquatic and soil environments. Additionally, microbial communities characterized through 16 S rRNA gene sequencing from a river and natural soil were exposed to the antibiotics to assess changes in population growth and metabolic profiles using Biolog EcoPlates™. Tetracyclines are harmful to Vibrio fisheri (LC50 ranges of 15-25 µg/mL), and the other three antibiotics seem to only affect D. magna, especially, SDZ. None of the antibiotics produced mortality in E. fetida at concentrations below 1000 mg/kg. NMC and CTC had the highest phytotoxicities in A. cepa (LC50 = 97-174 µg/mL, respectively). Antibiotics significantly reduced bacterial metabolism at 0.1-10 µg/mL. From the highest to the lowest toxicity on aquatic communities: OTC > FF > SDZ ≈ CTC > NMC and on edaphic communities: CTC ≈ OTC > FF > SDZ > NMC. In river communities, OTC and FF caused substantial decreases in bacterial metabolism at low concentrations (0.1 µg/mL), impacting carbohydrates, amino acids (OTC), and polymers (FF). At 10 µg/mL and above, OTC, CTC, and FF significantly decreased metabolizing all tested metabolites. In soil communities, a more pronounced decrease in metabolizing ability, detectable at 0.1 µg/mL, particularly affected amines/amides and carboxylic and ketonic acids (p < 0.05). These new ecotoxicity findings underscore that the concentrations of these antibiotics in the environment can significantly impact both aquatic and terrestrial ecosystems.

Plant and microbial response in constructed wetland treating tetracycline antibiotic polluted water: Evaluating the effects of microplastic size and concentration.

Microplastics (MPs) and antibiotics are novel water pollutants that have attracted increasing attention. Constructed wetlands (CWs) are widely applied treating various types of polluted water. How these two new pollutants affect plants and microorganisms in CWs, especially deciphering the unknown roles of MPs size and concentration, is of great essential. Here, five CW treatments with submerged macrophyte Myriophyllum aquaticum were established to treat oxytetracycline (OTC) antibiotic-polluted water. The effects of polystyrene (PS) nanoplastics (NPs) (700 nm) and MPs (90-110 µm) on plant and microbial communities at 10 µg/L and 1 mg/L, respectively, were systematically evaluated. PS reduced the nitrogen and phosphorus removal efficiencies and inhibited OTC removal. Low doses (10 µg/L) of NPs and high doses (1 mg/L) of MPs had the greatest effects on plant and microbial responses. The overall effect of MPs was greater than that of NPs. Compared with high NPs concentration (1 mg/L), low concentrations (10 µg/L) had higher catalase (CAT), superoxide dismutase (SOD), and malondialdehyde (MDA) content. However, the activity and content of MPs at low concentrations (10 µg/L) were lower than those at high concentrations (1 mg/L). The coexistence of OTC and MPs/NPs decreased the microbial diversity and abundance. Low doses of NPs and high doses of MPs decreased the relative abundance of Abditibacteriota, Deinococccota, and Zixibacteria. Redundancy and network analyses revealed a strong correlation between pollutant removal and plant and microbial responses. NH4+-N and OTC removal was positively and negatively correlated with CAT, SOD, and MDA content, respectively. MDA positively correlated to chlorophyll content, whereas SOD showed a negative correlation with Chloroflexi. This study highlighted the scale effect of MPs in wastewater treatment via CWs. It enhances our understanding of the response of plants and microorganisms to the remediation of water co-polluted with MPs and antibiotics.

Effects of Zn and oxytetracycline on mobile genetic elements, antibiotic resistance genes, and microbial community evolution in soil.

Antibiotics and heavy metals added to livestock and poultry feed are excreted in manure, which is added to agricultural soil and causes severe pollution. However, the effects of oxytetracycline (OTC) and zinc (Zn), which are present at relatively high levels in feed additives, on antibiotic resistance genes (ARGs), mobile genetic elements (MGEs), and microbial communities have not been comprehensively studied. This study evaluated the effects of OTC and Zn on environmental factors, microorganisms, MGEs, and ARGs. The expression of MGEs in soil was stimulated by adding Zn at concentrations of 500 and 1000 mg/kg or OTC at concentrations of 30 and 100 mg/kg; however, the addition of their combination hindered the expression of MGEs in soil. The abundance of total MGEs and ARGs tended to decrease with increasing concentrations of Zn and OTC and the number of incubation days. Low and high OTC concentrations strongly inhibited sul and tet resistance genes, respectively. Network analysis showed that changes in the population of Firmicutes and Proteobacteria had the greatest impact on ARG abundance. Redundancy analysis revealed that MGEs, particularly intI2, facilitated the transfer and spread of ARGs and had the greatest impact on changes in ARG abundance. These findings provide reference values for the prevention and resolution of ecological and environmental risks posed by the presence of Zn and OTC in organic manure soil.

The toxicological effect on pak choi of co-exposure to degradable and non-degradable microplastics with oxytetracycline in the soil.

Microplastics and antibiotics are emerging as ubiquitous contaminants in farmland soil, harming crop quality and yield, and thus threatening global food security and human health. However, few studies have examined the individual and joint effects of degradable and/or non-degradable microplastics and antibiotics on crop plants. This study examined the individual and joint effects of polyethylene (PE) and polylactic acid (PLA) microplastics and the antibiotic oxytetracycline (OTC) on pak choi by measuring its growth, photosynthesis, antioxidant enzyme activity, and metabolite levels. Microplastics and/or oxytetracycline adversely affected root weight, photosynthesis, and antioxidant enzyme (superoxide dismutase, catalase, and ascorbate peroxidase) activities. The levels of leaf metabolites were significantly altered, causing physiological changes. Biosynthesis of plant secondary metabolites and amino acids was altered, and plant hormones pathways were disrupted. Separately and together, OTC, PE, and PLA exerted phytotoxic and antagonistic effects on pak choi. Separately and together with OTC, degradable microplastics altered the soil properties, thus causing more severe impacts on plant performance than non-degradable microplastics. This study elucidates the effects on crop plants of toxicity caused by co-exposure to degradable or non-degradable microplastic and antibiotics contamination and suggests mechanisms.

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.

Oxytetracycline-induced inflammatory process without oxidative stress in blue mussels Mytilus trossulus.

Potentially harmful compounds including pharmaceuticals are commonly found in marine waters and sediments. Amongst those, antibiotics and their metabolites are detected worldwide in various abiotic (at concentrations as high as µg/L) and biotic matrices at ng/gram of tissue, posing a risk to non-target species exposed to them such as blue mussels. Amongst those, oxytetracycline (OTC) belongs to the most detected antibiotics in the marine environment. In this work, we concentrated on studying the potential induction of oxidative stress, activation of cellular detoxification processes (including Phase I and Phase II xenobiotic biotransformation enzymes) and multixenobiotic resistance pumps (Phase III) as well as changes in the aromatisation efficiency in Mytilus trossulus exposed to 100 µg/L OTC. Our results show that 100 µg/L OTC concentration did not provoke cellular oxidative stress and did not affect the expression of genes involved in detoxification processes in our model. Moreover, no effect of OTC on aromatisation efficiency was found. Instead, phenoloxidase activity measured in haemolymph was significantly higher in OTC exposed mussels than in those from the control (30.95 ± 3.33 U/L and 17.95 ± 2.75 U/L, respectively). OTC exposed mussels were also characterised by a tissue-dependant activation of major vault protein (MVP) gene expression (1.5 times higher in gills and 2.4 times higher in the digestive system) and a decreased expression of the nuclear factor kappa B-a (NF-κB) gene (3.4 times lower in the digestive system) when compared to those from the control. Additionally, an elevated number of regressive changes and inflammatory responses in tissues such as gills, digestive system and mantle (gonads) was observed underlining the worsening of bivalves' general health. Therefore, instead of a free-radical effect of OTC, we for the first time describe the occurrence of typical changes resulting from antibiotic therapy in non-target organisms like M. trossulus exposed to antibiotics such as OTC.

Non-thermal plasma activated peroxide and percarbonate for tetracycline and oxytetracycline degradation: Synergistic performance, degradation pathways, and toxicity evaluation.

Tetracycline (TC) and Oxytetracycline (OTC) are common antibiotics increasingly detected in the environment, posing a potential risk to human and aquatic lives. Although conventional methods such as adsorption and photocatalysis are used for the degradation of TC and OTC, they are inefficient in removal efficiency, energy yield, and toxic byproduct generation. Herein, a falling-film dielectric barrier discharge (DBD) reactor coupled with environmentally friendly oxidants (hydrogen peroxide (HPO), sodium percarbonate (SPC), and HPO + SPC) was applied, and the treatment efficiency of TC and OTC was investigated. Experimental results showed that moderate addition of the HPO and SPC exhibited a synergistic effect (SF > 2), significantly improving the antibiotic removal ratio, total organic removal ratio (TOC), and energy yield by more than 50%, 52%, and 180%, respectively. After 10 min of DBD treatment, the introduction of 0.2 mM SPC led to a 100% antibiotic removal ratio and a TOC removal of 53.4% and 61.2% for 200 mg/L TC and 200 mg/L OTC, respectively. Also, 1 mM HPO dosage led to 100% antibiotic removal ratios after 10 min of DBD treatment and a TOC removal of 62.4% and 71.9% for 200 mg/L TC and 200 mg/L OTC, respectively. However, the DBD + HPO + SPC treatment method had a detrimental effect on the performance of the DBD reactor. After 10 min of DBD plasma discharge, the removal ratios for TC and OTC were 80.8% and 84.1%, respectively, when 0.5 mM HPO + 0.5 mM SPC was added. Moreover, principal component and hierarchical cluster analysis confirmed the differences between the treatment methods. Furthermore, the concentration of oxidant-induced in-situ generated ozone and hydrogen peroxide were quantitatively determined, and their indispensable roles during the degradation process were established via radical scavenger tests. Finally, the synergetic antibiotic degradation mechanisms and pathways were proposed, and the toxicities of the intermediate byproducts were evaluated.

Oxytetracycline-induced oxidative liver damage by disturbed mitochondrial dynamics and impaired enzyme antioxidants in largemouth bass (Micropterus salmoides).

Oxytetracycline (OTC), a commonly used tetracycline antibiotic in aquaculture, has been found to cause significant damage to the liver of largemouth bass (Micropterus salmoides). This study revealed that OTC can lead to severe histopathological damage, structural changes at the cellular level, and increased levels of reactive oxygen species (ROS) in M. salmoides. Meanwhile, OTC impairs the activities of antioxidant enzyme (such as T-SOD, CAT, GST, GR) by suppressing the activation of MAPK/Nrf2 pathway. OTC disrupts mitochondrial dynamics and mitophagy through via PINK1/Parkin pathway. The accumulation of damaged mitochondria, combined with the inhibition of the antioxidant enzyme system, contributes to elevated ROS levels and oxidative liver damage in M. salmoides. Further investigations demonstrated that an enzyme-treated soy protein (ETSP) dietary supplement can help maintain mitochondrial dynamic balance by inhibiting the PINK1/Parkin pathway and activate the MAPK/Nrf2 pathway to counteract oxidative damage. In summary, these findings highlight that exposure to OTC disrupts mitochondrial dynamics and inhibits the antioxidant enzyme system, ultimately exacerbating oxidative liver damage in M. salmoides. We propose the use of a dietary supplement as a preventive measure against OTC-related side effects, providing valuable insights into the mechanisms of antibiotic toxicity in aquatic environments.

Stress of cupric ion and oxytetracycline in Chlorella vulgaris cultured in swine wastewater.

Chlorella culturing has the advantages in treatment of wastewater including swine wastewater from anaerobic digesters due to the product of biolipids and the uptake of carbon dioxide. However, there often exist high concentrations of antibiotics and heavy metals in swine wastewater which could be toxic to chlorella and harmful to the biological systems. This study examined the stress of cupric ion and oxytetracycline (OTC) at various concentrations on the nutrient removal and biomass growth in Chlorella vulgaris culturing in swine wastewater from anaerobic digesters, and its biochemical responses were also studied. Results showed that dynamic hormesis of either OTC concentration or cupric ion one on Chlorella vulgaris were confirmed separately, and the presence of OTC not only did not limit biomass growth and lipids content of Chlorella vulgaris but also could mitigate the toxicity of cupric ion on Chlorella vulgaris in combined stress of Cu2+ and OTC. Extracellular polymeric substances (EPS) of Chlorella vulgaris were used to explain the mechanisms of stress for the first time. The content of proteins and carbohydrates in EPS increased, and the fluorescence spectrum intensity of tightly-bound EPS (TB-EPS) of Chlorella vulgaris decreased with increasing concentration of stress because Cu2+ and OTC may be chelated with proteins of TB-EPS to form non-fluorescent characteristic chelates. The low concentration of Cu2+ (≤1.0 mg/L) could enhance the protein content and promote the activity of superoxide dismutase (SOD) while these parameters were decreased drastically under 2.0 mg/L of Cu2+. The activity of adenosine triphosphatase (ATPase) and glutathione (GSH) enhanced with the increase of OTC concentration under combined stress. This study helps to comprehend the impact mechanisms of stress on Chlorella vulgaris and provides a novel strategy to improve the stability of microalgae systems for wastewater treatment.

Effect of antibiotic cocktail exposure on functional disturbance of nitrifying microbiome.

The present study quantitatively determined the degree and type of functional disturbance in the nitrifying microbiome caused by exposure to a single oxytetracycline (OTC) and a two-antibiotic mixture containing OTC and sulfamethoxazole (SMX). While the single antibiotic had a pulsed disturbance on nitritation that was recoverable within three weeks, the antibiotic mixture caused a more significant pulsed disturbance on nitritation and a potential press disturbance on nitratation that was not recoverable for over five months. Bioinformatic analysis revealed significant perturbations for both canonical nitrite-oxidizing (Nitrospira defluvii) and potential complete ammonium-oxidizing (Ca. Nitrospira nitrificans) populations that were strongly associated with the press perturbation on nitratation. In addition to this functional disturbance, the antibiotic mixture reduced the biosorption of OTC and altered its biotransformation pathways, resulting in different transformation products compared with those produced when OTC was treated as a single antibiotic. Collectively, this work elucidated how the antibiotic mixture can affect the degree, type, and duration of the functional disturbance on nitrifying microbiome and offer new insights into the environmental consequences of antibiotic residues (e.g., their fate, transformation, and ecotoxicity) when present as an antibiotic mixture rather than single antibiotics.

An alternative approach to assess ecotoxicological effects of agrochemical combinations used in Brazilian aquaculture farms.

Agrochemicals used for treating and preventing aquaculture diseases are usually present in combination with other compounds, and the toxicity resulting from their chemical interactions presents an important reason to assess the ecotoxicity of compound mixtures in view to better understanding the joint action of chemicals and avoiding their environmental impacts. In this study, we evaluated the acute aquatic ecotoxicity of several compounds used in Brazilian fish farming (Oxytetracycline [OXT], Trichlorfon [TRC], and BioFish® [BIO]), both individually and in binary and ternary mixtures. Initial test concentrations were prepared according to the recommended concentrations for aquaculture application, and from these, a geometric dilution series was tested on two important fresh water quality indicator species, the microcrustacean Daphnia magna and the bacterium Aliivibrio fischeri. At the recommended pond application rate, TRC and BIO applied individually showed toxicity to the tested organisms in terms of the lowest-observed-effect concentration (LOEC), and D. magna was always more sensitive than A. fischeri. For the two test organisms, the results obtained with the binary mixtures showed that the TRC and BIO mixture was more toxic than TRC and OXT, which in turn was more toxic than OXT and BIO. The toxicity from all agrochemicals in the ternary mixture was more than that of the agrochemical combinations in the binary mixtures. Given the results presented in this study, it is evident that the mode of action and availability of the tested compounds undergo changes that increase toxicity when they are present in combination, and therefore, aquaculture wastewater treatment should be adopted to ensure decontamination of agrochemical residues.

Early environmental exposure to oxytetracycline in Danio rerio may contribute to neurobehavioral abnormalities in adult zebrafish.

The common antibiotic oxytetracycline (OTC) is nowadays commonly found in natural aquatic environments. However, the underlying mechanisms of low-dose OTC exposure and its neurotoxic effects on aquatic animals remain unknown. In this study, we exposed zebrafish larvae to environmental concentrations of OTC in early life and performed neurobehavioral, 16S rRNA gene sequencing, and transcriptomic analyses. OTC exposure resulted in hyperactivity of larvae and a significant reduction in the number of neurons in the midbrain. The expression levels of 15 genes related to neural function changed. Additionally, the composition of 65 genera of the gut microbiota of larvae was altered, which may be one of the reasons for the abnormal neural development. We further studied the long-term outcomes among adult fish long after cessation of OTC exposure. OTC treatment caused adult fish to be depressive and impulsive, symbolizing bipolar disorder. Adult fish exposed to OTC had significantly fewer neurons and their gut bacteria composition did not recover 104 days after terminating OTC exposure. Finally, we analyzed the correlation between the gut microbiota of larvae, genes related to neural function, and metabolites of adult fish brain tissue. The results showed that the abundance of several members of the biome in larvae was related to the transcription levels of genes related to neural function, which were related to the metabolic levels in the adult brain. In conclusion, our study showed that early-life exposure to environmental concentrations of OTC can lead to persistent neurobehavioral abnormalities until adulthood through dysbiosis in the gut microbiota.

Assessment of Nonalcoholic Fatty Liver Disease Symptoms and Gut-Liver Axis Status in Zebrafish after Exposure to Polystyrene Microplastics and Oxytetracycline, Alone and in Combination.

BACKGROUND: Environmental pollution may give rise to the incidence and progression of nonalcoholic fatty liver disease (NAFLD), the most common cause for chronic severe liver lesions. Although knowledge of NAFLD pathogenesis is particularly important for the development of effective prevention, the relationship between NAFLD occurrence and exposure to emerging pollutants, such as microplastics (MPs) and antibiotic residues, awaits assessment. OBJECTIVES: This study aimed to evaluate the toxicity of MPs and antibiotic residues related to NAFLD occurrence using the zebrafish model species. METHODS: Taking common polystyrene MPs and oxytetracycline (OTC) as representatives, typical NAFLD symptoms, including lipid accumulation, liver inflammation, and hepatic oxidative stress, were screened after 28-d exposure to environmentally realistic concentrations of MPs (0.69mg/L) and antibiotic residue (3.00µg/L). The impacts of MPs and OTC on gut health, the gut-liver axis, and hepatic lipid metabolism were also investigated to reveal potential affecting mechanisms underpinning the NAFLD symptoms observed. RESULTS: Compared with the control fish, zebrafish exposed to MPs and OTC exhibited significantly higher levels of lipid accumulation, triglycerides, and cholesterol contents, as well as inflammation, in conjunction with oxidative stress in their livers. In addition, a markedly smaller proportion of Proteobacteria and higher ratios of Firmicutes/Bacteroidetes were detected by microbiome analysis of gut contents in treated samples. After the exposures, the zebrafish also experienced intestinal oxidative injury and yielded significantly fewer numbers of goblet cells. Markedly higher levels of the intestinal bacteria-sourced endotoxin lipopolysaccharide (LPS) were also detected in serum. Animals treated with MPs and OTC exhibited higher expression levels of LPS binding receptor (LBP) and downstream inflammation-related genes while also exhibiting lower activity and gene expression of lipase. Furthermore, MP-OTC coexposure generally exerted more severe effects compared with single MP or OTC exposure. DISCUSSION: Our results suggested that exposure to MPs and OTC may disrupt the gut-liver axis and be associated with NAFLD occurrence. https://doi.org/10.1289/EHP11600.

Induced development of oxytetracycline tolerance in bacterial communities from soil amended with well-aged cow manure.

The use of animal manure as organic fertilizer is a common agricultural practice that can improve soil health and crop yield. However, antibiotics and their metabolites are often present in animal manure and, hence, in manure-amended soil. The aim of this study was to assess the induced development of oxytetracycline (OTC) tolerance in soil bacterial communities as a result of the addition of OTC to soil amended with well-aged cow manure. To this purpose, soil amended with well-aged cow manure was repeatedly - three times - spiked with different OTC concentrations (0, 2, 20, 60, 150, and 500 mg OTC kg-1 dry weight soil, each time) according to a pollution-induced community tolerance (PICT) assay. The PICT detection phase was conducted in Biolog EcoPlatesTM in the presence of the following OTC concentration gradient in the wells: 0, 5, 20, 40, 60, and 100 mg L-1. For all treatments, the application of OTC in the PICT selection phase resulted in lower values of bacterial metabolic activity (i.e., lower values of average well color development) in the PICT detection phase. A significant increase in OTC tolerance was observed in soil bacterial communities that had been exposed three times to ≥ 20 mg OTC kg-1 DW soil during the PICT selection phase. In general, higher levels of OTC exposure during the PICT selection phase resulted in bacterial tolerance to higher OTC concentrations during the PICT detection phase, pointing to a dose-dependent induced tolerance. It is important to (i) rationalize the amount of antibiotics administered to livestock, and (ii) treat properly the antibiotic-containing manure before its application to agricultural soil as fertilizer.