Dietary Lactobacillus plantarum ST-III alleviates the toxic effects of triclosan on zebrafish (Danio rerio) via gut microbiota modulation.

The probiotics, Lactobacillus plantarum ST-III, plays an important role in modulating microbiota and alleviating intestinal metabolic disorders. Herein, we reported that Lactobacillus increases biodiversity of zebrafish gut flora, and attenuates toxic effects from chronic triclosan (TCS) exposure. Lactobacillus-feeding recovered the species and amount of microorganisms in the intestines of zebrafish, and inhibited toxin production by saprophytic bacterial growth. Abnormal physiological indexes and malonaldeyhde content resulting from TCS exposure were effectively alleviated. Additionally, lipid-metabolism disorders, such as increased triglyceride and total cholesterol levels, were attenuated by a probiotics diet. The number of CD4+ T cell lymphocytes in the lamina propria of the duodenal mucosa was decreased in zebrafish receiving a Lactobacillus diet compared to the TCS-exposed group, showing a consistent expression trend for six immune genes (NF-κB, IL-1ß, TNF-α, lysozyme, TLR4α, IL-10) in the intestinal mucosa. Histopathological observations of intestines, spleen and kidney showed that TCS exposure produced severe damage to the morphology and structure of immune and metabolism-related organs. Lactobacillus was capable of mitigating this damage, but bile salt hydrolase, an active extract of Lactobacillus, was not an effective mitigation strategy. The Lactobacillus-induced decrease in the number of inflammatory cells confirmed its role in preventing inflammatory injury. Three behavioral tests (T-maze, bottom dwelling and social interaction) indicated that a probiotics diet improved zebrafish movement and learning/memory capacity, effectively alleviating anxiety behavior due to TCS exposure. These findings inform development of beneficial strategies to alleviate intestinal metabolic syndromes and neurodegenerative diseases resulting from exposure to environmental contaminants through modifying gut flora with a probiotics diet.

Effects of miR-181a-5p abnormal expression on zebrafish (Danio rerio) vascular development following triclosan exposure.

Triclosan (TCS), one of the important bactericides, is widely used in personal care products, and its chronic exposure leads to severe toxic effects on the growth and development of blood vessels in zebrafish (Danio rerio). Herein, we screened out three differentially expressed miRNAs (miR-181a-5p, miR-132-3p and miR-128-3p) by sequencing and qRT-PCR analyses of 4-96-hpf TCS-exposed zebrafish, among which miR-181a-5p was found to regulate many signaling pathways involved in fatty acid biosynthesis and phosphatidylimositol signaling systems. By O-dianisidine staining, TCS-exposure resulted in decreased distribution of red blood cells and induced blood hypercoagulable state and thrombotic effects. Defective subintestinal veins (SIVs), and decreased branching and curvature of blood vessels were observed with increasing TCS-exposure concentrations. After microinjection of miR-181a-5p mimic and inhibitor, zebrafish malformation type and percentage were prominently increased such as distorted SIV vessels along with reduced venation and abnormal branches by ALP staining. Overexpressed miR-181a-5p had a greater effect on development and branching patterns of arteries and veins than its knockdown. By laser confocal microscopy observation, the 72-hpf Tg (flk1: mCherry) zebrafish obviously displayed vascular proliferation and ablation in the miR-181a-5p mimic group. Microinjection of miR-181a-5p mimics and inhibitors led to abnormal expressions (20-50%) of two key target genes (pax2a and vash2) by WISH, and increased malformation percentages (18-45%) by IOD analysis. Overexpression of vash2 led to the inhibitory or promoting effects on the expression of PI3K signaling pathway-related genes, proving that the effect of vash2 on development of blood vessels could be realized by inhibiting PI3K signaling pathway. These observations lay theoretical foundation for deep insight into the molecular mechanisms on TCS-induced cardiovascular diseases.

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.