Exposure to methylparaben at environmentally realistic concentrations significantly impairs neuronal health in adult zebrafish.

Methylparaben (MeP) is an emerging aquatic pollutant that is found to impact neural functions. However, it still lacks a comprehensive understanding about its neurotoxicology. The present study exposed adult zebrafish to environmentally realistic concentrations (0, 1, 3, and 10 µg/L) of MeP for 28 days, with objectives to elucidate the neurotoxic effects and mechanisms. Proteomic profiling found that MeP pollutant induced distinct mechanism of neurotoxicity as a function of sex. MeP pollutant appeared to preferentially target the neurotransmission cascade via synapse junctions. In male brain, glutamatergic neural signaling was enhanced by 10 µg/L of MeP in characteristics of higher glutamate neurotransmitter content (by 61.9%) and up-regulated glutamate receptor expression by 2.6-fold relative to the control. In MeP-exposed female brain, biomarker proteins of synapse formation and regeneration had significantly lower abundance, accounting for the blockage of synaptic neurotransmission. Furthermore, under the stress of MeP pollutant, both male and female zebrafish initiated a negative feedback mechanism along stress neuroendocrine axis by down-regulating the transcriptions of corticotropin-releasing hormone and its binding protein, which subsequently decreased blood cortisol concentrations. MeP subchronic exposure also disturbed innate immune function. In particular, significant increases in lipopolysaccharide (LPS) content by 15.6% were caused by MeP exposure in male brain, thereby inducing the synthesis of pro-inflammatory cytokines. In contrast, female brain was able to adaptively up-regulate the protein expression of blood brain barrier to inhibit the infiltration of LPS endotoxin into brain. Overall, the present findings pinpoint the potent neurotoxicity of MeP pollutant even at environmentally realistic concentrations.

Dysfunction of liver-gut axis in marine medaka exposed to hypoxia and perfluorobutanesulfonate.

With objectives to explore the interactive mode on the function of liver-gut axis, adult marine medaka were exposed for 7 days to environmentally realistic concentrations of perfluorobutanesulfonate (PFBS) (0 and 10 µg/L) under normoxia or hypoxia condition. Furthermore, PFBS exposure was extended to 21 days to reveal the temporal progression in toxicity. The results showed that hypoxia exposure significantly disturbed lipid metabolism, caused oxidative damage, and induced inflammation in the livers of medaka. The composition of gut microbiota was also drastically shifted by hypoxia acute exposure. In contrast, the effect of PFBS was much milder. Hypoxia was thus the determinant of the combined toxicity. Depending on the exposure duration, a time-course recovery from PFBS innate toxicity was generally noted. Overall, the present study underlines the hypoxic and temporal variation in the dysregulation of liver-gut axis by PFBS, which is expected to support a comprehensive ecological risk assessment.

Toxicity of perfluorobutanesulfonate on gill functions of marine medaka (Oryzias melastigma): A time course and hypoxia co-exposure study.

Perfluorobutanesulfonate (PFBS) is found in hypoxia regions. Results of previous studies have shown that hypoxia was capable of altering the inherent toxicity of PFBS. However, regarding gill functions, hypoxic influences and time course progression of toxic effects of PFBS remain unclear. In this study, with the aim to reveal the interaction behavior between PFBS and hypoxia, adult marine medaka Oryzias melastigma were exposed for 7 days to 0 or 10 µg PFBS/L under normoxic or hypoxic conditions. Subsequently, to explore the time-course transition in gill toxicity, medaka were exposed to PFBS for 21 days. The results showed that hypoxia dramatically increased the respiratory rate of medaka gill, which was further enhanced by exposure to PFBS; although exposure to PFBS under normoxic conditions for 7 days did not alter respiration, exposure to PFBS for 21 days significantly accelerated the respiration rate of female medaka. Concurrently, both hypoxia and PFBS were potent to interrupt the gene transcriptions and Na+, K+-ATPase enzymatic activity that play pivotal roles in the osmoregulation in gills of marine medaka, consequently disrupting homeostasis of major ions in blood, such as Na+, Cl-, and Ca2+. In addition, composition and diversity of the microbiome residing on surfaces of the gill were profiled by using amplicon sequencing. Acute exposure to hypoxia for only 7 days caused a significant decrease in diversity of the bacterial community of gill whatever the presence of PFBS, while PFBS exposure for 21 days increased the diversity of gill microbial community. Principal component analysis revealed that, compared with PFBS, hypoxia was the predominant driver of gill microbiome dysbiosis. Depending on duration of exposure, a divergence was caused in the microbial community of gill. Overall, the current findings underline the interaction between hypoxia and PFBS on gill function and demonstrate the temporal variation in PFBS toxicity.

Variability in fecal metabolome depending on age, PFBS pollutant, and fecal transplantation in zebrafish: A non-invasive diagnosis of health.

To protect the wellbeing of research animals, certain non-invasive measures are in increasing need to facilitate an early diagnosis of health and toxicity. In this study, feces specimen was collected from adult zebrafish to profile the metabolome fingerprint. Variability in fecal metabolite composition was also distinguished as a result of aging, perfluorobutanesulfonate (PFBS) toxicant, and fecal transplantation. The results showed that zebrafish feces was very rich in a diversity of metabolites that belonged to several major classes, including lipid, amino acid, carbohydrate, vitamin, steroid hormone, and neurotransmitter. Fecal metabolites had functional implications to multiple physiological activities, which were characterized by the enrichment of digestion, absorption, endocrine, and neurotransmission processes. The high richness and functional involvement of fecal metabolites pinpointed feces as an abundant source of diagnostic markers. By comparison between young and aged zebrafish, fundamental modifications of fecal metabolomes were caused by aging progression, centering on the neuroactive ligand-receptor interaction pathway. Exposure of aged zebrafish to PFBS pollutant also significantly disrupted the metabolomic structure in feces. Of special concern were the changes in fecal hormone intermediates after PFBS exposure, which was concordant with the in vivo endocrine disrupting effects of PFBS. Furthermore, it was intriguing that transplantation of young zebrafish feces efficiently mitigated the metabolic perturbation of PFBS in aged recipients, highlighting the health benefits of therapeutic strategies based on gut microbiota manipulation. In summary, the present study provides preliminary clues to evidence the non-invasive advantage of fecal metabolomics in the early diagnosis and prediction of physiology and toxicology.

Endocrine disruption and reproductive impairment of methylparaben in adult zebrafish.

Methylparaben (MeP) is one of the most frequently used preservatives in our daily products. However, it is becoming an aquatic pollutant of emerging concern. To reveal the endocrine disruption mechanism and reproductive impairment of MeP, the present study exposed adult zebrafish to 0, 1, 3, and 10 µg/L (0, 6.6, 19.7, and 65.7 nM) of MeP for 28 days. The results showed that subchronic exposure to 10 µg/L of MeP significantly increased the gonadosomatic index in zebrafish. Spermatogenesis and oogenesis were blocked by MeP at concentrations as low as 1 µg/L. Furthermore, parental exposure to MeP induced developmental deficits in offspring larvae, by increasing mortality, stimulating precocious hatching, and elevating heart rate. Blood concentrations of estradiol, testosterone, and 11-keto-testosterone were consistently lowered in MeP exposure groups. Transcriptional results evidenced that the disturbance in steroidogenesis and feedback regulation mechanisms along the hypothalamic-pituitary-gonadal axis underlay the imbalance of sex hormones. In line with the low estradiol level, hepatic production of vitellogenin (VTG) was significantly down-regulated, subsequently leading to a deficiency of VTG supply during oogenesis. To our knowledge, this is the first study to provide systemic insight about the antiestrogenic activity and reproductive toxicity of MeP.

Transcriptomic Interaction between Young Fecal Transplantation and Perfluorobutanesulfonate in Aged Zebrafish Gonads.

The transfer of young fecal microbiota has been found to significantly refresh the reproductive endocrine system and effectively ameliorate the toxicity of perfluorobutanesulfonate (PFBS) in aged zebrafish recipients. However, the mechanisms underlying the antagonistic action of young fecal microbiota against the reproductive endocrine toxicity of PFBS remain largely unknown. In this study, the aged zebrafish were transplanted with feces from young donors and then exposed to PFBS for 14 days. After exposure, the shift in the transcriptomic fingerprint of the gonads was profiled by using high-throughput sequencing, aiming to provide mechanistic clues into the interactive mode of action between young fecal transplantation and PFBS's innate toxicity. The results showed that the gene transcription pattern associated with protein and lipid synthesis in the gonads of the aged individuals was quite different from the young counterparts. It was intriguing that the transplantation of young feces established a youth-like transcriptomic phenotype in the elderly recipients, thus attenuating the functional decline and maintaining a healthy aging state of the gonads. A sex specificity response was clearly observed. Compared to the aged females, more metabolic pathways (e.g., glycine, serine, and threonine metabolism; glyoxylate and dicarboxylate metabolism; pyrimidine metabolism) were significantly enriched in aged males receiving young feces transplants. PFBS dramatically altered the transcriptome of aged testes, while a much milder effect was observable in aged ovaries. Accordingly, a suite of biological processes related to germ cell proliferation were disrupted by PFBS in aged males, including the ECM-receptor interaction, retinol metabolism, and folate biosynthesis. In aged ovaries exposed to PFBS, mainly the fatty acid and arginine biosynthesis pathway was significantly affected. However, these transcriptomic disorders caused by PFBS were largely mitigated in aged gonads by transferring young feces. Overall, the present findings highlighted the potential of young fecal transplantation to prevent the functional compromise of gonads resulting from aging and PFBS.

Young fecal transplantation mitigates the toxicity of perfluorobutanesulfonate and potently refreshes the reproductive endocrine system in aged recipients.

The aging process leads to the gradual impairment of physiological functions in the elderly, making them more susceptible to the toxicity of environmental pollutants. In this study, aged zebrafish were first transplanted with the feces from young donors and subsequently exposed to perfluorobutanesulfonate (PFBS), an emerging persistent toxic pollutant. The interaction between young fecal transplant and PFBS inherent toxicity was investigated, focusing on reproductive performance and the underlying endocrine mechanism. The results showed that PFBS single exposure increased the percentage of primary oocytes in aged ovaries, implying a blockage of oogenesis. However, transplantation of young feces completely abolished the effects of PFBS and promoted oocyte growth, as inferred by the obviously lower percentage of primary oocytes, accompanied by a higher percentage of cortical-alveolar oocytes. Measurement of sex hormones found that PFBS significantly increased the blood concentration of estradiol and disrupted the balance of sex hormones in the elderly, which were, however, efficiently ameliorated by young fecal transplantation. Based on gene transcription along the hypothalamic-pituitary-gonadal axis, hierarchical clustering analysis showed similar profiles of the reproductive endocrine system between young zebrafish and their aged counterparts transplanted with young feces, implying that young fecal transplantation might refresh the endocrine system of aged recipients, regardless of PFBS exposure. The increased transcription levels of mRNAs encoding vitellogenin, activinBA, and membrane bound progestin receptors would cooperatively enhance the growth and maturation of oocytes in the ovaries of aged zebrafish receiving young fecal transplantation. Overall, the findings highlighted the potent efficacy of young fecal transplantation to improve the reproductive function of the elderly and to mitigate the endocrine disruption of an environmental pollutant. These findings are expected to broaden our understanding of the efficacy, mechanisms, and application of fecal transplantation.

Hepatotoxicity caused by methylparaben in adult zebrafish.

Parabens are a class of aquatic pollutants of emerging concern, among which methylparaben (MeP) causes severe pollution worldwide. However, aquatic toxicology of MeP remains largely unknown, which hinders ecological risk evaluation. In the present study, adult zebrafish were exposed to environmentally realistic concentrations (0, 1, 3, and 10 µg/L) of MeP for 28 days, with objectives to reveal the hepatotoxicity based on transcriptional, biochemical, metabolomics, and histopathological evidences. The results showed that MeP subchronic exposure induced the occurrence of hepatocellular vacuolization in zebrafish. The most severe symptom was noted in 10 µg/L MeP-exposed female liver, which was characterized by rupture of cell membrane and small nuclei. In addition, MeP exposure disturbed the balance between oxidative stress and antioxidant capacity. Lipid metabolism dynamics across gut, blood, and liver system were significantly dysregulated after MeP exposure by altering the transcriptions of lipid nuclear receptors and concentrations of key metabolites. Metabolomic profiling of MeP-exposed liver identified differential metabolites mainly belonging to fatty acyls, steroids, and retinoids. In particular, hepatic concentration of cortisol was increased in male liver by MeP pollutant, implying the activation of stress response. Exposure to MeP also inhibited the synthesis and conjugation of primary bile acid (e.g., 7-ketolithocholic acid and taurochenodeoxycholic acid) in female liver. Furthermore, degradation of biologically active molecules, including retinoic acid and estradiol, was enhanced in the liver by MeP. Overall, the present study highlights the hepatotoxicity caused by MeP pollutant even at environmentally realistic concentrations, which necessitates an urgent and accurate risk assessment.

Disturbed glucose metabolism by perfluorobutanesulfonate pollutant and benefit of young fecal transplantation in aged zebrafish.

Perfluorobutanesulfonate (PFBS) is an environmental pollutant of emerging concern, which significantly impacts the metabolism and health of animals. Because of the loss of functional capacity, the aged animals are regarded more susceptible to the toxicity of environmental pollutants. In the present study, aged zebrafish were employed as the toxicological animal and transplanted with the feces collected from young donors for 14 days, after which the acclimated elderly were exposed to PFBS at environmentally relevant concentrations (0 and 100 µg/L) for another 14 days. When the exposure was concluded, glucose metabolic disturbances of PFBS in the aged and efficacy of young fecal transplant to mitigate the toxicity of PFBS were explored along the gut-liver axis. The results showed that PFBS exposure significantly inhibited the enzymatic activity of α-amylase in the gut, but increased the alanine aminotransferase (ALT) activity in the blood of the aged zebrafish, suggesting the impairment of intestinal digestive functions of carbohydrates and the induction of liver damage by PFBS. However, young fecal transplantation successfully ameliorated the toxicity of PFBS on α-amylase and ALT, underlining the benefits conveyed to the health of the elderly. In addition, transplantation of young feces was efficient to alleviate the hyperglycemia symptom in the elderly via stimulating the secretion of insulin. PFBS exposure increased blood glucagon level, disrupted insulin receptor transcription, and depleted hepatic glycogen store, which were all mitigated by young fecal transplant. Hepatic proteomic analysis also found dynamic interactions between young fecal transplantation and PFBS pollutant on the metabolic pathways of glucose and glycogen, involving glycolysis, gluconeogenesis, glycogenesis, and glycogenolysis. Overall, the present findings highlighted the beneficial effects of young fecal transplantation to protect the aged from the glucose metabolism toxicity of PFBS, thus providing a plausible measure to improve the health aging status.

Significant impairment of intestinal health in zebrafish after subchronic exposure to methylparaben.

Methylparaben (MeP) is a ubiquitous pollutant in aquatic environment, which has caused severe pollution worldwide. However, aquatic toxicology of MeP is still largely unknown. In the present study, adult zebrafish were exposed to environmentally realistic concentrations of MeP (0, 1, 3, and 10 µg/L) for 28 days. In terms of the antimicrobial nature, dysregulation of gut microbiota and zebrafish health by MeP were elucidated after exposure. High-throughput amplicon sequencing showed that MeP subchronic exposure was able to disrupt the composition and diversity of gut microbial community, which was characterized by the alterations in alpha diversity and divergent distribution by principal component analysis. In addition, MeP exposure increased the body length and body weight of female fish, implying stimulated growth at low doses. In male intestine, consistent increases were notable in goblet cell density, tight junction protein (TJP) 2 expression, and serotonin neurotransmitter concentration after MeP exposure. In contrast, female intestine exposed to MeP had lower density of goblet cells, inhibited expression of TJP2, reduced concentration of serotonin, but up-regulated transcription of pro-inflammatory cytokine. Under the stress of MeP pollutant, intestinal catalase antioxidant enzyme was activated, thus contributing to the removal of oxidative free radicals. Correlation analysis verified the modulation of TJP2 expression by Lactobacillus probiotic bacteria. Disturbances in goblet cell, tight junctions, and serotonin by MeP may be combined to interfere with gut barrier function. Overall, the present study highlights the impairment of intestinal health by environmentally realistic concentrations of MeP, which necessitates an urgent risk assessment.

Fecal transplantation from young zebrafish donors efficiently ameliorates the lipid metabolism disorder of aged recipients exposed to perfluorobutanesulfonate.

Aging is a biological process that is accompanied by the gradual loss of physiological functions. Under the context of ubiquitous and persistent environmental pollution, the elderly will be more vulnerable to the detrimental effects of toxic pollutants than the young. With objectives to explore effective measures to ameliorate the double stress of aging and toxicants, the present study transplanted the feces from young zebrafish donors to aged recipients, which were concurrently exposed to perfluorobutanesulfonate (PFBS), an emerging environmental pollutant of international concern. After exposure, growth, hepatic structural organization, and lipid metabolism were examined. The results showed that, irrespective of PFBS toxicity, transplantation of young feces significantly enhanced the growth of the aged. In the livers of aged and PFBS-exposed zebrafish, vacuolization symptom was prevalently observed, while young fecal transplantation alleviated the structural defects in aged livers. In the gut of the elderly, digestive activity of lipids was promoted after the transplantation of young feces. The blood of the aged females accumulated significantly higher concentration of triglyceride (TG) than the young counterparts (2.6-fold), implying that the elderly were at high risk of cardiovascular diseases. PFBS treatment of the aged further increased blood TG levels by 2.0-fold relative to the aged control group, pointing to the aggravation of the health of the elderly by environmental pollution. However, it is intriguing that young fecal transplantation efficiently inhibited the metabolic toxicity of PFBS and restored the normal level of blood TG, which provided more evidence about the benefit of young fecal transplant to improve the health of the aged individuals. In the aged livers transplanted with young feces, mitochondrial ß-oxidation of fatty acids was consistently activated. Overall, the present study verified the efficacy of young fecal transplantation to mitigate the metabolic disorders resulting from aging and an environmental pollutant.

Metabolomic profiles in zebrafish larvae following probiotic and perfluorobutanesulfonate coexposure.

Probiotic supplements are able to attenuate the developmental toxicity of perfluorobutanesulfonate (PFBS) pollutant. However, the underlying mechanisms remain unexplored. To this end, the present study acutely exposed zebrafish larvae for 4 days to 0 and 10 mg/L of PFBS, with or without the addition of probiotic Lactobacillus rhamnosus in the rearing water. The metabolomics approach was used to reveal the combined effects of PFBS and probiotics on metabolic dynamics, based on which gene transcriptions, enzymatic activities, and behavioral endpoints were further examined. The results showed that probiotic supplements were the major driver of the metabolomic fingerprints in coexposed zebrafish larvae. The addition of probiotic bacteria significantly decreased the methylation potential whilst up-regulating the demethylation process of genomic DNA, which may globally stimulate the gene expression to improve somatic growth. Acute exposure to PFBS significantly increased the cortisol concentration in zebrafish larvae, subsequently inducing stress response and hyperactive behavior. In contrast, probiotic supplementation promoted the degradation of cortisol, thus alleviating the stressful state. Antagonistic action of probiotics against PFBS developmental toxicity was also noted regarding the locomotor behavior. In addition, gut microbiota-mediated production of secondary bile acids was remarkably enhanced by probiotic supplements regardless of PFBS exposure. Overall, the present study underlines the efficacy of probiotic bacteria to protect zebrafish larvae from the metabolic disturbances of PFBS, thereby providing more evidence to support the application of probiotics in aquaculture and fishery as an environmentally-friendly choice.

Probiotics inhibit the stunted growth defect of perfluorobutanesulfonate via stress and thyroid axes in zebrafish larvae.

Perfluorobutanesulfonate (PFBS) is an emerging pollutant in aquatic environments and potently disrupts the early developmental trajectory of teleosts. Considering the persistent and toxic nature of PFBS, it is necessary to develop in situ protective measures to ameliorate the toxic damage of PFBS. Probiotic supplements are able to mitigate the growth retardation defects of PFBS. However, the interactive mechanisms remain elusive. To this end, this study acutely exposed zebrafish larvae to a concentration gradient of PFBS (0, 1, 3.3 and 10 mg/L) for 4 days, during which probiotic bacteria Lactobacillus rhamnosus were added in the rearing water. After exposure, alterations in gene transcriptions and key hormones along the hypothalamus-pituitary-interrenal (HPI), growth hormone/insulin-like growth factor (GH/IGF) and hypothalamus-pituitary-thyroid (HPT) axes were examined. The results showed that PFBS single exposure significantly increased the cortisol concentrations, suggesting the induction of stress response, while probiotic supplementation effectively decreased the cortisol levels in coexposed larvae in an attempt to relieve the stress of PFBS toxicant. It was unexpected that probiotic additive significantly decreased the larval GH concentrations independent of PFBS, thereby eliminating the contribution of GH/IGF axis to the growth improvement of probiotics. In contrast, probiotic bacteria remarkably increased the concentration of thyroid hormones, particularly the thyroxine (T4), in zebrafish larvae. The pronounced down-regulation of uridinediphosphate glucoronosyltransferases (UDPGT) gene pointed to the blocked elimination process of T4 by probiotics. Furthermore, proteomic fingerprinting found that probiotics were potent to shape the protein expression pattern in PFBS-exposed zebrafish larvae and modulated multiple biological processes that are essential for the growth. In summary, the present findings suggest that HPI and HPT axes may cooperate to enhance the growth of fish larvae under PFBS and probiotic coexposures.

Probiotic supplementation mitigates the developmental toxicity of perfluorobutanesulfonate in zebrafish larvae.

Perfluorobutanesulfonate (PFBS) is an emerging pollutant of international concern, which is found to impair the early embryonic development of fishes. In the context of ubiquitous and persistent pollution, it is necessary to explore mitigatory strategies against the developmental toxicity of PFBS. In this study, zebrafish larvae were acutely exposed to 0, 1, 3.3 and 10 mg/L of PFBS till 168 h post-fertilization (hpf), during which probiotic Lactobacillus rhamnosus bacteria were administered via the exposure media. After the singular or combined exposure, interaction between PFBS and probiotics on the growth of zebrafish larvae was measured. PFBS exposure significantly decreased the larval body weight, weight gain and specific growth rate, while probiotic supplementation efficiently inhibited the growth retardation caused by PFBS. Furthermore, PFBS and probiotic combinations remarkably activated the antioxidant capacity to timely scavenge the reactive oxidative species and protect the larvae from lipid peroxidation. Biochemical assay and fluorescent staining verified that PFBS exposure significantly promoted the production of bile acids, which were further enhanced by the probiotics. In coexposed zebrafish larvae, up-regulation of peroxisome proliferator-activated receptor (PPARb) would enhance the ß-oxidation of fatty acids to meet the energy demand from larval growth, subsequently decreasing fatty acid concentrations. In addition, probiotic supplements masked the dysbiosis of PFBS and potently shaped the gut microbiota, which closely modulated the production of bile acids. Overall, the present findings underline the beneficial effects of probiotics to protect the developing larvae from the aquatic toxicities of PFBS, thus highlighting the potential application values of probiotic recipe in aquaculture and ecological reservation.

Fingerprinting fecal DNA and mRNA as a non-invasive strategy to assess the impact of polychlorinated biphenyl 126 exposure on zebrafish.

In toxicological studies, experimental animals are generally subjected to dissection to obtain the tissues of concern, which causes great harm to the animals. In this regard, it is necessary to test and develop a non-invasive strategy to prevent the animals from anthropic injury when achieving scientific objectives. Therefore, zebrafish fecal DNA and mRNA pools were assessed by using metagenomic and transcriptomic analyses based on their potential to diagnose toxicological impairment of polychlorinated biphenyl (PCB) 126, a model persistent organic pollutant. The results showed that there was abundant zebrafish DNA and mRNA in the feces, which were, however, associated with contrasting profiles of physiological activities. As compared to DNA fragments, fecal mRNA provided a better representation of zebrafish physiological status. PCB126 exposure dramatically shifted the composition of fecal zebrafish DNA and mRNA as a function of sex. The differential mRNA caused by PCB126 clearly identified the toxicological fingerprint of PCB126. In summary, this study provides preliminary clues about the potential of fecal genes (mRNA in particular) in the development of non-invasive toxicological approaches. In the future, it is expected that more works will be conducted to screen sensitive diagnostic biomarkers from feces to increase the rate and reduce the cost of ecological risk assessment.

Probiotic intervention mitigates the metabolic disturbances of perfluorobutanesulfonate along the gut-liver axis of zebrafish.

Probiotic supplementation is effective to modulate the metabolic disorders caused by perfluorobutanesulfonate (PFBS). However, the underlying mechanisms remain unclear. To this end, the present study exposed adult zebrafish to PFBS (0 and 10 µg/L), probiotics, or their binary combinations for 40 days. After the exposure, the nutritional stores, intestinal organization, and metabolic activities along the gut-liver axis were investigated. The results showed that PFBS exposure decreased the nutrient reserves significantly, especially the lipid content, which was alleviated by the probiotic administration. Intestinal mucus secretion was promoted remarkably in the presence of the probiotic, which enhanced epithelial protection against PFBS damage. Metagenomic analysis showed that PFBS alone induced gut microbial dysbiosis, which was efficiently antagonized by the probiotic bacteria. Intestinal metabolomic profiling revealed that ferroptosis occurred because of the unrestricted lipid peroxidation following PFBS exposure. However, probiotic administration prevented the ferroptotic symptoms induced by PFBS, further highlighting the beneficial effects of the probiotic on the host. In PFBS-exposed livers, high levels of bile acid metabolites (e.g., taurochenodeoxycholic acid) accumulated, implying the induction of cholestasis. Notably, probiotic addition recovered the metabolomic homeostasis under PFBS stress, probably resulting from the activation of detoxification pathways based on the pentose and glucuronate interconversion. Overall, the present study provides systematic evidence of the antagonistic interaction between PFBS and the probiotic regarding the metabolic activities along the microbe, gut and liver axis, highlighting the application values of probiotic recipe in aquaculture industry and ecological reservation.

Antagonistic interaction between perfluorobutanesulfonate and probiotic on lipid and glucose metabolisms in the liver of zebrafish.

Perfluorobutanesulfonate (PFBS) and probiotic bacteria can interact to induce hepatic hypertrophy. However, the molecular events occurring in the hypertrophic liver are still unknown. Therefore, we performed this follow-up study using adult zebrafish that were exposed for 40 days to 0 and 10 µg/L PFBS, with or without dietary supplementation of probiotic Lactobacillus rhamnosus. After PFBS or/and probiotic exposures, proteome perturbation, histological pathogenesis and glucose metabolism were investigated in the livers. Proteomic analysis showed potent intervention of PFBS or/and probiotic with hepatic functions. PFBS single exposure caused marked disturbances in lipid metabolisms, which may underlie the severe vacuolization in male liver. The addition of probiotic alleviated the lipid metabolic disorders of PFBS. Furthermore, probiotic supplementation enhanced ATP energy production using glucose in mitochondrial respiratory chain of male fish. However, PFBS alone caused remarkable increase in blood glucose level (by 2.5-fold relative to the control), underlining the onset of hyperglycemia symptom. In contrast, the liver of male fish from the coexposure group functioned appropriately, which immediately increased insulin levels by 2.2-fold to reduce the glucose accumulation in blood. In female liver, PFBS alone significantly decreased the blood glucagon concentration by 2.9-fold. The deficiency of glucagon hormone consequently contributed to the accumulation of glycogen (3.2-fold) therein. Vigorous antagonistic interaction between PFBS and probiotic was noted with respect to glucose metabolism, which restored ATP, glucose, glycogen and glucagon to the control levels. Overall, the present study finds that probiotic L. rhamnosus is efficient to mitigate the metabolic disorders of PFBS on lipid and glucose, highlighting the potential values of probiotic bacteria to protect the aquatic ecosystem.

Shift in skin microbiota and immune functions of zebrafish after combined exposure to perfluorobutanesulfonate and probiotic Lactobacillus rhamnosus.

Dysbiosis of fish skin microbiome and immunity by environmental pollutants are rarely studied in toxicological research in spite of their importance for fish health. In the present study, adult zebrafish were exposed to 0 and 10 µg/L of perfluorobutanesulfonate (PFBS) for 40 days, with or without the supplementation of probiotic Lactobacillus rhamnosus, with objectives to explore the interaction between PFBS pollutant and probiotic bacteria on skin mucosal microbiota and immune response. Amplicon sequencing analysis found that PFBS alone significantly disturbed the microbial community composition and abundance on the skin, favoring the growth of stress-tolerant bacteria (e.g., Deinococcus and Enhydrobacter genera). However, the administration of probiotic inhibited the dysbiosis of PFBS and shaped the skin microbiome in the combined exposure group. PFBS single exposure also promoted the production of mucus on the skin of male zebrafish, which may be related to the growth of Limnobacter bacteria. In contrast, probiotic supplements remarkably improved the immune functions in male skin mucus from the combined group, as evidenced by the consistent increases in lysozyme activity, immunoglobulin concentrations and peroxidase activity. Overall, the present study provides the first clue about the singular and combined effects of PFBS and probiotic on skin microbiota and immunity, highlighting the beneficial action of probiotic L. rhamnosus against PFBS stress.

Interaction between probiotic additive and perfluorobutanesulfonate pollutant on offspring growth and health after parental exposure using zebrafish.

Perfluorobutanesulfonate (PFBS) pollutant and probiotic bacteria can interact to affect the reproductive outcomes of zebrafish. However, it is still unexplored how the growth and health of offspring are modulated by the combination of PFBS and probiotic. In the present study, adult zebrafish were exposed to 0 and 10 µg/L PFBS for 40 days, with or without dietary supplementation of probiotic Lactobacillus rhamnosus. After parental exposure, the development, growth and viability of offspring larvae were examined, with the integration of molecular clues across proteome fingerprint, growth hormone/insulin-like growth factor (GH/IGF) axis, calcium homeostasis, hypothalamic-pituitary-adrenal (HPA) axis and nutrient metabolism. Parental probiotic supplementation significantly increased the body weight and body length of offspring larvae. Despite the spiking of PFBS, larvae from the combined exposure group still had longer body length. RNA processing and ribosomal assembly pathways may underlie the enhancement of offspring growth by probiotic bacteria. However, the presence of PFBS remarkably increased the concentrations of cortisol hormone in offspring larvae as means to cope with the xenobiotic stress, which required more energy production. As evidenced by the proteomic analysis, the addition of probiotic bacteria likely alleviated the energy metabolism disorders of PFBS, thus allocating more energy for the larval offspring growth from the combined group. It was noteworthy that multiple molecular disturbances caused by PFBS were antagonized by probiotic additive. Overall, the present study elucidated the intergenerational interaction between PFBS and probiotic on offspring growth and health after parental exposure.

Probiotic Lactobacillus rhamnosus modulates the impacts of perfluorobutanesulfonate on oocyte developmental rhythm of zebrafish.

Potent interaction between probiotic bacteria and perfluorobutanesulfonate (PFBS), an aquatic pollutant of emerging concern, was previously reported on reproduction of zebrafish. However, the underlying mechanism is largely unexplored. In this regard, the present study continued to focus on the interactive modes between probiotics and PFBS. Adult zebrafish were exposed for 28 days to 0 and 10 µg/L PFBS with or without dietary supplementation of probiotic Lactobacillus rhamnosus. With the relevance to fecundity outcome, a suite of reproductive indices at transcriptional, hormonal, proteomic and histological levels of biological organization were measured herein. The fecundity monitoring results showed that probiotic additive shifted the impacts of PFBS on egg spawn, gradually approaching the control level. Based on ovary histological observation, oocyte growth was significantly promoted by probiotics or/and PFBS exposures, while the presence of probiotic bacteria partially antagonized the effects of PFBS on oocyte growth. The combination of probiotics and PFBS increased the concentration of maturation inducing hormones in ovary. Despite the enhanced hormonal signals, gene transcriptions of ovarian local auto/paracrine factors were consistently decreased in all exposure groups, suggesting the blocked transition from oocyte growth phase toward oocyte maturation phase. Ovary proteomic analysis found that PFBS exposure with or without probiotic bacteria mainly affected the RNA metabolic processes, although the addition of probiotics exerted extra influences on amino acid metabolism. Overall, the present study provided more mechanistic evidence about the interactive behavior between probiotic bacteria and PFBS pollutant. Feed additive of probiotic bacteria modulated the impacts of PFBS on egg production rhythm through oocyte growth and maturation phases.