Effects of atrazine on movement, metabolism and gene expression in Pelophylax nigromaculatus larvae under global warming.

Global warming and environmental pollutants both pose a threat to the behavior and physiology of animals, but research on the combined effects of the two is limited. Atrazine, a widely used herbicide, has toxic effects on organisms. In this study, the effects of environmental concentrations of atrazine exposure (100 µg/L) for seven days on the movement, metabolism and gene expression related to motility of Pelophylax nigromaculatus larvae (GS8) were investigated under global warming. The results showed that compared to the optimal growth temperature (18 °C), atrazine treatment under global warming (21 °C) significantly increased the average speed (about 11.2 times) and maximum acceleration (about 1.98 times) of P. nigromaculatus larvae, altered the relative abundance of 539 metabolites, including Formyl-5-hydroxykynurenamine, 2,4-Dihydroxybenzophenone, and FAPy-adenine, and changed the nucleotide metabolism, pyrimidine metabolism, glycerophospholipid metabolism, and purine metabolism, as well as increased the gene expression of SPLA2 (about 6.46 times) and CHK (about 3.25 times). In summary, atrazine treatment under global warming caused metabolic disorders in amphibian larvae and increased the expression of some movement-related genes in the brain, resulting in abnormally active.

Integrated physiological, intestinal microbiota, and metabolomic responses of adult zebrafish (Danio rerio) to subacute exposure to antimony at environmentally relevant concentrations.

The available information regarding the impact of antimony (Sb), a novel environmental pollutant, on the intestinal microbiota and host health is limited. In this study, we conducted physiological characterizations to investigate the response of adult zebrafish to different environmental concentrations (0, 30, 300, and 3000 µg/L) of Sb over a period of 14 days. Biochemical and pathological changes demonstrated that Sb effectively compromised the integrity of the intestinal physical barrier and induced inflammatory responses as well as oxidative stress. Analysis of both intestinal microbial community and metabolome revealed that exposure to 0 and 30 µg/L of Sb resulted in similar microbiota structures; however, exposure to 300 µg/L altered microbial communities' composition (e.g., a decline in genus Cetobacterium and an increase in Vibrio). Furthermore, exposure to 300 µg/L significantly decreased levels of bile acids and glycerophospholipids while triggering intestinal inflammation but activating self-protective mechanisms such as antibiotic presence. Notably, even exposure to 30 µg/L of Sb can trigger dysbiosis of intestinal microbiota and metabolites, potentially impacting fish health through the "microbiota-intestine-brain axis" and contributing to disease initiation. This study provides valuable insights into toxicity-related information concerning environmental impacts of Sb on aquatic organisms with significant implications for developing management strategies.

Highly efficient LaMO3 (M = Co, Fe) perovskites catalyzed Fenton's reaction for degradation of direct blue 86.

Perovskite-structured catalysts LaMO3 (M = Co, Fe) were successfully synthesized and attempted to catalyze hydrogen peroxide (H2O2) for the degradation of Direct Blue 86 (DB86), a carcinogenic phthalocyanine dye. The heterogeneous Fenton-like reaction revealed that the oxidative power of the LaCoO3-catalyzed H2O2 (LaCoO3/H2O2) process was higher than that of LaFeO3/H2O2. When LaCoO3 was calcined at 750 °C for 5 h, 100 mg/L of DB86 could be completely degraded within 5 min via LaCoO3/H2O2 system under H2O2 0.0979 mol/L, initial pH 3.0, LaCoO3 0.4 g/L, and 25 °C. The oxidative LaCoO3/H2O2 system has a low activation energy (14.68 kJ/mol) for DB86 degradation, indicating that it is a fast reaction process with highly favorable at high reaction temperatures. For the first time, a cyclic reaction mechanism of catalytic LaCoO3/H2O2 system was proposed based on the evidence of coexisting CoII and CoIII on the LaCoO3 surface and the presence of HO• radicals (major), O2•- radicals (minor), and 1O2 (minor). LaCoO3 perovskite catalyst was reusable and still maintained reactive with a satisfactory degradation efficiency within 5 min even after five consecutive uses. This study shows that the as-prepared LaCoO3 is a highly efficient catalyst for phthalocyanine dye degradation.

Effects of antimony stress on growth, structure, enzyme activity and metabolism of Nipponbare rice (Oryza sativa L.) roots.

Some antimony (Sb) contaminated areas are used for rice cultivation in response to economic demands. However, little is known about the effects of Sb stress on the growth and metabolism of rice roots. Thus, a hydroponic experiment was carried out on the growth, root anatomy, enzyme activity, and metabolism of Nipponbare rice (Oryza sativa L. ssp. japonica cv. Nipponbare) under varying levels of Sb (III) stress (0 mg L-1, 10 mg L-1, and 50 mg L-1). With the increase of Sb concentration, rice root length and root fresh weight declined by 67.8 % and 90.5 % for 10 mg L-1 Sb stress and 94.1 % and 98.4 % for 50 mg L-1 Sb stress, respectively. Anatomical analysis of cross-sections of Sb-treated roots showed an increase in cell wall thickness and an increase in the number of cell mitochondria. The 10 mg L-1 and 50 mg L-1 Sb stress increased the activity of enzyme superoxide dismutase (SOD) in root cells by 1.94 and 2.40 times, respectively. Compared to the control, 10 mg L-1 Sb treatment increased the activity of catalase (CAT) and peroxidase (POD), as well as the concentrations of antioxidant glutathione (GSH) in the root by 1.46, 1.38, and 0.52 times, respectively. However, 50 mg L-1 Sb treatment significantly decreased the activity or content of CAT, POD and GSH by 28.1 %, 13.5 % and 28.2 %, respectively. Nontargeted LC/MS-based metabolomics analysis identified 23 and 13 significantly differential metabolites in rice roots exposed to 10 mg L-1 and 50 mg L-1 Sb, respectively, compared to the control. These differential metabolites were involved in four main metabolic pathways including the tricarboxylic acid cycle (TCA cycle), butanoate metabolism, alanine, aspartate and glutamate metabolism, and alpha-linolenic acid metabolism. Taken together, these findings indicate that Sb stress destroys the structure of rice roots, changes the activity of enzymes, and affects the metabolic pathway, thereby reducing the growth of rice roots and leading to toxicity.

Assessment of rhizosphere bacterial diversity and composition in a metal hyperaccumulator (Boehmeria nivea) and a nonaccumulator (Artemisia annua) in an antimony mine.

AIMS: Heavy metal hyperaccumulators are widely used in mining restoration due to their ability to accumulate and transport heavy metals, compared to nonaccumulators. Rhizosphere bacteria in metal hyperaccumulators play a key role in the uptake of heavy metals from soil; however, assessments of the differences of rhizosphere bacteria between metal hyperaccumulators and nonaccumulator are scarce. METHODS AND RESULTS: To understand the difference of bacterial composition between hyperaccumulator and nonaccumulator in rhizosphere, the diversity and composition of rhizosphere bacteria in a metal hyperaccumulator (Boehmeria nivea) and a nonaccumulator (Artemisia annua) grown in the same field in Xikuangshan were evaluated using Illumina Miseq high-throughput sequencing technology. Boehmeria nivea and A. annua had 3926 overlapping OTUs, 19,736 and 17,579 unique OTUs, respectively. Boehmeria nivea had lower Chao1 index, Shannon index and Pielou index than A. annua. The dominant phyla and genera of rhizosphere bacteria in B. nivea and A. annua were similar, but some rhizosphere bacterial communities with heavy metal remediation ability mainly appeared in the rhizosphere of the hyperaccumulator. Compared to A. annua, B. nivea showed a significantly higher relative abundance of rhizosphere bacteria, such as Acidobacteria and Bacteroidete at the phylum level and RB41 at the genus level. Some specific rhizosphere bacteria with the ability to bind metal, such as Leifsonia and Kibdelosporangium, were only found in the rhizosphere of B. nivea. CONCLUSION: Results indicated that B. nivea, as a metal hyperaccumulator, has a key function in governing metal-resistant rhizosphere bacteria in response to antimony compound pollution stress. SIGNIFICANCE AND IMPACT OF STUDY: Understanding the diversity of rhizosphere bacteria between hyperaccumulators and nonaccumulators is beneficial to formulate strategies to improve metal uptake efficiency by selecting specific plant species and rhizosphere bacteria grown on polluted soil.

Effect of Lead Exposure on Jumping Ability in Pelophylax nigromaculata.

Jumping ability determines the likelihood of survival and reproduction of amphibians. To study the toxic effect of lead (Pb) on the jumping ability of amphibians, Pelophylax nigromaculata were treated with 0, 1 and 10 mg/L Pb for 10 days and 20 days each. The results showed that for both treatment time (10 days and 20 days), compared to the control group, the 1 mg/L Pb treatment had no significant effect on jumping parameters. Compared to the control group, the maximum jumping height, distance, speed and acceleration in the 10-day 10 mg/L Pb treatment group increased significantly, but after 20 days the jumping parameters decreased significantly. With an increase in duration and concentration, the correlation coefficient and the correlation between these jumping parameters decreased. Our results indicate that an exposure to 10 mg/L Pb for 20 days has a significant negative effect on the jumping ability of amphibians.

Chronic effects of environmentally-relevant concentrations of lead in Pelophylax nigromaculata tadpoles: Threshold dose and adverse effects.

Lead (Pb) is a common heavy metal in the natural environment, but its concentration has been increasing alongside widespread industrial and agricultural development in China. The dark-spotted frog Pelophylax (formerly Rana) nigromaculata (Anura: Ranidae) is distributed across East Asia and inhabits anthropogenic habitats such as farmland. Here, P. nigromaculata tadpoles (Gosner stage 19-46) were exposed to Pb at different concentrations (0, 40, 80, 160, 320, 640 and 1280µg/L) and Pb-induced survival, metamorphosis time, development, malformations, mobility and gonad structure were monitored. The results showed that above the threshold concentration of Pb, adverse effects were obvious. As the concentration of Pb increased, the adverse effects on different traits followed different patterns: the effects on hindlimb length, survival rate, metamorphosis rate, total malformation rate, swimming speed and jumping speed largely exhibited a linear pattern; the effects on snout-vent length, body mass and forelimb length largely exhibited a bimodal pattern. Sex ratio and gonadal histology were not affected by Pb, suggesting that Pb is not strongly estrogenic in P. nigromaculata.

Effects of continuous and pulse lead exposure on the swimming behavior of tadpoles revealed by brain-gut axis analysis.

Lead (Pb) is present in aquatic environments with a continuous or pulse form due to the regular or irregular discharge of wastewater. These two modes of exposure result in different toxicological effects on aquatic animals. To compare the effects of Pb exposure mode on the swimming behavior of amphibian larvae, this study proposed a combination method to examine the brain-gut axis (gut bacteria, histopathology, metabolomics, and ethology) in order to evaluate the ecotoxic differences in Pelophylax nigromaculatus tadpoles (Gs 21-28) when exposed to continuous (CE100) versus pulse exposure (PE100) of environmental concentrations of Pb (100 µg/L). The results showed that: 1) CE100 significantly decreased the movement distance and swimming activity of the tadpoles compared to PE100 and the control, while there were no significant differences between the control group and PE100. 2) At the phyla level, compared to PE100, CE100 treatment significantly decreased the abundance of Actinobacteria, Firmicutes, Proteobacteria, and Bacteroidetes and increased the abundance of Fusobacteria in the gut. At the genus level, compared to PE100, CE100 significantly increased the abundance of U114 and decreased the abundance of Anaerorhabdus, Exiguobacterium and Microbacterium. 3) Compared to PE100, CE100 changed the metabolites of the brain-gut axis pathway, such as quinolinic acid, L-valine, L-dopa, L-histidine, urocanic acid, L-threonine, γ-aminobutyric acid (GABA), L-glutamate (Glu), acetylcholine (Ach), L-tyrosine (Tyr), L-tryptophan (Trp), and levodopa (DOPA). 4) CE100 and PE100 played a repressive role in the histidine metabolism and tyrosine metabolism pathways and played a promoting role in the purine metabolism and pyrimidine metabolism pathways. This study provides a method for evaluating the toxic effects of heavy metal exposure via two different exposure modes (pulse versus continuous) which tadpoles may encounter in the natural environment from a combined study examining the brain-gut axis.

Endocrine system, cell growth and death, and energy metabolism induced by Sb(III) exposure in earthworm (Pheretima guillemi) revealed by transcriptome and metabolome analysis.

Antimony (Sb) is known for its severe and extensive toxicity, and earthworms are considered important indicator organisms in soil ecosystems. Therefore, the present study investigated the mechanism of toxicity of the Sb at different concentrations (50, 200 mg/kg) on earthworms using biochemical indicators, pathological sections, as well as metabolomics and transcriptomics analyses. The results showed that as the exposure concentration increased, both the antioxidant system of earthworms, extent of intestinal damage, and their metabolomic characteristics were significantly enhanced. In the 50 and 200 mg/kg Sb treatment group, 30 and 177 significant differentially changed metabolites (DCMs) were identified, respectively, with the most DCMs being down- and up-regulated, respectively. Metabolomics analysis showed that the contents of dl-tryptophan, glutamic acid, glycine, isoleucine, l-methionine, involved in the protein digestion and absorption as well as aminoacyl-tRNA biosynthesis were significantly up-regulated under the 200 mg/kg treatment. At the transcriptional level, Sb mainly affected the immune system, nervous system, amino acid metabolism, endocrine system, and carbohydrate metabolism in earthworms. The integration of transcriptomic and metabolomic data indicated that high doses of Sb regulated the metabolites and genes related to the oxidative phosphorylation pathway in earthworms. Overall, these results revealed global responses beyond the scope of conventional toxicity endpoints and facilitated a more in-depth and comprehensive assessment of the toxic effects of Sb.

Influences of continuous and pulse atrazine exposure on intestinal flora and metabolites of Pelophylax nigromaculatus tadpoles.

Atrazine, a widely used herbicide, has adverse effects on the growth and metabolism of amphibians. Due to the cyclical application use of the pesticide atrazine in agricultural production, atrazine concentrations in water occur in the form of pulses. However, knowledge of the effects of atrazine pulse exposure on the gut microbiota and metabolism of amphibians is limited. In this study, Pelophylax nigromaculatus tadpoles (Gs 26) were exposed to continuous and pulse atrazine (100 µg/L) for 60 days. The results showed that continuous exposure and pulse exposure had different effects on the diversity of gut microbiota. At the phyla level, pulse exposure significantly increased the relative abundance of Actinobacteria, and decreased the relative abundance of Firmicutes compared to continuous exposure. At the genus level, continuous and pulse exposure to atrazine significantly altered the relative abundance of Acetobacterium, Microbacterium, Bacteroides, Eulopiscium and Leuconostoc. Compared to continuous exposure, pulse exposure significantly increased the relative abundance of Microbacterium, and significantly decreased the relative abundance of Acetobacterium and Eplopiscium. In terms of metabolism, pulse exposure significantly increased the relative abundance of creatine, guanine, and inosine and significantly decreased the relative abundance of 3-hydroxysebacic acid, ganoderic acid F, hypoxanthine, and withaperuvin H compared to continuous exposure. Continuous and pulse exposure to atrazine significantly altered the relative abundance of metabolites of the pymidine metabolism, purine metabolism, beta-alanine metabolism and other pathways in the gut of P. nigromaculatus tadpoles. In addition, changes in most metabolites had a significant correlation with changes in gut microorganisms. In conclusion, our study confirmed that pulse exposure to atrazine has a greater effect on the composition of the gut microflora and the metabolism of P. nigromaculatus tadpoles than continuous exposure.

Alleviation of metal stress in rape seedlings (Brassica napus L.) using the antimony-resistant plant growth-promoting rhizobacteria Cupriavidus sp. S-8-2.

This study investigated an effective strategy for remediating antimony (Sb)-contaminated soil using the bacterial strain screened from Sb-contaminated fern rhizospheres due to its superior growth-promoting, heavy-metal(loid) resistant, and antibiotic-tolerant characteristics. The strain that belongs to Cupriavidus sp. was determined by 16S rRNA sequencing and showed no morphological changes when grown with high concentrations of Sb (608.8 mg/L). The strain showed prominent indole acetic acid (IAA), phosphate-solubilizing abilities, and ACC deaminase activity under Sb stress. Moreover, IAA and soluble phosphate levels increased in the presence of 608.8 mg/L Sb. Inoculation of rape seedlings with Cupriavidus sp. S-8-2 enhanced several morphological and biochemical growth features compared to untreated seedlings grown under Sb stress. Inoculation of Cupriavidus sp. S-8-2 increased root weight by more than four-fold for fresh weight and over two-fold for dry weight, despite high environmental Sb. The strain also reduced Sb-mediated oxidative stress and malondialdehyde contents by reducing Sb absorption, thus alleviating Sb-induced toxicity. Environmental Scanning Electron Microscope (ESEM) imaging and dilution plating technique revealed Cupriavidus sp. S-8-2 is localized on the surface of roots. Identifying the Sb-resistant plant growth-promoting bacterium suggested its usefulness in the remediation of contaminated agricultural soil and for the promotion of crop growth. We highly recommend the strain for further implementation in field experiments.

Ecotoxicological Differences of Antimony (III) and Antimony (V) on Earthworms Eisenia fetida (Savingy).

In this study, we assessed the acute and chronic toxic effects of Sb (III) and Sb (V) on Eisenia fetida (Savingy) (E. fetida) by applying the filter paper contact method, aged soil treatment, and avoidance test experiment. In the acute filter paper contact test, the LC50 values for Sb (III) were 2581 mg/L (24 h), 1427 mg/L (48 h), and 666 mg/L (72 h), which were lower than Sb (V). In the chronic aged soil exposure experiment, when the Sb (III)-contaminated soil was aged 10 d, 30 d, and 60 d after exposure for 7 d, the LC50 value of E. fetida was 370, 613, and >4800 mg/kg, respectively. Compared to Sb (V) spiked soils aged only for 10 d, the concentrations causing 50% mortality significantly increased by 7.17-fold after 14 days of exposure in soil aged for 60 d. The results show that Sb (III) and Sb (V) could cause death and directly affect the avoidance behavior of E. fetida; yet, the toxicity of Sb (III) was higher than that of Sb (V). Consistent with the decrease in water-soluble Sb, the toxicity of Sb to E. fetida was greatly reduced with time. Therefore, in order to avoid overestimating the ecological risk of Sb with varying oxidative states, it is important to consider the forms and bioavailability of Sb. This study accumulated and supplemented the toxicity data, and provided a more comprehensive basis for the ecological risk assessment of Sb.

Effects of short-term continuous and pulse cadmium exposure on gut histology and microbiota of adult male frogs (Pelophylax nigromaculatus) during pre-hibernation.

Cadmium (Cd) is an environmental endocrine-disrupting pollutant which mainly occurs in pulsed manner in natural waters, while traditional toxicology experiments have less examined the effects of pulsed exposure. Here, we studied the effects of short-term (7 days) continuous and pulse exposure to 100 µg/L Cd on gut morphology and microbiota of frogs (Pelophylax nigromaculatus) during pre-hibernation. Compared to continuous exposure, Cd pulse exposure significantly increased individual mortality and decreased the villi height and the ratio of villi height to crypt depth of the gut. Cd continuous and pulse exposure both changed the community structure and relative abundance of intestinal microbiota. Compared to continuous exposure, Cd pulse exposure significantly decreased the relative abundance of beneficial bacteria (e.g., Cetobacterium and Aeromonas genus), and significantly increased the relative abundance of harmful bacteria (e.g., Parabacteroides, Odoribacter, and Acinetobacter genus). This study shows that the gut histology and microbiota of amphibians during pre-hibernation are more susceptible to Cd pulse exposure than continuous exposure.

Short-term continuous and pulse Pb exposure causes negative effects on skin histomorphological structure and bacterial composition of adult Pelophylax nigromaculatus.

Lead (Pb) is a toxic heavy metal often present in the environment as a pulse in water. Traditional toxicity tests are usually carried out under conditions of continuous concentration, without considering the impact of pulse exposure on aquatic organisms. This study aimed to evaluate the effects of short-term continuous and pulse Pb exposures on the skin bacteria and histomorphological structure of Pelophylax nigromaculatus. Results showed that compared to the control (CON) and Pb continuous exposure group (CEPb), the Pb pulse exposure group (PEPb) showed the smallest size of granular glands, which would interfere with the permeability and secretory function of skin, making the individual more sensitive to external pollution. Lead exposure significantly changed the composition and diversity of skin bacteria. Compared to the CON and CEPb groups, the PEPb group showed a significant increase in the abundance of harmful bacteria (e.g., Bacteroidetes and Chryseobacterium) and a decrease in the abundance of beneficial bacteria (e.g., Pseudomonas). PICRUSt software showed that there were differences in the metabolic pathway of skin bacteria among the three groups (CON, CEPb, and PEPb). Overall, this study indicates that Pb pulse exposure can aggravate the toxicity of Pb for frog skin, providing a new framework for simulating short-term heavy metal exposure in the context of frog health.

Diversity and composition of soil bacteria between abandoned and selective-farming farmlands in an antimony mining area.

Background and aims: Land abandonment and selective farming are two common management methods to restore the soil conditions of low-pollution farmland in mining areas. The soil bacterial community plays an important role in farmland soil restoration; however, few studies have compared the composition and diversity of soil bacteria between the abandoned farmlands (AFS) and selective-farming farmlands (FFS). Here, the effects of AFS and FFS on soil properties and bacterial diversity were evaluated in an antimony (Sb) mining area in southern China. This study aimed to identify effective land management methods in terms of positive or negative changes in soil environment and bacterial diversity. Methods: 16S rRNA high-throughput sequencing was used to compare the diversity and composition of soil bacteria between AFS and FFS in the Xikuangshan (the largest Sb mine in the world). Results: Compared to AFS, FFS had higher Sb concentration and nutritional properties (e.g., available N, P, and K) and lower Zn concentration (p < 0.05). The bacterial alpha diversity including Chao1 index, Simpson index, Shannon index and Pielou-e index in FFS was higher than AFS (p < 0.05). At the phylum level, FFS had higher relative abundances of Chloroflexi, Acidobacteria, Gemmatimonadetes, and Rokubacteria, and lower relative abundances of Firmicutes, Actinobacteria, and Bacteroidetes. At the genus level, FFS had higher relative abundances of Acidothermus, Haliangium, and Rokubacteriales, and lower relative abundances of Bacillus, Rhodococcus, Sphingomonas, and 67-14. Redundancy analysis indicated that soil heavy metal content and soil fertility were closely correlated with the soil bacterial community. Altogether, selective farming of low-pollution farmland in the mining area can improve soil properties and soil bacterial diversity.

Atrazine exposure and recovery alter the intestinal structure, bacterial composition and intestinal metabolites of male Pelophylax nigromaculatus.

The pesticide atrazine poses a potential threat to the health of frogs living in farmland areas. The exposure concentration in traditional pesticide experiments is usually constant, while pesticide pollution in actual water may fluctuate due to periodic or seasonal application. We examined the effects of different concentrations of atrazine (50, 100 and 500 µg/L) over a 14-day exposure and a 7-day recovery on intestinal histology, bacterial composition and intestinal metabolites of male Pelophylax nigromaculatus. HE staining revealed that after a 14-day atrazine exposure, the 100 µg/L and 500 µg/L groups showed obvious cysts and significantly decreased intestinal crypt depth and villus height. After a 7-day recovery, the damaged intestine in the 100 µg/L group was partially recovered, while in the 500 µg/L exposure group there was no improvement. 16S rRNA gene analysis of intestinal bacteria showed that 500 µg/L atrazine exposure significantly caused a persistent decrease in bacterial α diversity. Compared to the control and other atrazine exposure groups, the 500 µg/L group showed significant changes in the relative abundance of predominant bacteria. In addition, most dominant bacteria in the 500 µg/L recovery group showed significant differences with the 50 µg/L and 100 µg/L recovery groups. Nontargeted metabolomics profiling based on UPLC/MS analysis showed that atrazine exposure and recovery induced changes in the intestinal metabolic profile. The changes in metabolites were mainly related to purine/pyrimidine metabolism, glycine, serine and threonine metabolism, and arginine and proline metabolism. In general, these pathways were closely related to energy metabolism and amino acid metabolism. These results suggest that the short-term exposure to 500 µg/L atrazine causes persistent harm to intestinal health. This study is an important step toward a better understanding of the toxic effects of atrazine exposure and recovery in frog intestines.

Toxicity of Pb continuous and pulse exposure on intestinal anatomy, bacterial diversity, and metabolites of Pelophylax nigromaculatus in pre-hibernation.

Amphibians are often exposed to pulse pollution due to fluctuational inputs of pollutants in water. Traditional ecotoxicology research is mainly performed under constant exposure conditions, which is not consistent with the true environmental pollution. Frogs are sensitive to changes in water pollutants in pre-hibernation. Thus, to understand the toxicity difference to continuous and pulse exposure in environmental concentrations of Pb (100 µg/L), Pelophylax nigromaculatus adults were exposed to short-term treatments (8 days) in pre-hibernation. Individual mortality, intestinal anatomical structure, bacterial diversity, and metabolites were measured in a control group (CON), a Pb continuous treatment group (CEPb) and a Pb pulse treatment group (PEPb). The results showed that PEPb significantly increased individual mortality, compared to the control group and CEPb. PEPb induced pathological changes in the small intestinal tissues, such as mucosal erosion, swollen and distorted villi, large vacuoles, and the proliferation of goblet cells. In addition, PEPb altered the structure and diversity of intestinal bacteria, resulting in an increase in some pathogenic bacteria (e.g. Bacteroides and Ruminococcus) and a decrease in beneficial bacteria (e.g. Cetobacterium and Akkermansia). Both CEPb and PEPb significantly changed intestinal metabolites and metabolic pathways. Moreover, PEPb has a significant effect on the metabolism of amino acids by increasing the content of 5-Aminopentanoic acid, cis-4-Hydroxy-l-proline, Glycocholic acid, l-Alanine, and l-Isoleucine. We concluded that PEPb may lead to intestine impairment of P. nigromaculatus in pre-hibernation by inducing intestinal structural integrity destruction, bacterial imbalance, and metabolic dysfunction, resulting in a significant increase in mortality. The study provides new insights for understanding the intestinal responses of frogs to pulse metal exposure.

The effect of atrazine on intestinal histology, microbial community and short chain fatty acids in Pelophylax nigromaculatus tadpoles.

The intestine is the main organ for nutrient absorption in amphibians. It is sensitive to atrazine, which is a herbicide widely used in agricultural areas. At present, there is a lack of systematic research on the effect of atrazine on the amphibian intestine. In this study, we evaluated the effects of atrazine exposure (0, 50 µg/L, 100 µg/L, and 500 µg/L) for 20 days on intestinal histology, microbiota and short chain fatty acids in Pelophylax nigromaculatus tadpoles. Our research showed that 500 µg/L atrazine exposure significantly decreased the height of microvilli and epithelial cells, and altered the composition and diversity of intestinal microbiota in P. nigromaculatus tadpoles compared to the control. At the phylum level, the abundance of Bacteroidetes and Fusobacteria increased significantly, while that of Verrucomicrobia and Firmicutes decreased significantly in the 500 µg/L atrazine treatment group. At the genus level, Akkermansia and Lactococcus had significantly lower abundance in the 100 µg/L and 500 µg/L atrazine exposure group, while Cetobacterium was only detected in the 100 µg/L and 500 µg/L atrazine treated group. Also, function prediction of intestinal microbiota showed that atrazine treatment significantly changed the metabolism pathways of P. nigromaculatus tadpoles. In addition, 500 µg/L atrazine exposure changed the content of short chain fatty acids by significantly increasing the content of total SFCAs, butyric acid, and valeric acid, and decreasing the content of isovaleric acid in the intestine. Taken together, atrazine exposure could affect the intestinal histology and induce intestinal microbiota imbalance and metabolic disorder in amphibian tadpoles.

Effects of atrazine short-term exposure on jumping ability and intestinal microbiota diversity in male Pelophylax nigromaculatus adults.

Atrazine, a common chemical pesticide, has toxicity to adult and juvenile amphibians in natural ecosystems; however, it is more common to study its effects on larvae instead of adults. This study assessed the impacts of atrazine in water through short-term exposure (7 days) on male black spotted frog (Pelophylax nigromaculatus) adults fed every day. The jumping ability, including jumping height, distance, time, and speed, was measured by 3D motion analysis software, and the intestinal content microbiota was determined by 16S rRNA amplicon sequencing with QIIME software. The results showed that male P. nigromaculatus exposure to 200 and 500 µg/L atrazine significantly increased jumping distance and jumping time compared to control groups. Conversely, 500 µg/L atrazine treatments significantly decreased the diversity and changed the composition and structure of intestinal content microflora in male P. nigromaculatus compared to control groups. At the phylum level, Chlamydiae was only detected in the control group, and Actinobacteria, Bacteroidetes, Firmicutes, Fusobacteria, and Proteobacteria were the dominant microflora in the atrazine treatment groups. At the genus level, the abundance of Lactobacillus and Weissella significantly increased in atrazine treatment groups compared to control groups. This study can provide a new framework based on movement behavior and intestinal microbiota to evaluate the response of amphibians to short-term exposure to environmental pollution.

Changes in diversity and composition of rhizosphere bacterial community during natural restoration stages in antimony mine.

BACKGROUND: Open pit antimony (Sb) mining causes serious soil pollution, and phytoremediation is a low-cost approach to remediate heavy metal contaminated soil. Rhizosphere bacteria play an important role in ecological restoration in mining areas. There is a knowledge gap on how to find suitable rhizosphere microorganisms to improve the phytoremediation effect. Understanding the differences of rhizosphere bacterial diversity in different restoration stages is helpful to find suitable bacteria for ecological restoration. METHODS: A method of the substitution of "space" for "time" was used to study the effect of natural restoration on rhizosphere bacterial community. According to the dominant vegetation types (herb, shrub, and tree) in the natural restoration area of Sb mining, the early restoration (ER), middle restoration (MR), and later restoration (LR) from the largest Sb mine (Xikuangshan mine) in the world were selected to evaluate the differences in the composition and diversity of rhizosphere bacteria during three natural restoration stages. Each restoration stage had five samples. To determine the relationship between restoration stages and bacterial diversity in the rhizosphere, high throughput sequencing of PCR amplified were used. RESULTS: Alpha diversity, as assessed by Chao indices, appeared lowest in ER but this trend was not seen with other diversity metrics, including the Simpson and Shannon. Beta diversity analysis suggested there were differences in rhizobacterial community structure associate with restoration stage. At the phylum level, natural restoration led to a significant increase in the relative abundance of Actinobacteria in the MR, and a significant decrease in the relative abundance of Patescibacteria in the LR. Additionally, Calditrichaeota, Deferribacteres and Epsilonbacteraeota were only found in ER. At the genus level, the relative abundance of RB41 and Haliangium were highest in LR plots, while that of Bacillus and Gaiella were highest in ER plots. Additionally, the Azorhizobium genus was only detected in the ER phase. Overall, our findings suggested that several rhizosphere microbial communities had significant differences among three natural restoration stages (ER, MR, and LR) and the rhizosphere bacterial communities mainly appeared in the early restoration stage can be preferred for remediation of pollution soil in Xikuangshan.