Effects of cadmium exposure on thyroid gland and endochondral ossification in Rana zhenhaiensis.

Discovery of elevated concentrations of cadmium in the natural environment has increased awareness because of their potential threats. Amphibians are negatively affected due to their moderate sensitivity to cadmium. Here, we conduct acute and subchronic toxicity tests to examine whether, and to what extent, cadmium exposure disturbs metamorphosis, growth, and kinetic ability of Rana zhenhaiensis. We set different concentration treatment groups for the subchronic toxicity test (0, 10, 40, 160 µg Cd L-1). Our findings demonstrate that cadmium exposure reduces growth parameters and the cumulative metamorphosis percent of R. zhenhaiensis. Decreases in follicular size and follicular epithelial cell thickness of thyroid gland are found in the treatment group. Further, subchronic exposure to cadmium decreases ossification ratio of hindlimbs in all treatment. Also, adverse effects of cadmium exposure on aquatic tadpoles can result in the reduced physical parameters and weak jumping ability in adult frogs. In this sense, our study suggests that cadmium adversely influences body condition and metamorphosis of R. zhenhaiensis, damages thyroid gland and impairs endochondral ossification. Meanwhile, we speculated that cadmium-damaged thyroid hormones inhibit skeletal development, resulting in the poor jumping ability, which probably leads to reduced survival of R. zhenhaiensis.

Altered temperature affect body condition and endochondral ossification in Bufo gargarizans tadpoles.

Bufo gargarizans is one kind of economic animals with higher medicinal value in China. In this study, B. gargarizans (Bufo gargarizans) tadpoles were reared at three different water temperature (15, 22 and 29 °C) from Gosner stages 28-46. We investigated the effects of temperature on growth, development, survival, metamorphic duration, size and skeletal ossification at Gosner stage 40, 42, and 46, as well as thyroid tissue reached metamorphic climax (Gs42). Besides, we examined the transcription levels of endochondral ossification-related genes in hind limb at metamorphic climax (Gs42). Our results showed that the growth and development of tadpoles conform to the temperature-size rule (TSR). While warm temperature resulted in the decrease in body size and hind limb length, and shorten larval period, cold temperature led to increase in body size and hind limb length but prolonged larval period. Histological examinations revealed that warm and cold temperatures caused damage to thyroid tissue. Also, warm and cold temperatures inhibited the degree of ossification with the double staining methodology. Additionally, the real-time PCR results suggested that warm and cold temperatures significantly up-regulated Runx2, VEGF and VEGFR mRNA levels, and down-regulated TRß, MMP9, MMP13 and Runx3 mRNA levels. The up-regulation of Dio2 level and down-regulation of Dio3 level were observed in warm temperature. TRα mRNA level was significantly increased in warm temperature, but decreased in cold temperature. Collectively, these observations demonstrated that warm and cold temperatures affected endochondral ossification in B. gargarizans tadpoles, which might influence their capacity to terrestrial locomotion.

Effects of temperature on growth, development and the leptin signaling pathway of Bufo gargarizans.

Climate change is one of the most important causes of the decline in amphibians. Changes in temperature have an important effect on the growth and development and energy metabolism of amphibians. The aim of this study is to unravel the effects of temperature on the leptin signaling pathway of Bufo gargarizans and its molecular mechanisms. Our results showed that high temperature accelerated the development rate of tadpoles, but reduced body size and mass, while low temperature deferred the development of tadpoles, but increased size and mass. Both high temperature and low temperature exposure caused pathological damage of the liver in B. gargarizans. The results of RT-qPCR revealed that the high temperature treatment significantly upregulated the transcript levels of genes related to thyroid hormone (DIO2 (D2), Thyroid Hormone Receptor-α (TRα)) and the leptin signaling pathway (Leptin Receptor (LepR), Janus kinase 1 (JAK1), Janus kinase 2 (JAK2), Tyrosine kinase 2 (TYK2), Signal Transducer And Activator Of Transcription 3 (STAT3), Signal Transducer And Activator Of Transcription 3.1 (STAT3.1), and Signal Transducer And Activator Of Transcription 6 (STAT6)), while there was a decrease of mRNA expression of these genes (TRα, Thyroid Hormone Receptor-Beta (TRß), LepR, JAK1, and TYK2) in the liver of tadpoles exposed to high temperature compared with the intermediate temperature treatment. Therefore, our results suggested that temperature extremes might interfere with the thyroid and leptin signaling pathways and affect the growth and development of B. gargarizans. Furthermore, tissue injury of the liver could occur due to exposure to temperature extremes. This work promotes public awareness of environmental protection and species conservation needs, also provides valuable experimental data and a theoretical basis for the protection of amphibians.

Impacts of earthworm introduction and cadmium on microbial communities composition and function in soil.

Heavy metal contamination of soil has become a public concern. Earthworms are key players in the functioning and service of soil ecosystems, with comprehension of their introduction in the polluted soil offering new insights into the protection of soil resources. In the present study, we evaluated the effects of earthworm (Eisenia fetida) introduction and Cd (0, 10, 30, and 60 mg kg-1 of Cd) exposure upon soil microbial community using 16S rRNA gene amplicon sequencing. Our research demonstrated that Gemmatimonadetes and Deinococcus-Thermus upregulated significantly, while Chryseolinea showed an obvious decreasing trend after earthworm introduction. In Cd contaminated soil, many genera exhibited a greater presence of Cd-dependent bacteria, namely Cd-tolerant bacteria such as Altererythrobacter and Luteimonas, and a decrease of sensitive bacteria, such as Amaricoccus and Haliangium. Moreover, functional prediction analysis of soil microbiota indicated that earthworm introduction and Cd exposure changed functional pathways of soil microorganisms. The results obtained in this study are beneficial for understanding soil microbial community impacted by earthworm, and for exploring Cd resistant or tolerant bacteria, with potentially significant findings for soil biodiversity and Cd bioremediation.

Comparison of the characteristics of intestinal microbiota response in Bufo gargarizans tadpoles: Exposure to the different environmental chemicals (Cu, Cr, Cd and NO3-N).

Environmental pollutants could change the intestinal microbiota communities, while data concerning the dynamics of the intestinal microbiota in response to different environmental chemicals in amphibian are lacking. We compared the effects of Cu, Cd, Cr and NO3-N on intestinal microbiota of B. gargarizans tadpoles by using high-throughput 16S rRNA sequencing technology. Our results revealed that responses of intestinal microbiota to three metals and NO3-N showed different characteristics. At the phylum level, the most 100 OTUs were predominantly colonized by Proteobacteria, and meanwhile, expansion of Proteobacteria was observed in Cu 64 µg/L, Cd (100 µg/L and 200 µg/L) and NO3-N100 mg/L treatment groups. In addition, the abundance of Bacteroidetes significantly increased in the gut administrate with Cu, Cd, Cr, NO3-N 20 mg/L exposures, while declined abundance of Fusobacteria was observed in Cu64 µg/L Cd100 µg/L Cd200µg/L-exposed groups. At the genus level, several genera exhibited increased prevalence of abundance such as Shewanella, Azospira and Flavobacterium. The functional prediction revealed that exposures of three metals and NO3-N increase the risks of metabolic disorders and diseases. Our research could be an important step toward an assessment of the ecological risks of different chemicals to aquatic organisms using intestinal microbiota.