Toxicology research focuses on the cellular principles, biochemical and molecular mechanisms of the toxic effects of chemicals on living organisms, and the mechanism research results are very important in many fields of applied toxicology. The transcriptome can study gene function and gene function at the overall level, and reveal the molecular mechanism of specific biological processes and disease occurrence. This paper uses the unique degradation ability of white rot fungi to study its degradation mechanism with different doses of toxic pollutants.
Decabromodiphenylethane (DBDPE) is a new type of brominated flame retardant widely used, DBDPE can accumulate in organisms and pose a threat to the environment and even human health. As a biotechnology tool for environmental pollution control, white rot fungi ( Pleurotus ostreatus ) belong to the basidiomycetes Agaricus agaricus, and are one of the most representative fungi among cultivated edible white rot fungi in China. substances have unique degradability. Toxic substances are the substrates of microorganisms in the process of biodegradation, and the toxic reaction of microorganisms is accompanied by it. Therefore, it is imperative to study the stress response of Pleurotus ostreatus to toxic pollutants.
Journal: Journal of Hazardous Materials
Impact factor: 10.588
Release time: 2022
1. Experimental design
1. Experimental treatment
Collect P. otreatus (CICC 50166) spore suspension, add 1 mL spore solution and 19 mL medium into the flask, and then add 1, 2, 4, 5, 10, 20 and 50 mg/L of DBDPE, respectively. The mycelia were collected at 0, 12, 24, 36, 48, 72 and 96 h, respectively, and the mycelium particles without DBDPE treatment were used as the control.
2. Research methods
After observing mycelial morphology and measuring oxidative stress, samples exposed to 0 mg/L (control), 5 mg/L (low dose) and 20 mg/L (high dose) DBDPE for 72 hours were selected for transcriptome sequencing analysis .
Figure 1 Experimental design
2. Experimental results
1. Effect of DBDPE on fungal growth
The effect of DBDPE on growth was judged by measuring the biomass of fungi and the pH value of the medium. It was found that the accumulation of metabolites and proteins of fungi after exposure for 36 h could relieve the pressure of DBDPE at a lower concentration (5 mg/L).
Fig. 2 Effect of DBDPE on fungal biomass (a) and medium pH (b)
At the same time, the change trend of Ca2+, Mg2+-ATPase and Na+, K+-ATPase activity of Pleurotus ostreatus was measured, and it was found that the enzyme activity increased significantly with the increase of concentration and reached the maximum at 24 h. The activity of SOD, CAT and GSH and the determination of MDA content also found that the high concentration of DBDPE (5-50 mg/L) had a greater impact on Pleurotus ostreatus. By inhibiting the synthesis of GSH, it promotes the continuous increase of MDA content in cells, thereby inhibiting the GSH activity of Pleurotus ostreatus and accelerating lipid peroxidation.
At 0, 5 and 20 mg/L DBDPE concentrations, the effect of DBDPE on the morphology of Pleurotus ostreatus mycelium was studied by SEM. When the exposure concentration of DBDPE reached 20 mg/L, it was observed that almost all mycelia were destroyed (Fig. 3), which may lead to the efflux of intracellular material.
Figure 3 Morphological observation of Pleurotus ostreatus exposed to DBDPE
2. Transcriptome analysis results
The results of transcriptome analysis showed that compared with the control group, there were 1193 and 1107 DEGs in the 5 and 20 mg/L DBDPE treatment groups, respectively, and the number of down-regulated genes was far more than the number of up-regulated genes (Fig. 6a, b and c). The Venn diagram shows the amount of the same DEG at different exposure concentrations of DBDPE, DBDPE can change the gene expression of Pleurotus ostreatus, and the effect of higher concentration (20 mg/L) on the fungus is obvious.
Fig.4 DEG scatter diagram of Pleurotus ostreatus treated with different DBDPE
GO analysis of DEGs (Fig. 5) showed that DEGs were significantly annotated into 50 different GO terms. Compared with the low concentration treatment group, there were more down-regulated DEGs in the high concentration treatment group. Compared with the control group, most DEGs in the low-concentration group were enriched in biological processes and cellular components, including fatty acid catabolism, purine nucleoside triphosphate biosynthesis, etc., and most DEGs exposed to high concentrations were enriched in biological processes (Fig. 6). The downregulation of these gene expression indicates that DBDPE can affect biological processes and cellular components by inhibiting electron transport, mitochondrial ATP synthesis, oxidoreductase activity, and transporter activity.
Fig. 5 GO annotations of Pleurotus ostreatus DEG in different DBDPE treatment groups
Figure 6 GO enrichment analysis of DEG in Pleurotus ostreatus treated with different DBDPE groups
KEGG enrichment analysis showed (Fig. 7) that energy metabolism (oxidative phosphorylation), global metabolism (carbon metabolism) and carbohydrate metabolism (TCA cycle) were the main metabolic pathways of Pleurotus ostreatus. As shown in Table 1, the expression of genes related to these complexes were all down-regulated, which indicated that electron transport was weakened and oxidative phosphorylation might be inhibited by DBDPE stress, thereby reducing fungal growth. It may explain the inhibition of fungal growth and the reduction of DBDPE degradation by Pleurotus ostreatus.
Table 1 The main KEGG enrichment pathways of Pleurotus ostreatus DEGs exposed to 5 and 20 mg/L DBDPE, respectively.
Figure 7 KEGG enrichment of Pleurotus ostreatus DEG in different DBDPE treatment groups
Three research conclusions
The article investigated the toxicological response and transcriptomic changes of P. otreatus exposed to DBDPE. DBDPE was found to inhibit fungal growth and ATPase activity. Transcriptomic analysis showed that DBDPE can inhibit oxidative phosphorylation, TCA cycle and carbon metabolism. These findings help to develop a biodegradable system for DBDPE.
references
Toxicity evaluation of decabromodiphenyl ethane (DBDPE) to Pleurotus ostreatus: Oxidative stress, morphology and transcriptomics. Journal of Hazardous Materials , 2022.