Studies On Metabolome, Colonization Of Tetrahydrofuran-degrading Rhodococcus Sp. YYL And Its Interaction With Bacillus Cereus HZX

Tetrahydrofuran (THF) was widely used and a kind of persisent environmental pollutants. Microbial degradation was one of the main mechanisms responsible for the mineralization of THF pollutant. With the aims of studying tetrahydrofuran degradation mechanisms by strain Rhodococcus sp. YYL, we focused on assessing the basal metabolism in strain YYL in the presence of THF, studying the effects of trehalose on THF degradation by strain YYL and colonization in sequencing batch reactor (SBR) system treating THF, and illustrating the interaction between strain YYL and Bacillus cereus HZX during THF degradation process. The main results were as following:(1) Nuclear magnetic resonance (NMR)-based metabolomics analyses were performed to explore the metabolic profiling response of strain YYL exposing to THF. Exposure to THF slightly influenced the metabolome of strain YYL when yeast extract was provided in the medium, which means no obvious THF toxicity to strain YYL. The metabolic profile changes of strain YYL over time were also investigated using THF as the sole carbon source to identify the metabolites associated with high-efficiency THF degradation. Strain YYL could accumulate several disturbances in osmoregulation (trehalose), accelerate tricarboxylic acid cycle (TCA) (from succinate to malic acid) and enhance protein, nucleotide synthesis.As an osmoregulation factor, trehalose accumulation was positively correlated with THF degradation efficiency. Then it was added into the cultural medium to investigate whether an appropriate amount of trehalose could promote THF degradation. The results indicated that trehalose played much more important roles in protecting strain YYL under the initial pH 7.0 than that in the optimal initial pH 8.26. 2 mM Trehalose could enhance mRNA expression of THF degrading monooxygenase gene (thm), enzyme activity of succinate dehydrogenase (SDH), ATPase, pyruvate kinase (PK), total superoxide dismutase (T-SOD), and glutathione (GSH) concentration.(2) Trehalose, as a biostimulant in bioaugmention system with strain YYL, could help strain YYL better colonizing in THF treating SBR. The higher abundance of strain YYL in SBRs, the lower concentration of THF existed. Lower concentration of THF resulted in higher abundance, adversity and activity of total bacteria and extracellular polymeric substance (EPS) in activated sludge.Soluble di-iron monooxygenases (SDIMOs) gene, particularly group-5 SDIMOs (SDIMO) (i.e., tetrahydrofuran and propane monooxygenases), play a significant role in the initiation of the ring cleavage of tetrahydrofuran. Cytochrome P450 Monooxygenases (P450s) genes, such as CYP153, have a significant role in catalyzing the region-and stereospecific oxidation of non-activated hydrocarbons, and THF was reported to inhibit P450s in the environment. Denaturing gradient gel electrophoresis (DGGE), clone library and quantitative PCR were employed to study abundances and diversities of genes SDIMO and CYP153.2 mM Trehalose could help activated sludge keeping higher genes SDIMO and CYP153 abundances and diversities. Therefore, trehalose as a biostimulant was helpful for enhancing the microbial community’s ability to respond to environmental stress.During the process of bioremediation, degradation of pollutants could also probably have an effect on the nitrogen removal efficiency. Biostimulation with 2 mM trehalose exhibited different impacts on the functional bacteria involving in nitrogen cycle (nitrifier/denitrifier). The results demonstrated that trehalose could help activated sludge possessing higher diversities and abundances of amoA and nirK; however, there was no obvious influence on nosZ and nirS. Generally, the higher in diversity and abundance could result in higher biological activity of gene. Biostimulation with 2mM trehalose could alleviate the negative effects of THF stress on nitrifier/denitrifier community.(3) During THF degradation, the interaction between THF-degrading strain YYL and no THF-degrading strain Bacillus cereus HZX depended on the cultural environment for strain YYL. Under the optimal initial pH 8.26, strain YYL could develop high THF-degrading ability on its own, and the presence of strain HZX didn’t exhibit beneficial effects on THF degradation. Under the adverse initial pH 7.0, high abundance of strain HZX could stimulate gene thm expression to accelerate THF degradation, which resulted in decreasing pH and higher ratio of strain HZX.According to the transcriptome analysis of strain YYL and strain HZX, strain YYL and HZX exhibited more closely relationship at pH 7.0 than that at pH 8.26 in co-culture for THF degradation. Under pH 7.0, strain HZX could acquire energy from THF degradation by strain YYL; In return, strain HZX could help/replace strain YYL to perform some physical and metabolic processes, such as amino acids biosynthesis, transformation of sugar,2-Oxocarboxylic acid metabolism and linoletic acid biosynth -esis. With the help of strain HZX, strain YYL can be in good physiology in response to environmental stress, and functions well for THF degradation.