Zhenmei Lv

Bioaugmentation with the nicotine-degrading bacterium Pseudomonas sp. HF-1 in a sequencing batch reactor treating tobacco wastewater: degradation study and analysis of its mechanisms.

The highly effective nicotine-degrading bacterium Pseudomonas sp. HF-1 was augmented in an SBR system that is used to treat tobacco wastewater. Compared to the non-bioaugmented (non-BA) system, the bioaugmented (BA) system exhibited considerably stronger pollution disposal abilities, with 100% nicotine degradation and more than 84% chemical oxygen demand (COD) removal within 12h. Nicotine degradation had a significant effect on COD removal in SBRs (r=0.928, p<0.01). The mechanisms of bioaugmentation were systematically investigated using a combination of polymerase chain reaction and denaturing gradient gel electrophoresis (PCR-DGGE) and a toxicity assay (protein carbonyl (PC) and DNA-protein crosslinking (DPC)). DGGE fingerprint profiles showed that the number of bands and the Shannon-Wiener index decreased at a nicotine load of 250mg/L compared to a 40-130mg/L nicotine load in the non-BA system. However, a stepwise increase in the Shannon-Wiener index was found during all periods in the BA system. A comparison of sequences excised from DGGE gels demonstrated significant differences in the dominant microbial species between the two SBRs. This result suggested that bioaugmentation of strain HF-1 could select cooperators for treating complicated tobacco wastewater. The PC content and the DPC coefficient increased significantly at levels higher than 80mg/L in the non-BA system; nevertheless, no increase was observed in the BA system during the stepwise nicotine load. This indicated that bioaugmentation of strain HF-1 resulted in the maintenance of high treatment activity by minimizing the nicotine toxicity for other microbes in the BA system. In conclusion, the rapid nicotine degradation of strain HF-1 performed a vital function in SBR by influencing the microbial community structure, dynamics and activity of the activated sludge system.

Isolation, identification and characterization of a novel Rhodococcus sp. strain in biodegradation of tetrahydrofuran and its medium optimization using sequential statistics-based experimental designs.

Statistics-based experimental designs were applied to optimize the culture conditions for tetrahydrofuran (THF) degradation by a newly isolated Rhodococcus sp. YYL that tolerates high THF concentrations. Single factor experiments were undertaken for determining the optimum range of each of four factors (initial pH and concentrations of K(2)HPO(4).3H(2)O, NH(4)Cl and yeast extract) and these factors were subsequently optimized using the response surface methodology. The Plackett-Burman design was used to identify three trace elements (Mg(2+), Zn(2+)and Fe(2+)) that significantly increased the THF degradation rate. The optimum conditions were found to be: 1.80 g/L NH(4)Cl, 0.81 g/L K(2)HPO(4).3H(2)O, 0.06 g/L yeast extract, 0.40 g/L MgSO(4).7H(2)O, 0.006 g/L ZnSO(4).7H(2)O, 0.024 g/L FeSO(4).7H(2)O, and an initial pH of 8.26. Under these optimized conditions, the maximum THF degradation rate increased to 137.60 mg THF h(-1) g dry weight in Rhodococcus sp. YYL, which was nearly five times of that by the previously described THF degrading Rhodococcus strain.

Isolation and characterization of phenanthrene-degrading Sphingomonas paucimobilis strain ZX4

Phenanthrene-degrading bacterium strain ZX4 was isolated from an oil-contaminated soil, and identified as Sphingomonaspaucimobilis based on 16S rDNA sequence, cellular fatty acid composition, mol% G + C and Biolog-GN tests. Besides phenanthrene, strain ZX4 could also utilize naphthalene, fluorene and other aromatic compounds. The growth on salicylic acid and catechol showed that the strain degraded phenanthrene via salicylate pathway, while the assay of catechol 2, 3-dioxygenase revealed catechol could be metabolized through meta-cleavage pathway. Three genes, including two of meta-cleavage operon genes and one of GST encoding gene were obtained. The order of genes arrangement was similar to S-type meta-pathway operons. The phylogenetic trees based on 16S rDNA sequence and meta-pathway gene both revealed that strain ZX4 is clustered with strains from genus Sphingomonas.

Bioaugmentation of activated sludge with Acinetobacter sp. TW enhances nicotine degradation in a synthetic tobacco wastewater treatment system.

Bioaugmentation (BA) using Acinetobacter sp. TW with high nicotine-degrading efficiency was applied in a bioreactor receiving a load of COD (3,200 ± 50 mg/L) and nicotine (1.0 ± 0.1g/L). The results showed that because of the colonization of strain TW, the COD removal was stable at 80-90%, while nicotine removal reached 98% in the BA system. Furthermore, according to PCR-DGGE fingerprinting, compared with the originally activated sludge, more bacteria existed in the BA systems while some bacteria disappeared from the non-BA system. In terms of the quorum sensing, short chain AHLs increased to assist colonization of strain TW, and long chain AHLs were secreted and helped to resist the nicotine toxicity. Compared with the non-BA system, the amounts of ROS, protein carbonyls and 8-OHdG were significant lower in the BA systems, which suggested that strain TW played an important role in eliminating the nicotine toxicity from the bioreactors.

MTH1745, a protein disulfide isomerase-like protein from thermophilic archaea, Methanothermobacter thermoautotrophicum involving in stress response

MTH1745 is a putative protein disulfide isomerase characterized with 151 amino acid residues and a CPAC active-site from the anaerobic archaea Methanothermobacter thermoautotrophicum. The potential functions of MTH1745 are not clear. In the present study, we show a crucial role of MTH1745 in protecting cells against stress which may be related to its functions as a disulfide isomerase and its chaperone properties. Using real-time polymerase chain reaction analyses, the level of MTH1745 messenger RNA (mRNA) in the thermophilic archaea M. thermoautotrophicum was found to be stress-induced in that it was significantly higher under low (50°C) and high (70°C) growth temperatures than under the optimal growth temperature for the organism (65°C). Additionally, the expression of MTH1745 mRNA was up-regulated by cold shock (4°C). Furthermore, the survival of MTH1745 expressing Escherichia coli cells was markedly higher than that of control cells in response to heat shock (51.0°C). These results indicated that MTH1745 plays an important role in the resistance of stress. By assay of enzyme activities in vitro, MTH1745 also exhibited a chaperone function by promoting the functional folding of citrate synthase after thermodenaturation. On the other hand, MTH1745 was also shown to function as a disulfide isomerase on the refolding of denatured and reduced ribonuclease A. On the basis of its single thioredoxin domain, function as a disulfide isomerase, and its chaperone activity, we suggest that MTH1745 may be an ancient protein disulfide isomerase. These studies may provide clues to the understanding of the function of protein disulfide isomerase in archaea.

Trehalose promotes Rhodococcus sp. strain YYL colonization in activated sludge under tetrahydrofuran (THF) stress.

Few studies have focused on the role of compatible solutes in changing the microbial community structure in bioaugmentation systems. In this study, we investigated the influence of trehalose as a biostimulant on the microbial community in tetrahydrofuran (THF)-treated wastewater bioaugmentation systems with Rhodococcus sp. YYL. Functional gene profile changes were used to study the variation in the microbial community. Soluble di-iron monooxygenases (SDIMO), particularly group-5 SDIMOs (i.e., tetrahydrofuran and propane monooxygenases), play a significant role in the initiation of the ring cleavage of tetrahydrofuran. Group-5 SDIMOs genes are enriched upon trehalose addition, and exogenous tetrahydrofuran monooxygenase (thmA) genes can successfully colonize bioaugmentation systems. Cytochrome P450 monooxygenases (P450s) 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. The CYP153 family was chosen as a representative P450 to study the inhibitory effects of THF. The results demonstrated that CYP153 family genes exhibited significant changes upon THF treatment and that trehalose helped maintain a rich diversity and high abundance of CYP153 family genes. Biostimulation with trehalose could alleviate the negative effects of THF stress on microbial diversity in bioaugmentation systems. Our results indicated that trehalose as a compatible solute plays a significant role for environmental strains under extreme conditions.

Influence of poisonous gases on the bio-oxidation of coalmine gas

Carbon monoxide (CO), hydrogen sulphide (H2S) and sulphur dioxide (SO2), which commonly appear and mix in coalmine gas, were investigated in terms of their effects on methane oxidation of methanotroph systems using single-factor and multi-factor experiments. The single-factor experiments that were performed showed that the methane oxidation rate and methane monooxygenase (MMO) activity of the test group at different concentrations had no evident variability when compared with the control group. Multi-factor experiments indicated that a mixture of CO, H2S and SO2 at various concentrations slightly inhibited the methane oxidation process of the methanotroph system. A pairwise comparison on the basis of sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) protein profiles demonstrated a high Jaccard index (ranging from 79 to 93%) similar to that of the methanotroph system, while the existence of CO, H2S and SO2, did not have evident effects on methane oxidation in the methanotroph system. Therefore, the possible concentrations of these gases in coalmines have no obvious influence on the methane oxidation of the methanotroph system. This system can be used effectively in the treatment of coalmine gas.

Genetic characterization and modification of a bioethanol-producing yeast strain

Yeast Saccharomyces cerevisiae strains isolated from different sources generally show extensive genetic and phenotypic diversity. Understanding how genomic variations influence phenotypes is important for developing strategies with improved economic traits. The diploid S. cerevisiae strain NY1308 is used for cellulosic bioethanol production. Whole genome sequencing identified an extensive amount of single nucleotide variations and small insertions/deletions in the genome of NY1308 compared with the S288c genome. Gene annotation of the assembled NY1308 genome showed that 43 unique genes are absent in the S288c genome. Phylogenetic analysis suggested most of the unique genes were obtained through horizontal gene transfer from other species. RNA-Seq revealed that some unique genes were not functional in NY1308 due to unidentified intron sequences. During bioethanol fermentation, NY1308 tends to flocculate when certain inhibitors (derived from the pretreatment of cellulosic feedstock) are present in the fermentation medium. qRT-PCR and genetic manipulation confirmed that the novel gene, NYn43, contributed to the flocculation ability of NY1308. Deletion of NYn43 resulted in a faster fermentation rate for NY1308. This work disclosed the genetic characterization of a bioethanol-producing S. cerevisiae strain and provided a useful paradigm showing how the genetic diversity of the yeast population would facilitate the personalized development of desirable traits.

Periodically spilled-oil input as a trigger to stimulate the development of hydrocarbon-degrading consortia in a beach ecosystem

In this study, time-series samples were taken from a gravel beach to ascertain whether a periodic oil input induced by tidal action at the early stage of an oil spill can be a trigger to stimulate the development of hydrocarbon-degrading bacteria under natural in situ attenuation. High-throughput sequencing shows that the microbial community in beach sediments is characterized by the enrichment of hydrocarbon-degrading bacteria, including Alcanivorax, Dietzia, and Marinobacter. Accompanying the periodic floating-oil input, dynamic successions of microbial communities and corresponding fluctuations in functional genes (alkB and RDH) are clearly indicated in a time sequence, which keeps pace with the ongoing biodegradation of the spilled oil. The microbial succession that accompanies tidal action could benefit from the enhanced exchange of oxygen and nutrients; however, regular inputs of floating oil can be a trigger to stimulate an in situ “seed bank” of hydrocarbon-degrading bacteria. This leads to the continued blooming of hydrocarbon-degrading consortia in beach ecosystems. The results provide new insights into the beach microbial community structure and function in response to oil spills.

Effects of tetrahydrofuran on dissolved oxygen, pH, culturable aerobic microbes and enzyme activity in wastewater

Tetrahydrofuran (THF) is easy to volatilize and hard to biodegrade. Little is known about the influence and fate of tetrahydrofuran in the environment. The effects of THF were examined on dissolved oxygen, pH, culturable aerobic microbes (bacteria, actinomycetes, and fungi) and four selected enzymes (protease, phosphatase, amylase, and lipase) in wastewater. The wastewater containing some activated sludge was taken from the aeration basin in Sibao sewage plant in Hangzhou, China. THF at lower concentrations of enhanced aerobic respiration and higher concentrations decreased utilization of oxygen in wastewater. The short-term influence of THF on pH was not obvious, but the long-term effect was apparent. Inhibition of THF on culturable aerobic microbes rose with increasing THF concentrations but decreased with prolongation of incubation time. No effect of THF on the activity of protease, phosphatase, amylase, and lipase was found. Data indicate that lower concentrations of THF exert no marked influence on wastewater in an urban sewage plant.