微生物学报最新文献

H2S is the third gaseous signaling molecule next to nitric oxide and carbon monoxide, but studies on its physiological functions in bacteria are just emerging. In this paper, we review recent findings regarding endogenous production and physiological functions of H2S in facultative anaerobic bacteria, partly based on our own research on Shewanella oneidensis. There are two principal H2S producing pathways in S. oneidensis:one is through cysteine degradation, and the other is via inorganic sulfur respiration. Endogenous H2S could either benefit mutual growing bacteria by supplying energy and inorganic, or inhibit competing bacteria. Our review attaches particular importance to the role of H2S in bacterial oxidative stress response. On one hand, H2S is able to directly inhibit heme-containing catalase, enhancing killing by H2O2. On the other hand, H2S could activate oxidative response as a signaling molecule, leading to cell protection from the oxidative stress due to elevated expression of H2O2 scavenging and repairing systems. Intriguingly, the dominance of either role is determined by H2S-treating time, that is, inhibition is the immediate response whereas activation of oxidative stress response needs extended treatment. The elucidation of endogenous production and its physiological function of H2S in facultative anaerobic bacteria would improve understanding of biogeochemical sulfur recycling, and facilitate control of infectious bacterial pathogens.

Members of the genus Microlunatus exhibit many potential advantages in managing the environmental pollution caused by phosphorus. The genus was proposed by Nakamura and co-workers with the name Microlunatus phosphovorus as the type species in 1995. Up to date, the genus Microlunatus encompasses seven validly described species, which were isolated from various environments. Members of the genus Microlunatus share the following genus-specific characteristics, possessing LL-2, 6-diaminopimelic acid in the cell wall peptidoglycan, MK-9(H4) as the predominant menaquinone and diphosphatidylglycerol and phosphatidylglycerol as the phospholipid pattern. Based on the taxonomic results of two newly isolated strains of the genus Microlunatus and the related reference reports, this review summarizes the research advances of the genus Microlunatus, including the genus establishment, taxonomic characteristics, their distribution in the environments, as well as the application prospect in chemical and medical industry.

Objective: To identify non-coding RNAs in Haloferax mediterranei through high-throughput RNA sequencing, bioinformatics analysis and molecular techniques.

Methods: After H. mediterranei cells under log phase of growth were treated with different salt concentrations for 30 minutes, total RNA was extracted for the following strand-specific RNA sequencing and differential RNA sequencing. These RNA-seq data were used to identify the genome-wide ncRNAs and to predict the 5' and 3'-ends of the transcripts by bioinformatics analysis. A few selected ncRNAs were further confirmed by Northern blotting and Circularized RNA reverse transcription-PCR analysis.

Results: We identified 105 highly credible ncRNAs. Expression of four ncRNAs showed difference in different salt concentrations. We confirmed the expression, length of transcripts, transcription start and termination sites of incRNA1436 and incRNA1903 by Northern blotting and CR-RT-PCR.

Conclusion: We identified the ncRNAs of H. mediterranei in a genome-wide scale, including identification of a few ncRNAs involved in the responses of H. mediterranei to different salt concentrations. Our results have provided fundamental data and novel insights for future study of the function of ncRNA in haloarchaea.

Objective: To establish a pipeline for unknown transcriptional start site (TSS) identification without radioactivity, we used genetic fragment analysis system and replenished two steps regarding prediction and evaluation.

Methods: We used unknown TSSs of GroEL genes from M. xanthus as a case. Firstly, we predicted the potential TSSs through bioinformatics databases. According to the prediction, we designed and synthesized fluorescence labeled primers to carry out the reverse transcription reactions. Further, we took advantage of the genetic fragment analysis system to identify TSSs with internal standards. Finally, we applied the normal distribution theory to evaluate the data.

Results: We determined the numbers, abundances and accurate sites of the TSSs:GroEL1 has one promoter and the site is TSS(286), whereas GroEL2 has two promoters, and the sites are TSS548 and TSS(502). TSS(286) is 14.3 times more abundant than TSS(548) and TSS(548) is 13.8 times more than TSS(502).

Conclusion: The bioinformatics analyzing indicates the range for the experimental design. TSS determination through genetic fragment analysis system is safer, more automatic and accurate. Normal distribution theory further refines the reliability of results. Combination of the three techniques establishes a more complete pipeline of primer extension for unknown TSS determination.

Objective: To identify and clone the polymerase Ⅲ U6 promoter from Cryptococcus neoformans (CnU6 promoter), and verify if CnU6 promoter can effectively transcribe shRNA and gRNA of CRISPR/Cas9 system.

Methods: Combining the C. neoformans genome information published in GenBank database and RNA-seq library data from our laboratory, we obtained the U6 RNA sequence with high transcriptional level by bioinformatics analysis. The putative CnU6 promoter was ligated upstream of shRNA and gRNA by EasyGeno and overlapping PCR respectively. Based on shRNA-mediated target gene silence phenotype by RNAi and gene mutation by gRNA-guided Cas9 nuclease mediated target sites editing by CRISPR/Cas9 system, we could identify if CnU6 promoter could drive the transcription of short RNA.

Results: CnU6 promoter could drive the transcribtion of shRNA, which could silence the target gene, and gRNA, which could guide Cas9 nuclease to cut the target site.

Conclusion: The CnU6 promoter from C. neoformans was successfully identified and cloned, which could drive the transcription of shRNA and gRNA efficiently.

Objective: Hydrocarbon microseepage is a natural phenomenon that hydrocarbon gases of subsurface petroleum accumulations migrate upward by reservoir pressure. The detection of the activity and distribution of these highly specialized populations can be used to forecast the existence of oil and gas deposits. However, the hydrocarbon-oxidizing bacterial population are usually not predominant in soil samples above the typical onshore oil and gas reservoirs. It is hard to assess the abundance of hydrocarbon-oxidizing bacteria.

Methods: In this study, changes of microbial abundance and functional genes were studied.

Results: Under gaseous hydrocarbon condition, changes of methane and butane oxidizing bacteria were different. Furthermore, changes of functional genes indicated that genome analysis was more proper for microbial anomalies detection.

Conclusion: The profiling data of this study provide a comprehensive insight into gene expression profiles and lay the foundation for optimizing the microbial prospecting technology.

Objective: To provide scientific data for studying the ecology of cyanophage, we studied the genetic diversity of psbA of cyanophage from paddy floodwater in northeast China and its phylogenetic positions.

Methods: Membrane separation and concentration of cyanophage, PCR-cloning-sequencing were applied to study the diversity of psbA of cyanophage from paddy floodwater in northeast China.

Results: In total 17 psbA sequences of cyanophage were obtained. Novel cyanophages were found by phylogenetic analysis. Compared to those of Japanese paddy floodwater, marine and lakes, psbA gene assemblage of paddy floodwater in northeast China was significantly different.

Conclusion: This is the first report to study genetic diversity of cyanophage from paddy floodwater in northeast China with a molecular marker of psbA by PCR-cloning-sequencing. The novel psbA assembly of cyanophage was found in paddy floodwater in northeast China.

Objective: The complete genome of the extreme environmental resistant bacterium Deiococcus radiodurans R1 was analyzed by sequence comparative method and putative ferritin-like protein DRA0258 was screened. Molecular techniques were applied to validate and analyze its function.

Methods: We applied sequence alignment to analyze amino acid sequence of the hypothetical protein DRA0258 and detected its iron binding activity after purification. We used triple-fraction-ligation method to construct dra0258 null mutant and detected its survival rate under H2O2 treatment, catalase activity and total antioxidant capacity, using QRT-PCR to examine the relative transcriptional level change of the antioxidant relative enzymes and iron transport relative proteins.

Resutls: We confirmed DRA0258 obtained a certain iron binding activity. The survival rate assay with H2O2 treatment suggested that deletion of dra0258 reduced the cellular antioxidant activity of D. radiodurans. The attenuation of catalase activity, total antioxidant capacity as well as the reduction of relative transcriptional levels of antioxidant related genes verified that both the oxidative stress response systems and the iron regulation network were damaged.

Conclusion: This study verified DRA0258 is an iron-binding protein. Deletion of this gene would affect cellular iron transport system and reduce cellular antioxidant capability.

Objective: To study the regulation of sporulation controlled by two-component system (TCS) YvcPQ.

Methods: β-galactosidase experiment was used to verify the regulation of YvcP on kapD expression; bacterial one-hybrid assay, EMSA and RT-qPCR were applied to study the regulation of AbrB on yvcPQ expression; markerless gene deletion coupled with spore count was used to reveal the influence of yvcPQ and kapD expressions on sporulation.

Results: transcriptional regulator AbrB up-regulated the expression of yvcPQ; YvcP promoted the expression of kapD to inhibit sporulation.

Conclusion: AbrB up-regulated the transcription of yvcPQ operon, then the increased YvcP strengthened the transcriptional acitivation of sporulation inhibitor gene kapD, and subsequently inhibited sporulation.

Bacteria get a survival advantages in the complex environment through the chemotaxis system. Chemotaxis plays an important role in colonization and pathogenicity of bacteria, the legume–rhizobia symbiosis, and plant-microbe interactions. Phosphatase CheZ is an indispensable regulatory protein in the process of chemotactic signal adaptation. This review focused on the structure, mechanism of action, functional regulation, protein targeting and the status of the evolutionary of CheZ. This work can benefit the study of other bacterial chemotaxis systems.