Abstract:[Background] Heterologous expression of many superoxide dismutases (SODs) from bacteria, yeast or plants have been reported to improve the salt tolerance of the host, but few from archaea. In particular, no SODs from archaea has been expressed in bacteria to improve their salt tolerance. [Objective] We are aiming to mine putative SODs encoding genes in archaea Haloferax sp. D1227, and to identify its biochemical function. It is also our intention to see possible enhancement of salt tolerance in para-nitrophenol utilizer Burkholderia sp. SJ98, by introducing the newly identified SOD. [Methods] The putative SOD encoding gene from strain D1227 was mined by bioinformatics analysis before its heterologous expression in E. coli BL21(DE3) and strain SJ98, which was further purified through AKTA purifier system. The specific activity of cell extracts and purified enzyme were measured by a spectrophotometry. Cells of strain SJ98 with the SOD gene and with vector only were cultured in M9 medium (with 0 and 500 mmol/L NaCl) with glucose or PNP as carbon source, respectively. Subsequently, the growth and degradation of these strains were detected by automatic growth curve analyzer and high-performance liquid chromatography. [Results] A putative SOD encoding gene from Haloferax sp. D1227 was found and designated sodA. SodAD1227 heterologous expressed in both E. coli BL21(DE3) and strain SJ98 exhibited SOD activity (with specific activities of 21.07±0.02 U/mg and 84.56±0.16 U/mg respectively in cell extracts). The specific activity of purified protein from E. coli was 179.46±3.43 U/mg. Strain SJ98[pBBR-sodA] grew well in M9 containing 500 mmol/L NaCl with glucose as carbon source, while strain SJ98[pBBR1MCS-2] almost lost its growth ability. When para-nitrophenot (PNP) was used as carbon source, strain SJ98[pBBR-sodA] still had a normal growth with a proper PNP degradation ability, while the growth and degradation ability of strain SJ98[pBBR1MCS-2] was almost lost. The structural analysis of SodAD1227 simulated by Phyre2 showed that SodAD1227 has the typical structural characteristics of the Fe/Mn-SOD family. [Conclusion] This study provides a potential feasibility for the use of archaeal SODs in transforming bacteria to adapt to the degradation of organic pollutants in high salinity environment.

Abstract:[Background] Genome editing method coupling the CRISPR (Clustered regularly interspaced short palindromic repeats) system with λ-Red recombination technology has become an important access to Escherichia coli genomic editing. Recently, there have been several genomic editing strategies in E. coli based on CRISPR system. However, these methods often require many processes, such as the elimination of single plasmid or multi-fragment assembly, and are still inefficient, tedious and time-consuming. [Objective] Establish a fast, continuous and efficient CRISPR genome editing method based on several different temperature-sensitive plasmids in E. coli, and improve the editing efficiency of CRISPR in E. coli. [Methods] The pTarget plasmid in traditional CRISPR method was modified to an RK2ts-type plasmid with better temperature sensitivity. Two plasmids with different resistance markers (pTW-A/S) were constructed and used alternately to eliminate false positives. And this realized the elimination of plasmids and next round of gene integration simultaneously. [Results] At the same temperature, RK2ts-type plasmid was eliminated more easily than pSC101ts-type plasmid, and could selectively eliminate pTW-A/S plasmid. Moreover, it eliminated pTW-A/S plasmid and transformed other plasmids and target fragments during next round of gene integration simultaneously. The efficiency of gene knock-out/integration reached 100%. Then, we used this method to construct BP03 by modifying gene BspanD and aspA on strain BP01 continuously, leading to increased production of β-alanine successfully. [Conclusion] A novel, efficient, convenient, continuous and flexible CRISPR/Cas9-mediated genome editing strategy has been established in E. coli. This coordinate use of multiple temperature-sensitive plasmids method solves the problem of incomplete elimination of pTarget plasmids in traditional CRISPR systems, and it also avoids the low connection efficiency in process of large plasmids construction. Therefore, it shortens the experimental hours and provides a powerful tool for the construction of metabolic engineering strains.

Abstract:[Background] Chiral phenylethl acetate is a crucial chiral flavor compound and possesses important applications in the fields of food and fine chemicals. The enzymatic synthesis of chiral phenylethl acetate is of great industrial application prospect. [Objective] To characterize a novel microbial esterase EstC11 for enantioselective resolution of (±)-1-phenylethl acetate. [Methods] A microbial esterase EstC11 identified from the deep sea of Western Pacific Ocean was cloned, expressed, and characterized. The optical purity of chiral product (R)-1-phenylethl acetate generated through enzymatic kinetic resolution was increased by optimizing the conditions of enzymatic reactions, such as pH, temperature and organic solvents. [Results] The optimum pH and temperature of EstC11 were 8.5 and 25 °C, respectively. Some metal ions and organic solvents could inhibit the hydrolytic activity of EstC11. Under the optimum reaction conditions (pH 9.0, 50 mmol/L Tris-HCl, 20 °C, 50 mmol/L substrate concentration), the optical purity of (R)-1-phenylethl acetate reached 98% with the yield of 39% after 3 h. [Conclusion] The optical purity and yield of (R)-1-phenylethl acetate were dramatically improved after the optimization of enzymatic reaction conditions. Thus, our study laid the foundation for the application of esterase EstC11 in industry.

Abstract:[Background] As a relatively new member of marine probiotics, Bdellovibro-and-like organisms (BALOs) have a wide application prospect. However, because of BALOs’ special reproductive mode and cycle, the pratical application of BALOs is greatly affected by their parasitic host characteristics and biological activities. Therefore, selecting appropriate parasitic hosts and maintaining the activity of BALOs preparation are very important for BALOs’ application. [Objective] to isolate a strain of BALOs which can use gram-positive probiotics Bacillus subtilis as host, and increase its Bdelloplast concentration in the culture for storage. [Methods] Bacillus subtilis was used as host bacterium to isolate BALOs from sea mud samples from Hainan by DNB (Dilute nutrient broth) double plate method. Transmission electron microscope and partial 16S rRNA gene sequence analysis were conducted to identify the Bdellovibrio strain, and its biological characteristics and lysis spectrums were studied. In addition, the effects of ampicillin, indole, Ca2+ and Mg2+ on the formation of bdelloplast were researched. [Results] A strain of BALOs (named BDE-1) which could prey on Bacillus subtilis was isolated and identified successfully. The optimal temperature, salinity and pH for BDE-1 were 25 °C, 2.0% and 7.0 respectively. Lysis experiments on 28 pathogenic or potential pathogenic strains showed that BDE-1 lysed 24 strains, corresponding to 85.7% of lysis rate. For the 13 strains of tested vibrios, its lytic rate was as high as 92.3%. While all four factors (indole, ampicillin, Ca2+ and Mg2+) promoted bdelloplast formation, the promotion effect of indole and Ca2+ was more significant. [Conclusion] This study not only provided a feasible solution for the optimal selection of parasitic hosts of Bdellovibro, but also provided a theoretical basis for the maintenance or enhancement of the application of Bdellovibro probiotic preparation.

Abstract:[Background] The problem of bacterial drug resistance is becoming more and more serious. The development of new antibiotics far lags behind the clinical needs. The discovery of microbial drug resources from special habitats is expected to solve this problem. [Objective] Biodiversity and bioactivity screening of actinomycetes from color desert in Dengpa, Tibet were performed, which would lay a good foundation for discovery of the resources of medicinal actinomycetes and new antibiotics. [Methods] The actinobacterial strains were cultured on 8 different media. Biodiversity of actinobacterial strains were analyzed according to 16S rRNA gene sequences. Detection of PKS-II (ketosynthase, KS), NRPS (adenylation A domain), AHBA and Halo genes was performed by PCR and agarose gel electrophoresis. Screening of antimicrobial activity was carried out by filter paper method. Antioxidant ability including total antioxidant activity, hydroxyl and oxygen free radical scavenging rate was tested by the chemical method. [Results] a total of 231 strains were isolated from four soil samples from color desert in Dengpa, Tibet. 16S rRNA gene analysis revealed these strains were affiliated with seven actinobacterial genera. Streptomyces was the dominant genus. Screening of biosynthetic gene clusters showed that 68 strains had at least one of four biosynthetic gene clusters and 6 strains had all the tested biosynthetic gene clusters. Furthermore, secondary metabolites produced by these strains showed antimicrobial activity against certain pathogens and antioxidant capacity towards hydroxyl and oxygen free radical scavenging. Therein, eight strains displayed broad-spectrum antimicrobial activity. Thirteen strains showed the positive total antioxidant activity. Ten strains had good hydroxyl free radical scavenging and three strains showed positive oxygen free radical scavenging. [Conclusion] These results exhibited that there are abundant actinomycetes medicinal resources in color desert in Dengpa, Tibet, which was promising for discovering new strains of actinomycetes and developing new antibiotics.

Abstract:[Background] Lhalu Wetland was the highest and largest urban natural wetland in China. Studying on microbial community structure in Lhalu Wetland could provide theoretical basis for exploitation and protection of biological resources in Qinghai-Tibet plateau, and lay foundation for the study of microbial diversity in wetland ecosystem. [Objective] investigate diversity of mycelial fungus and explore the main environmental factors which influenced the community structure of filamentous fungi in Lhalu Wetland. [Methods] Fungi were isolated from 11 water samples, and identification of fungi were completed using the sequence analysis of nrDNA ITS, combining with traditional classified method. SPSS and CANOCO were used to examine correlations between filamentous fungi diversity and environmental factors. [Results] Mycelial fungi isolated from Lhalu Wetland were belonging to 6 genera and 13 species. Mucor, Cladosporium, Galactomyces were the dominate genera in the wetland. M. hiemalis、M. racemosus and G. geotrichum were the dominate species. Statistical analysis indicated that TN has significantly negative correlation with the counts of filamentous fungi at the Lhalu Wetland (P<0.05), moreover, TN and TP had significant effect on distribution of the fungi, which positively correlated with M. racemosus but passively correlated with M. hiemalis. [Conclusion] Environmental variable has been one of the major factors that influencing wetland microorganism community structure, therefore, it was crucial concerning to study the relationship between environmental factors and diversity of microbe in Tibet.

Abstract:[Background] Urban river sediments are rich in microbial resources, and the surface sediment is a major site for nitrification. Microorganisms in surface sediment play an important role in nitrogen transformations of river ecosystems. [Objective] To compare and analyze the microbial communities of ammonia-oxidizing microorganism enrichments from sediments under different environmental conditions, sediment samples were collected at four sampling sites at the Shunao and Hengjiang Rivers, which are tributaries of the Wen-Rui Tang River located in Wenzhou. [Methods] Ammonia-oxidizing microorganism enrichments were obtained through field sampling and laboratory cultivation. The microbial composition, abundance and diversity of ammonia-oxidizing microorganism enrichments were studied by high-throughput sequencing. [Results] Proteobacteria and Bacteroidetes were the two dominant phyla in enrichments. The microbial communities in four samples contained three genera of ammonia-oxidizing bacteria (Nitrosomonas, Nitrosospira and Nitrosococcus) and one genus of ammonia-oxidizing archaea (Nitrososphaera), of which the genus Nitrosomonas was dominant in each sample. There were marked differences in the composition of ammonia-oxidizing microorganism OTUs (operational taxonomic units) among different enrichments. The number and relative abundance of ammonia-oxidizing bacteria OTUs were highest in the sample DA2 from the site that has many aquatic plants, while those of ammonia-oxidizing archaea OTUs were highest in the sample DA4 from the site where untreated municipal wastes are often dumped there. The number and relative abundance of ammonia-oxidizing microorganism OTUs were higher in the enrichment samples DA2 and DA4 from lotic-dominated sites compared to those from lentic-dominated sites. [Conclusion] The results revealed microbial diversity in enrichments from urban river sediments under four different environments, and described the dominant phyla in enrichments. Such data provides a reference for selecting the culture source of ammonia-oxidizing microorganism, and provides fundamental knowledge for isolation and further study of ammonia-oxidizing microorganism in urban river sediments.

Abstract:[Background] The wood lacquer is vulnerable to microbial corrosion in the environment of water preserved wood lacquer because of microorganisms. [Objective] In order to study the microbial community structure and microbial disease information of wood lacquer ware. [Methods] The community structure of bacteria in wood lacquer and water samples was analyzed by Illumina MiSeq high throughput sequencing technology. [Results] There were differences between wood samples and water samples in the microbial community structure on the gate level, wood samples of a total of 7 dominant bacteria (relative abundance, gate>1%) were Proteobacteria (64.00%), Acidobacteria (14.70%) and Actinobacteria (3.83%). There were 6 dominant phyla detected at the phylum level, Proteobacteria (61.26%), Acidobacteria (8.25%) and Planctomycetes (4.88%), respectively. A total of 8 dominant species (relative abundance>1%) were found in the woody samples, which were Phenylobacterium (16.24%), Acidobacteria-Gp6 (9.68%) and Rhodoplanes (6.45%), respectively. There were 10 dominant genera in water samples, which were Naxibacter (9.03%), Acidobacteria-Gp6 (3.84%) and Nevskia (3.27%), respectively. The unclassified bacteria in wood samples accounted for 8.70% and 50.68%, respectively. The unclassified bacteria in water samples accounted for 12.83% and 59.35% respectively in the door and genus. [Conclusion] The microbial community structure of wood samples and water samples is abundant in the water preservation environment of wood lacquerware, but the microbial diversity is more complex than that of wood samples in the water samples of the door and genus level. In addition, a large number of potential new bacteria were detected in the tested samples.

Abstract:[Background] The leakage expression of promoters is a major concern in metabolic engineering and synthetic biology. To explore promoters with tight regulation like a switch would resolve this problem. [Objective] In order to avoid the shortcomings of using plasmid as research auxiliary, the aim of this study is to construct and assess promoter with tight regulation on chromosome. [Methods] Four elements including tetO, lacO, araC and rhaR regulated by TetR, LacI, AraC and RhaR respectively, were selected for designing six promoters including PtetO2, PtetO3, PlacO2, PlacO3, PlacO+ara and PlacO+rha. CRISPR/Cas9 system was applied to integrate these promoters into chromosome of Escherichia coli ATCC 8739. Promoter strength and relative expression with or without inducing agents were determined by the fluorescent quantities of GFP protein. [Results] Hybrid promoter PlacO+rha was the best one with tight regulation. The expression strength of PlacO+rha was 0.02 in absence of the inducer, and the maximum expression intensity was 12-fold higher than that of the induced lacZ promoter, of which the range of expression level was 600-fold. [Conclusion] This study would provide a good foundation for precisely regulating gene expression in metabolic engineering and synthetic biology.

Abstract:[Background] Dictyostelids cellular slime molds are model organisms for biology, cytology and developmental biology. Although more than one hundred species of dictyostelids have been reported up to now, the developmental process of each species is still unclear. [Objective] To understand the ontogenetic characteristics of this group. [Methods] Dictyostelids were isolated and identified from soil samples collected from Cang Mountain in Yunnan Province. The ontogeny of isolated dictyostelid species were investigated. The 18S sequences of this species were sequenced. The morphological features and 18S sequence were used to identify the taxonomic status. Furthermore, the development from spore to spore including characteristics of spore, myxamoeba, aggregation, pseudoplasmodia, ascent, and sorocarp were microbiologically observed on bi-concavity slide and water agar in this paper. [Results] Dictyostelium macrocephalum was obtained and identified. It adapt a wide range of temperatures, which suggested that this dictyostelid species is common species distributing in subtropical and tropical area. The whole life cycle of D. macrocephalum extends of a period of less than 3 d (71 h). The spores germinated and released myxomoebae 20 h after inoculation. The aggregations formed 50 h after the inoculation, whereas the pseudoplasmodia formation needed 56 h. After that, the sorogen formed needed 56 h, and finally form fruiting sorocarps after 71 h. [Conclusion] This new record of Yunnan Province, D. macrocephalum, extented our understanding on the distribution of this species in subtropical areas of China. The developmental process of D. macrocephalum was clarified.

Abstract:[Background] The long-term excessive use of chemical nitrogen fertilizers in agriculture has already caused ecological destruction of farmland. The direct consequences are soil compaction, secondary salinization and heavy metal pollution. Moreover, a large amount of nitrogen leaching loss of field could lead to eutrophication and groundwater pollution. The nitrate content of agricultural products may exceed standard, and eventually human health would be threatened due to the food chain. Thus, developing the biological nitrogen-fixing (NF) bacterial agents to reduce the use of chemical nitrogen fertilizer is of great significance for environmental protection and sustainable agriculture. [Objective] This study aims at creating abundant mutants of an associative NF bacterial strain, Kosakonia radicincitans GXGL-4A, isolated from maize root by Tn5 insertional mutagenesis and the results will benefit for further exploration of nitrogen fixation mechanism, nitrogen metabolism and related regulatory network, and lay a solid foundation for the application of GXGL-4A in agricultural production. [Methods] The nitrite reductase gene nirBD was cloned from K. radicincitans GXGL-4A by PCR. Meanwhile, a Tn5 transposon plasmid pMOD-egfp-tet was constructed and then transferred into the wild type GXGL-4A bacterial cells by electroporation to generate Tn5 insertional mutants. [Results] Large quantities of mutants were obtained, and four mutants with significant decrease in nitrite reductase activities comparing with the wild type strain GXGL-4A were identified. Subsequently, DNA sequence flanking integration site in the genome of mutant M36 were also isolated. [Conclusion] It is feasible and applicable for Tn5 transposon mutagenesis in the associative NF K. radicincitans strain GXGL-4A, and an effective and stable insertional mutagenesis system of this microorganism has been successfully established in this study.

Abstract:[Background] Plant rhizosphere growth-promoting bacteria are one of the hotspots in the current research of agricultural microorganisms because they promote plant growth or have antagonistic effects on pathogenic bacteria. Among them, Pseudomonas chlororaphis HT66 is a non-pathogenic biocontrol strain that can efficiently synthesize phenazine-1-carboxamide (PCN) — a promising and environmental friendly agricultural antibiotic. [Objective] To study the physiological function of ompR gene in P. chlororaphis HT66 and its biocontrol efficiency. [Methods] The ompR gene deletion mutant of HT66 strain was constructed by scar-less gene knockout method. We studied the effects of the mutation on the growth rate, biofilm synthesis, tolerance to pH and osmotic pressure, swarming mobility and PCN production. [Results] Compared with the wild-type strain, the ompR gene deletion inactivated mutant showed a slight reduction in cell biomass. The biofilm decreased by 31.5%, whereas the swarming mobility of the strain and the tolerance to osmotic pressure and pH decreased. However, the yield of PCN increased by 57.8% in the mutant strain. In HT66 strain, the ompR gene has a certain degree of regulation of its motility, environmental tolerance and physiological and biological functions. [Conclusion] Our findings enriched the metabolic pathway of P. chlororaphis, and will provide a basis for the research and application of the subsequent PCN synthesis mechanism.

Abstract:[Background] Potato dry rot is a fungal disease, caused by Fusarium spp., which always happens in both field and storage period. It mainly leads to quality deterioration and lower economic value. [Objective] To isolate and identify effective biocontrol strains to control potato dry rot, and investigate the inhibitory effect on plant pathogens. [Methods] We used 109 strains as tested strains, which were isolated from the rhizosphere soil of potato from Dingxi region in Gansu province. Antagonistic bacteria were screened by plate confrontation method with Fusarium sulphureum as the target strain, strain identification was based on morphology, physiological and biochemical characteristics and 16S rRNA gene sequence analysis. The effects of cell-free fermentation filtrate on the mycelial growth, spore germination of F. sulphureum, the lesion diameter of wounded inoculated potato tubers, the incidence of dry rot and germination of mung bean seeds were evaluated. [Results] Strain YL11 was isolated from the rhizosphere soil of potato and exhibited an excellent antagonistic activity against F. sulphureum, which was identified as Pseudomonas sp., cell-free fermentation filtrate of YL11 strain significantly inhibited the mycelial growth, spore germination of pathogenic fungi, the lesion diameter of wounded inoculated potato tubers and the incidence of dry rot. Moreover, the inhibition rate of 20% cell-free filtrate to F. sulphureum was 87.3%; 75% cell-free filtrate completely inhibited spore germination; the cell-free fermentation dip significantly reduced the lesion diameter of inoculated potato tuber, the inhibition rate was 67.1% after 14 days inoculation; the incidence of dry rot reduced by 68.4% after 90 days inoculation. Meanwhile, it also decreased the mycotoxin activity of F. sulphureum. [Conclusion] Strain YL11 significantly inhibited the growth of the F. sulphureum and had a notably biological control effect on potato dry rot, and it would be a potential biological control agent.

Abstract:[Background] With the increase of living standards and health awareness, quality and safety of milk are acquiring great concern. [Objective] To investigate aerobic plate count (APC) and total number of Coliform Group (TCG) in the milk of lactating Chinese Holstein cows from an intensive farm located in the lower reaches of the Yangtze River, China, by using different bacteriological methods. The causative factors of APC and TCG were also explored. [Methods] Taking the milk of Chinese Holstein cows as the research object, mastitis detection using mastitis rapid diagnostic agents, APC, TCG and the number of bacteria in the environment detection method with reference to national standards. The collected raw milk samples were subjected to routine isolation and identification of the bacteria, and the milk composition change data was collected. [Results] The results of multivariate analysis of variance showed that different lactation stages and different milk yields had significant (P<0.05) and significant (P<0.01) effects on the total number of colonies in Holstein cows? raw milk. Body condition score, milk yield, stage of lactation, parity and udder quarters had no significant effect (P>0.05) on TCG; APC and TCG measured in summer were significantly (P<0.01) higher than other seasons. Effect of the number of bacteria in different environment on APC and TCG were significant (P<0.05) above. With the increase of mastitis degree, APC and TCG in milk increased, with a significant difference in aerobic plate count for different degree of mastitis (P<0.01). Bacterial identification results in the majority of Staphylococcus and Bacillus infections and almost no Streptococcus infection. The change of the total number of colonies had no significant effect on the rate of milk protein (P>0.05), but had a significant effect on milk fat percentage (P<0.01). [Conclusion] Different physiological and pathological conditions of the randomly sampled lactating cows and the external feeding conditions and hygiene environment had different effects on APC and TCG in the raw milk. The milk fat percentage of raw milk was negatively correlated with the change in total number of bacteria.

Abstract:[Background] Vps74/GOLPH3 is a key protein involved in the glycosylation of the Golgi protein, and an important phosphatidylinositol-4-kinase effector and phosphatidylinositol-4- phosphate sensor participating in a variety of intracellular signaling pathways. [Objective] To characterize the Vps74/GOLPH3 homologue in Candida albicans and investigate the role of Cavps74 in stress response, secretion, morphogenesis and pathogenicity. [Methods] Sequence alignment and analysis were performed by the relevant biosoftware. The gene deletion strain vps74?/? and reconstructed strain VPS74c were constructed by PCR-mediated gene disruption. With reverse genetic analysis, we further investigated the function of Vps74 in protein glycosylation, stress response, secretion, morphogenesis and pathogenicity. [Results] There is a typical homologue of Vps74/GOLPH3 in C. albicans which also plays important role in glycosylation process. Deletion of vps74 resulted in various effects, including decreased secretion ability, morphogenesis, adhesion and loss of pathogenicity. [Conclusion] Vps74 plays a key role in protein secretion, morphogenesis, adhesion and pathogenicity in C. albicans.

Abstract:[Background] Hyperuricemia is well-known as a kind of metabolic disease with abnormal higher serum uric acid level. It is becoming a new method for treating hyperuricemia by degrading exogenous purine components ingested from food through probiotics. [Objective] Thirty probiotic strains of thirteen species were investigated for their abilities of lowering serum uric acid, we preliminarily propose the mechanism of its effects on hyperuricemic rats. [Methods] The ability of degrading nucleotides (adenosine monophosphate, guanosine monophosphate) , nucleosides (adenosine, guanosine), purines (xanthine, hypoxanthine and guanine) and uric acid by different probiotics were determined with HPLC, the probiotic strain with the fastest degrading ability was selected as the candidate strain for further test on hyperuricemic rats. Metabolites in the process of degrading nucleosides and nucleotides by probiotics were detected by mass, the mechanism of probiotics lowering the blood uric acid was preliminarily explored. [Results] Lactobacillus casei ZM15 was selected with the highest degradation rates of nucleosides and nucleotides. The animal experimental results showed that ZM15 had effects on hyperuricemia. Mass spectrometry results showed that after degradation of nucleotides and nucleotides in cell, guanine, xanthine and hypoxanthine appeared both inside and outside ZM15 cells, three kinds of purine content were markedly higher than those in normal cells, uric acid and allantoin were not found both inside and outside. [Conclusion] Lactobacillus casei ZM15 has the fasted degradation ability of nucleotide and nucleotide, and it has obvious effects of lowering uric acid on hyperuricemic model rats by mechanism of competition.

Abstract:The infection of Helicobacter pylori is increasingly drawing the world’s attention, and the radical treatment is the main measure taken by most countries aiming to reduce the rate of infection and subsequent health problems such as peptic ulcer, gastritis and gastric cancer. However, with the increase of drug resistance, especially the increase of macrolide drug resistance, the standard triple therapy is losing its power in controlling the bacteria and new therapies are in urgent need. But most of the new therapies use a longer course, a larger dose and a wider range of antimicrobial agents, and thus can bring severe influences on the structure and quantity of the gut microflora, which may result in severe side effects. Moreover, the deteriorating situation of the antimicrobial resistance will be exacerbated. This review summarized researches in the recent 20 years focusing on the effects that the eradication therapy may bring to the gut microflora. We also summarized some new experimental therapies, thus try to draw a clear profile of our current situation and give some clues for the treatment of Helicobacter pylori.

Abstract:Microbial communities play central roles in global climate regulation, human health and industrial biotechnology. The quantification of microbial diversity is important for the understanding of ecological characteristics of communities, their dynamics and functions. Herein, we introduced the commonly used alpha-diversity indices, including richness, Shannon-Weaver index, Simpson diversity indices and Hill’s diversity number. Also, we summarized diversity evaluation ways which are used in estimating the coverage of molecular methods (e.g., rarefaction curve and good’s coverage) and community richness (e.g., Chao1 and ACE indices and taxa abundance distribution curve). Then we showed the application of mathematic methods in the research on microbial diversity by taking wastewater treatment plant (WWTP) as an example which is the largest application of bioprocess engineering. Current investigations showed that taxa richness and Shannon diversity of activated sludge microbial communities in full-scale WWTPs increased with the increase in sampling sizes of different methods. However, the disparity between sample size and community size is a common problem in microbial investigations. By reconstructing microbial communities using DNA sampling data based on certain taxa abundance distribution curves, diversity of the communities was evaluated. It was shown that microbial richness was characterized with large uncertainty. Shannon diversity, and especially Simpson diversity indices which are weakly dependent on low-abundance taxa could be estimated accurately. They are good tools to evaluate and compare microbial taxonomic diversity. Developing novel modeling approaches and advances in sequencing technology can improve the accuracy of microbial richness evaluation. In addition, clarifying phylogenetic and functional diversity of microbial communities are also of substantial importance for the understanding of microbial ecology.

Abstract:The largest Zika virus (ZIKV) outbreak in South America in 2015 raised considerable worldwide health concern, owning to its unexpected link to microcephaly in the newborns and Guillain-Barré syndrome in adults. The WHO has declared the outbreak a global public health emergency. Currently no vaccines and therapeutics are available. The analysis of the crystal structure of ZIKV and viral proteins will deepen the understanding of viral replication and pathogenesis, which provides important targets for the development of vaccines and drugs. Here, we review the latest advances in research on the structural biology of ZIKV and viral proteins.

Abstract:Microbial metal responsive proteins are a class of DNA transcriptional regulators with metal sensing effect. There are seven families of regulators now been characterized (Ars R-Smt B etc.). Different representatives of each sensor families can regulate gene expression in response to different metals, they not only modulate the expression of genes directly associated with metal homeostasis of microbial cells, but can also alter metabolism to reduce the cellular demand for metals in short supply. At present, the metal response protein research has been some success, and the residues that form the sensory metal-binding sites have been defined in a number of these proteins. This review summarized the different families of bacterial metal-sensing transcriptional regulators and discussed current knowledge regarding the mechanisms of metal-regulated gene expression. Details of the structural features of sensory metal-binding sites focusing on the Ars R-Smt B family and the mechanisms of Fur gene expression. In addition, recent progress in understanding the coordination of the different sensors to control microbial cellular metal levels was discussed, as well as in the application of bio-metallurgy and environmental management prospects.

Abstract:Filamentous fungi not only are pathogenic but also play a significant role in heterologous expressing enzymes, chemicals, and pharmaceuticals. Along with the implementation and promotion of Human Genome Project, life science has entered functional genomics era. Proteomics study is a type of functional genomics research in order to understand cellular process mechanism, protein functions and their interactions, which provides a vast potential for future production of industrial enzymes and recombinant pharmaceutical proteins by filamentous fungi. This review summarized the research contents and methods of proteomics study, and its applications in filamentous fungi including pathogenic, antibiotic producing and cellulase producing strains. Furthermore, future studies in transcriptomics, proteomics and metabolomics could offer a full scale of understanding the metabolic network and regulations in filamentous fungi.

Abstract:English taught courses for undergraduate students have achieved rapid development in domestic universities. Microbiology is a professional compulsory course with international universality, and more and more universities implement all-English to deliver the course currently. However, there still exist some problems for English taught courses such as, language barriers to free communication and total understanding, imperfect evaluation system and so on. In order to address these problems and facilitate the sound development of English taught course for Microbiology, we proposed to exploit a blended “bilingual teaching+ Massive open online courses (MOOC) sharing course” teaching mode, which is in line with the characteristics of Chinese students, and can be used to overcome the difficulties faced by teachers and students in the teaching process of microbiology.

Abstract:Research and application of microbiology are of increasing importance in the development of human society. Microorganisms are closely related with the manufacture and living of mankind. Therefore, quality education in related field at university level, helping students learn the importance of microbiology, is of great significance. To this end, we set up the course Walking into the Microbial World, as a quality education course for students of non-life-science major. The course pays attention to the systematism and relevance of knowledge. Contents of basic theory, production and application are introduced in a systematic way. The course aims to enrich the knowledge of students and encourage their innovation. Gradational and functional teaching are built with the use of multimedia, internet and interactive teaching of cloud platform to improve students’ study interest, feedback learning and teacher-student interaction. Over the past three years, this course has become a high-quality course with our outstanding teaching method.

Abstract:[Background] As a selective marker, carboxin has been used in plant and fungi transformation. [Objective] To construct the genetic transformation system of Lentinula edodes by using carboxin resistant gene as a selective marker. [Methods] The mycelia of L. edodes strain 411-4 were collected at the 4th day after cultivation and then digested by lywallzyme to obtain the protoplast. Sufficient protoplast was mixed with the plasmid pL-cbx DNA and the polyethylene glycol (PEG) solution for transformation. The mixture finally was spread on the regeneration selecting plate, and the generated colonies were picked up for further testing. [Results] When more than 108 protoplasts and 4 μg plasmid DNA were used, 40 transformants were obtained. After being subcultured on media under carboxin selective stress, and the PCR identification, 38 transformants were positive and stable, indicating that carboxin gene was integrated into the genome of L. edodes 411-4. [Conclusion] The PEG mediated transformation method was constructed by using L. edodes strain 411-4 and carboxin selective marker.