Abstract:Acetic acid bacteria (AAB) are obligately aerobic Gram-negative bacteria. Known for their ability to oxidize ethanol to acetic acid and robust tolerance to acetic acid, AAB have been widely used in industrial vinegar fermentation. Besides the incomplete oxidative ability, investigation of their resistance mechanisms to acetic acid is intriguing and crucial for high titer vinegar production. In this review, we evaluated a variety of resistant pathways involved in carbohydrate metabolism, protein metabolism, lipid metabolism, and stress response based on genomics and proteomics investigations in AAB. Specifically, the discovery in modules related to quorum sensing (QS) system in Komagataeibacter species and the emerging genome data of AAB opens a new window to screen acid resistance regulatory networks, which may promote industrial strain breeding and fermentation optimizing. We reviewed the latest research progress of quorum sensing in acetic acid bacteria based on the brief introduction of genomic and proteomic studies.

Abstract:Pneumolysin is a multifunctional virulence factor expressed by Streptococcus pneumoniae. Pneumolysin includes 4 domains and is a member of cholesterol-dependent cytolysins. Pneumolysin has extensive cytotoxicity to a range of host cells. Furthermore, pneumolysin can activate complement classical pathway, and induce macrophages and monocytes to produce proinflammatory cytokines, mediate host immune responses. Consequently, pneumolysin is a potential candidate target for research and development of vaccines and drugs. In this review, the latest research progresses on the structure and function of pneumolysin, and related vaccines are discussed.

Abstract:Quorum-sensing (QS) involved in the production of N-Acylhomoserine lactones (AHLs) is a universal way of communication of gram-negative bacteria. Complete AHL-QS system includes pairs of AHLs synthase belonging to LuxI family and cognate LuxR-family AHLs sensor-regulator. However, many gram-negative bacteria have evidenced the presence of AHL-QS related LuxR-type genes, which are unpaired to a cognate LuxI. These unpaired LuxRs have been called solos or orphans. LuxR solos are thought to be important for bacterial signal perception in eavesdropping, intra-species and inter-kingdom communication, which become research topic in the field of QS. Here, the finding, concept, protein structure, and main types of LuxR solos are illustrated. Furthermore, the function and important protein of LuxR solos associated with sensing AHLs or non-AHLs are reviewed. The prospect and significance of quorum sensing LuxR solos in bacteria are also discussed.

Abstract:[Objective] To reveal the morphological pattern of Isaria cicadae. [Methods] We observed 17 morphological characters and measured 75 strains of 15 populations in I. cicadae. Statistical analysis system (SAS) 8.1 was used to analyze the morphological data, the morphological pattern was analyzed in 15 populations of I. cicadae, using the descriptive statistical analysis, nested analysis and Q cluster analysis. [Results] Two types of asexual conidium (large and small conidium) were observed in I. cicadae. The gourd-shaped and bottle-shaped conidiogenous cells were observed in I. cicadae. Many chlamydospores of I. cicadae were easy to form in PDA medium. Many fusion hyphae were generated between hyphae, and some fusion hyphae between hypha and chlamydospore, the fusion hyphae between conidiogenous cells were also observed. The CV of 17 morphological characters was from 13.07 to 104.09% in I. cicadae, indicating an ample morphological diversity at the species level. The nested variation analysis of the 17 morphological characters indicated that about 11.29% of the variability was attributable to the differentiation among populations, the rest 15.27% of the variability was derived from individual strains, and the remaining 73.44% was resided in the observations in the same strain. [Conclusion] The phenotypic variation within strain was the main morphological variation of I. cicadae. The morphological characters had no significant relationship with geographical origin in I. cicadae.

Abstract:[Objective] The aim of this study was to characterize β-glucosidase from Citrobacter koser GXW-1 isolated from soil and to improve the enzyme by molecular modification. [Methods] A bacterial strain with β-glucosidase activity was screened from the soil around Wuming sugar mill in Guangxi by esculin-ferric ammonium citrate selecting plate. The 16S rDNA of the strain was obtained and analyzed. By searching GenBank database, the genes encoding β-glucosidase from the same genus Citrobacter were found. These sequences were aligned. Then, a gene encoding β-glucosidase was amplified by PCR. The recombinant plasmid pQE-cbgl was constructed. The recombinant protein was purified with Ni-NTA. The enzyme properties of the recombinant protein CBGL were studied in detail. At last, the wild enzyme CBGL was reformed by error-prone PCR and site-directed random mutagenesis. [Results] C. koser GXW-1 with β-glucosidase activity was isolated from the soil. A gene encoding β-glucosidase was cloned from the wild strain GXW-1. The properties of CBGL were identified. Its optimal pH and temperature were 6.0 and 45℃. Its K_m and V_max value were (11.280±1.073) mmol/L and (0.1704±0.0073) μmol/(mg·min), respectively. Its K_i values was (66.84±3.40) mmol/L. CBGL can hydrolyze α-pNPG, stevioside, daidzin and genistin. CBGL was modified by error-prone PCR and site directed random mutagenesis. A positive mutant W147F was obtained successfully. Its V_max was 2.54 times that of the wild enzyme CBGL. [Conclusion] CBGL not only can hydrolyze β-glycosidic bond, but also can hydrolyze the α-glycosidic bond in α-pNPG. Furthermore, CBGL can hydrolyze stevioside, daidzin and genistin. These characteristics indicate that the β-glucosidase CBGL has important applications in theoretical research and in industry.

Abstract:[Objective] Algal blooms occurred in some sections of Shennong bay after impounding of Three Gorges Reservoir. [Methods] Related environmental and hydrodynamic factors were monitored during the period of algal blooming season in 2014 (March 20, April 13, May 23) in Shennong bay, Three Gorges Reservoir. To study succession of planktonic algae, water stable coefficient, euphotic depth and mixed layer depth were used to analyze stratification and hydrodynamic characteristics. [Results] We identified 6 phyla, 38 species (genera) planktonic algae. The sensitive area of algal bloom was at SN05 (677.677×10⁵ cells/L) and SN06 (716.761×10⁵ cells/L), and the planktonic algae biomass during this period was significantly different (ANOVA, p<0.05). Moderate water temperature, adequate nutrients, weak stratification and poor mixing promoted the rapid growth and breakout of the diatom bloom with Cyclotella spp. as the dominant species in March. Further increase of water temperature, stronger stratification and decrease of dissolved silicate and mixing layer restricted the diatom growth. Chlorella spp. and Chlamydomonas spp. grew better in shallow mixed layer with rich nutrients and evident stratification. Then Chlorophycean bloom broke out with Chlorella spp. as the dominant species and Chlamydomonas spp. the next-dominant species. High biomass maintained in April. In May, algal bloom gradually vanished due to sharp fluctuation of water level and increase of velocity. Monitored maximum water velocity was 0.1141 m/s at 2 m depth, exceeded an optimal flow rate perfect for growth of planktonic algae. [Conclusion] Stratification and hydrodynamic characteristics had important effect on planktonic algae under the condition of adequate nutrients. Velocity became the main factor that inhibited the growth of algae in Shennong bay in pre-flood falling stage of the Three Gorges Reservoir.

Abstract:[Objective] We studied the biological and the epidemiological characteristics of the pathogen of hypertrophy sorosis scleroteniosis, which is a devastating fungal disease of mulberry. [Methods] We studied the asexual and sexual reproductive phase stages of C. shiraiana, including the infection ability of hyphal, dormancy of sclerotia, the structures, release, number and germination of ascospores from apothecia, as well as the phenology of sclerotial germination. [Results] In C. shiraiana, hyphae had no infection ability toward the female flowers of mulberry. Sclerotia of C. shiraiana must undergo cold treatment above 6 weeks, then the dormancy-breaking sclerotia could germinate to apothecia. One to fifteen apothecia were germinated from one sclerotium, and the number of ascospores in a 1.5 cm diameter apothecia could contain up to (5.6-6.3)×10⁷. Ascospore C. shiraiana had significantly higher germination rates in acid than in neutral and alkaline environments. From late January to middle April, sclerotia germinated to apothecia, and got the highest value in the middle of March. [Conclusion] C. shiraiana is a formidable pathogen to cause epidemic disease and damage in mulberry.

Abstract:[Objective] The present study aims to analyze the chemotaxis genes and proteins of several PAH-degrading Novosphingobium strains, and the chemotaxis of these strains toward aromatic compounds and intermediates. [Methods] Based on genome comparative analysis, we identified the chemotaxis genes organization and proteins distribution. We used drop and swarm plate assays to detect the chemotaxis of these strains toward aromatic compounds and intermediates of TCA cycle. [Results] We found that all these Novosphingobium strains showed chemotaxis, but the chemotatic ability varied. The completed genome sequenced strains N. pentaromativorans F2, N. pentaromativorans US6-1, N. pentaromativorans PP1Y, Novosphingobium sp. AP12, Novosphingobium sp. Rr 2-17, and Novosphingobium nitrogenifigens DSM 19370 contained MCP, CheW, CheA, CheB, CheR and CheY. Strain F2, US6-1 and PP1Y, shared a consistent order of chemotaxis genes in "che" cluster. The chemotatic system of these Novosphingobium strains belonged to the Fla chemotactic system. [Conclusion] These strains all contained a complete chmotaxis pathway. Their chemotactic ability toward aromatic compounds and intermediates varied, and the chemotaxis of US6-1 was obvious.

Abstract:[Objective] Endophytes are widespread in plants and build long-term mutually beneficial symbiotic relationship with the host. However, the mechanism of their interactions with the host needs further study. To explore the mechanism of endophytic bacterium ginkgo endophyte KM-1-2, we managed to forecast its secretory proteins based on its genome and explicit characteristics.[Methods] Signal peptide analysis software SignalP, transmembrane helical structure analysis software TMHMM and Phobius, cells position software PSORT, subcellar localization software TargetP and GPI anchor site analysis software big-PI Predictor were used to predict the scope of all secreted proteins, which were defined as secretome.[Results] Altogether 128 typical signal peptide secretory proteins were screened out of 5299 protein sequences in KM-1-2 genome, accounting for 2.4% of the whole genome. The shortest ORF encoding these proteins is 61 bp, the longest one is 2105 bp and the average is 373 bp. The length of the signal peptide guiding secretory protein was distributed between 15 to 37 aa, with the average length of 24 aa. Amino acid with the highest present frequency of signal peptide in proper order is alanine, leucine and valine. The type of signal peptide cleavage belongs to A-X-A which named SPI cleavage type. Among the total secretory proteins 66 pieces have functional description and 26 pieces were enzymes. These enzymes mainly include glycoside hydrolase, esterase transferase, REDOX enzyme and carbon oxygen lyase.[Conclusion] The predicted secretory proteins of Streptomyces lavendulae KM-1-2 were achieved through bioinformatics analysis. These secretory proteins involved some enzymes and other unknown functions. This result laid the foundation for further study between endophyte and host.

Abstract:[Objective] We studied the dynamic nuclear behavior of Hypomyces perniciosus on axenic culture and its disease progression after infection on different growth stages of Agaricus bisporus. [Methods] Infection process was initiated by inoculating different stages of A. bisporus fruit body, and different depths of compost and casing soil with H. perniciosus. Disease progression was studied by observing symptoms on the fruit body using light microscopy and scanning electron microscopy. The nuclear behavior of H. perniciosus was determined by observation using fluorescence light microscopy after binding of DNA specific fluorochrome dye (DAPI:4, 6-Diamidino-2-phenylindole dihydrochloride) to the nuclei. [Results] Inoculating H. perniciosus on different depths of compost and casing soil resulted in different disease rate as follows:on the surface of casing soil>in the center of casing soil>between the casing soil and the compost>in the center of compost. H. perniciosus can infect any stage of fruit body development, when young primordial (up to 3 mm) was infected, large, irregular and tumorous fungal masses were formed. H. perniciosus directly penetrated A. bisporus without the formation of appressorium-like structures. The germination of the conidia led to a necrotic brown lesion symptom on A. bisporus at the beginning stages of disease development. The mycelium of A. bisporus plasmolysed, hydropically degenerated, cytoplasmolysed, emptied of mycelium cytosol and eventual death as the disease advanced. H. perniciosus produced two types of conidia. Group I conidia had no septa, colorless and smooth containing one nucleus. Group II didymoconidium had septa, containing two nuclei, separated by septa. The first round of mitosis occurred in conidia with no nucleus in the germinal tube. Another kind of asexual spore for thicker cell wall wart convex chlamydospore, chlamydospore had two cells. The upper cell had two nuclear while the basal cell had one or two nuclear, when germinated, it produced one or two germinal tubes. The number of nuclear in the germinal tube was irregular, usually contained 0 to 2 nuclear. [Conclusion] H. perniciosus can infect any part of the A. bisporus fruit body and can cause tremendous cytology changed. If we perform single spore isolation to do genetic analysis, one must isolate conidia with no septa.

Abstract:[Objective] To study the effects of bacteria on the species and morphology of carbonate minerals. [Methods] We conducted a series of cultural experiments in the medium with initial Mg/Ca ratio of 2 but without carbonate ion using Curvibacter sp. strain HJ-1 for 50 days. During the cultivation, bacterial density, precipitate quantities, calcium and magnesium concentration were determined. The morphologies of precipitated carbonates were observed using scanning electron microscopy, and mineral species of carbonate were determined by X-ray diffraction. [Results] Strain HJ-1 could induce the precipitation of carbonate minerals, the quality of carbonate gradually increased with the incubation time. XRD patterns showed that the mineral precipitates consisted of high-Mg calcite and aragonite. The percentage of aragonite in the precipitates was up to 86%. The morphology of carbonate minerals was multiform, including rod-shaped, dumbbell-shaped, spherical, tabular, as well as irregular and flake. [Conclusions] The formation of aragonite under the condition of low Mg/Ca ratio has a close correlation with extracellular polysaccharide secreted by Curvibacter sp. strain HJ-1.

Abstract:[Objective] A flavonoid 3'-hydroxylase from tea plant was engineered to synthesize B-3',4'-dihydroxylated flavones such as eriodictyol, dihydroquercetin and quercetin. [Methods] Four articifical P450 constructs harboring both flavonoid 3'-hydroxylase gene from Camellia sinensis (CsF3'H) and P450 reductase gene from Arabidopsis thaliana (ATR1 or ATR2) were introduced into Escherichia coli strains TOP10, DH5α and BL21, resultantly engineering strains S1 to S12. The plasmid pYES-Dest52-CsF3'H harboring CsF3'H gene was introduced into yeast Saccharomyces cerevisiae WAT11 designated as strain S13. The plasmid pES-HIS-CsF3H::AtFLS 9 AA was constructed through fusing flavanone 3-hydroxylase gene from Camellia sinensis (CsF3H) and flavonol synthase gene from Arabidopsis thaliana (AtFLS). Plasmid pES-URA-CsF3'H and pES-HIS-CsF3H::AtFLS 9 AA were then co-introduced into yeast S. cerevisiae WAT11 designated as strain S14. [Results] Strain S6 generated highest bioconversion efficiency at 25℃ among all E. coli strains during 24 h fernentation. Supplemented with 1000 μmol/L naringenin, dihydrokaempferol and kaempferol, the maximum amounts of eriodictyol, dihydroquercetin and quercetin produced by strain S13 were 734.32 μmol/L, 446.07 μmol/L and 594.64 μmol/L respectively. Supplemented with 5 mmol/L naringenin, the maximum amounts of eriodictyol, kaempferol, quercetin, dihydroquercetin and dihydrokaempferol produced by strain S14 were 1412.16 μmol/L, 490.25 μmol/L, 445.75 μmol/L, 66.75 μmol/L and 73.50 μmol/L during 36-48 h fermentaion respectively. [Conclusion] CsF3'H was engineered for biosynthesis of B-3',4'-dihydroxylated flavone.