Abstract:DegP, also known as HtrA, is a protein widely distributed in eukaryotic and prokaryotic cells. It has both protease activity and chaperone activity. On one hand, DegP forms a capsule-like structure to perform its chaperone function, called "Cage assembly". On the other hand, the protease activity of DegP depends on protease active site and PDZ1 domain, called "molecular ruler". In Gram-negative bacteria periplasm, DegP protects misfolded proteins or degrades denatured proteins. For example, DegP takes part in the transport of outer membrane proteins has been well-studied to prove its functions. Additionally, DegP can be secreted to extracellular and participates in the regulation of biofilm formation. In this way, DegP endows the ability to survive from the harsh environments on bacteria. In this review we summarized the structures and functions of DegP. It provides new insights into vitro activities of DegP and how DegP works as the final step in protein quality control system.

Abstract:Riemerella anatipestifer is a Gram-negative bacterium causing serositis in ducks and belongs to Flavobacteriaceae. Since its discovery, its outbreak and epidemic happen from time to time in various duck industries of China. Furthermore, it has a variety of serotypes and resistance to multiple antibiotics, leading to a serious threat to the duck industries. In recent years, with the help of molecular biology and related new technologies, more and more researchers have studied the virulence, antibiotics resistance and antibiotics resistance mechanism. Our review summarizes the above research progress, provide enlightenment and reference for further understanding this bacterium to prevent this disease.

Abstract:Saccharomyces cerevisiae is a commonly used aging model. Aging in yeast is assayed primarily by measuring replicative lifespan (RLS) or chronological lifespan (CLS). Several longevity factors have been identified through RLS and CLS studies, including Sirtuins (Sir2 protein family members). Sirtuins are well-known aging factors and a class of NAD dependent protein deacetylases that are evolutionarily conserved from prokaryotes to higher eukaryotes. Sirtuins exhibit multiple intracellular functions, including stress response, gene transcription, cellular metabolism, and longevity. The Sir2 protein is the founding sirtuin family member that functions in transcriptional silencing of heterochromatin domains and as a pro longevity factor for replicative lifespan. Deleting SIR2 shortens replicative lifespan, while increased gene dosage causes extension. Furthermore, Sir2 homologs in higher eukaryotes have been implicated in mediating age related diseases as well. In this review, we summarize the role of Sir2 and its yeast homologs, Hst1 through Hst4, in replicative aging and chronological aging.

Abstract:Nowadays, studies are increasingly emerging on the extremophiles inhabiting the extreme environments on earth. Covering 3/4 of the Earth's surface area, the ocean nurtures a variety of extremophiles found in extreme marine environments such as hydrothermal vents, hot springs, saline lakes and deep sea. These extremophiles can adapt to extreme high temperature, extreme low temperature, high pressure, high salinity, high radioactivity and extreme acidity and alkalinity. Their special biodiversity, together with their genetic background and unique metabolic pathways have evoked vast interests of scientist and industries. Research on extremophiles from variety of submarine geological landforms will be helpful to gain the knowledge on the origin of life, biodiversity, discovery and utilization of new natural resources, and geobiology. On the other hand, application orientated studies on panel of enzymes and variety of other unique active substances with extremophiles, have already been attempted with their potency well revealed. This review covers the latest progress on the studies covering the diversity of extremophiles, the development and utilization of new resources, research methods and techniques, respectively.

Abstract:[Objective] To assess the anti-stress characteristics and to optimize the conditions affecting its cadmium ions adsorption efficiency of a cadmium-tolerant Bacillus methylotrophicus strain NTGB29. [Methods] The anti-stress characteristics were studied by plate colony-counting method. In addition, we used single factor experiment and response surface methodology to optimize the fermentation conditions of cadmium ions adsorption of the strain. Moreover, the available cadmium ions contents were used to verify the cadmium ions adsorption ability of the strain in the cadmium contaminated soils. [Results] The maximum tolerance of NTGB29 to NaCl, pH and concentration of Cd²⁺ was 10%, 11.0 and 50 mg/L, respectively. The optimum cadmium adsorption conditions were as follows:initial pH 6.4, culture temperature 37℃, NaCl concentration 4.2%, liquid volume 50 mL in the 250 mL flask, and culture time 24 h. Under the conditions, the cadmium ions adsorption percentage reached to 79.70%. In addition, the available cadmium ions of the cadmium contaminated soils was decreased significantly by adding strain NTGB29, and the adsorption percentage was 29.65%. [Conclusion] Strain NTGB29 has good environmental resistance and Cd²⁺ adsorption capacity, and it could be a kind of valuable strain source to develop the heavy metal contaminated soil conditioners and microbial functional agents.

Abstract:[Objective] To establish a duplex PCR assay for detection of the strains XSBZ03 and XSBZ14. [Methods] Strain-specific primers were designed according to the target sequence of XSBZ03 and XSBZ14. Subsequently, duplex PCR assay based on the primers was successfully established. [Results] The duplex PCR assay could detect accurately strains XSBZ03 and XSBZ14. The detection limit for XSBZ03 and XSBZ14 genomic DNA concentration was 1.7 pg/μL and 2.0 pg/μL. In artificial seawater samples, the detection limit for XSBZ03 and XSBZ14 strain concentration was 6×10³ CFU/mL and 8×10³ CFU/mL, respectively. [Conclusion] This assay provides a reliable method for the rapid diagnosis of Porites andrewsi White Syndrome (PAWS) and the Specific Pathogen Free (SPF) coral transplantation caused by these two pathogens.

Abstract:[Objective] Heat shock response is an important cellular defense strategy to environmental stresses. Identification of the function of heat shock protein in the process of baculovirus infection and revealing the molecular mechanism of its role will provide a theoretical basis for understanding the molecular basis of host-virus interaction. [Methods] Bmhsc70-4 gene was cloned by molecular cloning, and multiple sequence alignment was performed with BioEdit and GeneDoc. The Bmhsc70-4 gene was overexpressed by eukaryotic expression and knocked out by CRISPR/Cas9 gene editing system. The expression of the corresponding genes was detected by quantitative real-time PCR. The effect of Bmhsc70-4 gene on apoptosis was determined by detecting the activity of Caspase-9 and Caspase-3/7. The co-localization of BmHSC70-4 and BmIAP was verified by immunofluorescence, and their interaction was further verified by immunoprecipitation. [Results] The open reading frame of Bmhsc70-4 gene was 1950 bp, encoding 649 amino acids and it was highly conserved among insects. Bmhsc70-4 gene was up-regulated during BmNPV infection. Overexpression of Bmhsc70-4 gene could promote the proliferation of BmNPV, and knockout of Bmhsc70-4 gene inhibited the proliferation of BmNPV, indicating that the expression of Bmhsc70-4 gene was conducive to the proliferation of BmNPV. We confirmed that Bmhsc70-4 gene can inhibit the apoptosis of silkworm cells. Immunofluorescence assay showed that BmHSC70-4 and BmIAP were co-localized in the cytoplasm and immunocoprecipitation showed that they could interact with each other. Bmhsc70-4 gene could promote the expression of Bmiap gene during BmNPV infection. [Conclusion] Bmhsc70-4 gene had the function of inhibiting apoptosis of silkworm cells, and it could interact with BmIAP and promote BmNPV replication and proliferation during the process of BmNPV infecting silkworm cells.

Abstract:[Objective] To study the effect of amino acid mutation(s) in the G-H loop of VP3 of foot-and-mouth disease virus (FMDV) serotype O on its biological characteristics. [Methods] Two site-directed mutants, rHN^V3174Y and rHN^{D3173N+V3174E+N3179C}, were generated by using reverse genetic technique platform of FMDV. The phenotypic properties of the resulting viruses were characterized by plaque forming assays, one-step growth curves, determination of TCID₅₀ and LD₅₀, indirect immunofluorescence assays and laser confocal microscopies. [Results] There were no significant differences in the infectivity, plaque phenotype, and replication kinetics between rHN, rHN^V3174Y and rHN^{D3173N+V3174E+N3179C} for BHK-21 cells. Compared to the backbone virus rHN, however, the pathogenicity of rHN^V3174Y and rHN^{D3173N+V3174E+N3179C} was obvious decreased in suckling mice, and these two rescued viruses gained the ability to infect CHO-K1 cells via caveola-mediated endocytosis. [Conclusion] Individual amino acid mutations at position 3174 in VP3 have an influence on the virulence and endocytic pathway of FMDV serotype O in the infection of host cells, which may help us understand the potentially important functional roles of the G-H loop of VP3 in the 3D spatial conformation of FMDV virion.

Abstract:[Objective] To obtain the dissimilatory iron reducing microorganisms (DIRM) from deep sea hydrothermal fields, analyze their mineralization ability and mineralization products, to further understand their role in iron biogeochemical cycle. [Methods] We enriched and cultivated DIRM from polymetallic sulfides of South Mid-Atlantic Ridge hydrothermal fields with FeOOH as an electron acceptor, and acetic acid etc. as electron donor under the constant 60℃ temperature anaerobic condition. The morphology observation and elemental composition analysis on mineralized products were carried out by scanning electron microscope, transmission electron microscope, selected area electron diffraction and Energy Dispersive Spectrometer. [Results] We obtained a total of 139 iron reducing microbial cultures from 2 polymetallic sulfides. All of them could transform FeOOH (Fe³⁺ 90 mmol/L) into mineralized iron products with obvious crystal structure, mainly in cubic shape with side length ranged from 5.0 nm to 20.0 nm. According to Energy Dispersive Spectrometer analysis, the elements of all mineral crystals were iron and oxygen, presumably a mixed mineral composed crystal of siderite and magnetite. The time of formation of mineral crystals varies from 3 to 54 d, and most cultures can form crystals within 11 to 20 d. Microbial diversity indicated that the dominant microorganisms in the culture were mainly Firmicutes and Euryarchaeota, including Carboxydocella and Desulfotomaculum, a new species (16S rRNA Homology 89%-91%) and Geoglobus. [Conclusion] At 60℃, bacteria and archaea in hydrothermal fields could transform ferric iron to mixed iron oxides mineral with the simple organic compounds as electron donor. These results supported the potential of microorganisms to participate in the biogeochemical cycle and mineralization. However, it requires extensive research work to verify their roles in situ.

Abstract:[Objective] We examined the physiological characteristics of invasive cyanobacterium Cylindrospermopsis raciborskii helix isolated from Litopenaeus vannamei culture ponds. [Methods] The algal strain was isolated and purified from shrimp ponds in Chenghai, Shantou, and identified by morphology and 16S rRNA sequence analysis. Then, with the general culture media of CT and BG11, we optimized the culture conditions. Finally, we analyzed the effect of heavy metals Cu²⁺ (0-0.8 mg/L), Cd²⁺ (0-4 mg/L) and Pb²⁺ (0-80 mg/L) on the growth of the isolated strain. [Results] The identity of 16S rRNA sequence of the isolated strain was higher than 98% with that of typical strains of Cylindrospermopsis raciborskii from GenBank. Under laboratory culture conditions, the optimal culture medium for the growth of the isolated strain was the modified BG11 (N 62 mg/L, N:P=9:1), the exponential biomass and growth rate were up to (0.632±0.170)×10⁷/L, (0.063±0.001)/d, respectively. The isolated strain tolerated heavy metals Cu²⁺, Cd²⁺ and Pb²⁺, and the tolerance concentrations was Pb²⁺ 1-40 mg/L, Cd²⁺ 0-0.5 mg/L, Cu²⁺ 0-0.2 mg/L. Furthermore, Cu²⁺ and Cd²⁺ inhibited the growth of the isolated strain, and the inhibition increased with enhancing of heavy metal concentration and contact time, the 96 h EC₅₀ of Cu²⁺ and Cd²⁺ were 0.125 and 0.551 mg/L for the strain, respectively, but Pb²⁺(0-80 mg/L) had a dual effect on its growth, which promoted growth at low concentration (≤ 40 mg/L) and inhibited growth at high concentration (≥ 80 mg/L). [Conclusion] A strain of Cylindrospermopsis raciborskii isolated from L. vannamei culture ponds has the potential of adsorbing heavy metal ions in shrimp ponds.

Abstract:[Objective] To clone and recombinant express the gene Heparinase I from Bacteroides thetaiotaomicron, and then characterize the recombinant SUMO-Bt-HepI and Bt-HepI. [Methods] Codon optimization was done on the gene sequence of B. thetaiotaomicron heparinase I. The target gene was obtained by PCR amplification, inserted into the expression vectors pET-28a and pE-SUMO, and then transformed into E. coli Rosetta (DE3) to obtain recombinant products Bt-HepI and SUMO-Bt-HepI. Heparin sodium was used as substrate to study the enzymatic properties of the recombinant proteins. [Results] SDS-PAGE analysis showed that the molecular weights of Bt-HepI and SUMO-Bt-HepI were about 42.5 kDa and 55 kDa, respectively. Compared with Bt-HepI, the specific enzyme activity of Heparinase I increased by 48.9% after fusion SUMO-Tag. The enzymological properties showed that the optimum pH and temperature of Bt-HepI and SUMO-Bt-HepI were pH 9 and 45℃, and both recombinant enzymes were stable at pH 5-9, while the acid resistance of SUMO-Bt-HepI was obviously higher than Bt-HepI when the pH value was lower than 5. Besides, SUMO-Bt-HepI also showed higher activities than Bt-HepI under 50℃. In addition, Ca²⁺ and Mg²⁺ have obvious promoting effect on the recombinant heparinase I, while Cu²⁺, Mn²⁺ and Zn²⁺ show certain inhibiting effect, suggesting that in addition to the well-known Ca²⁺ binding site, Mg²⁺ binding sites may aslo exist in the structure of B. thetaiotaomicron Heparinase I. [Conclusion] Recombinant Heparinase I in B. thetaiotaomicron using SUMO fusion system significantly improved its specific enzyme activity for potential production and application.

Abstract:[Objective] The purpose of the study was to isolate and identify Bdellovibrio-and-like organisms (BALOs) strains from seawater and increase the known species richness of BALOs. [Methods] Vibrio alginolyticus LF TCBS 15 was used as host bacterium to isolate BALOs from seawater samples with seawater agar double-layer plate method. The seawater samples were collected from Daya bay of Shenzhen, China. The morphology of the cells was observed by light microscopy and transmission electron microscopy. 16S rDNA was used for phylogenetic analysis to complete molecular identification. The prey range on 16 different strains and the effects of NaCl, pH and temperature on the growth of strain BALOs10 were determined by double-layer plate filter paper method.[Results] A novel strain, BALOs10, was isolated using V. alginolyticus LF TCBS 15 as prey. The plaques are round, transparent and the edges were smooth and tidy. The cells are curved (0.21-0.44 μm diameter×1.25-1.87 μm long) with a single polar flagellum. Optimum growth occurs at 35-37℃, pH 7-8 and in the presence of 2%-3% (W/V) NaCl. Prey range on 16 tested strains showed that strain BALOs10 lysed 9 different strains, corresponding to 56.3% of lysis rate, and was especially susceptive to Vibrio and Marinobacter. 16S rDNA sequence analysis indicates that BALOs10 was most closely related to Halobacteriovorax marinus SJ with 92.14% similarity, demonstrating that the strain may represent a novel species. Therefore, Halobacteriovorax sp. BALOs10 is proposed.[Conclusion] strain BALOs10 may represent a novel predator of genus Halobacteriovorax, which will enrich the species resource of BALOs and lay the material foundation for subsequent application and theoretical research.

Abstract:[Objective] Through sequencing and genetic function analysis of the transcripts of Dunaliella salina, we studied its response to salt stress. [Methods] The transcriptome of Dunaliella salina under 9% and 24% NaCl concentration was obtained and sequenced by Illumina platform. All the unigenes were finally obtained by sequence splicing, redundancy removing based on the sequence clustering software. Functional notation and cluster analysis of unigenes were performed by comparing the unigenes to the database. [Results] A total of 40682 unigenes were obtained, including 10905 in the NR database, 2768 in the NT database, 7261 in the Swiss-prot database, and 6499 in the COG/KOG database. By comparing cells grown under hyper-and hyposaline conditions, 717 genes (41.46%) increased expression and 1012 genes (58.5%) decreased expression. Through the data integration analysis, 60 different expression genes of Dunaliella salina under salt stress were excavated. [Conclusion] Salinity stress up-regulated key genes in photosynthesis. Our findings provide reference for dissecting molecular mechanisms of salinity tolerance in algae and higher plants.

Abstract:[Objective] Isomaltases IMA1 play key roles in full utilization of oligosaccharides containing α-1,6-O-glucosidic bonds. [Methods] We cloned, overexpressed, purified and characterized four isomaltases IMA1 from four strains of S. cerevisiae including three acidophilic ones. [Results] They showed similar pH and temperature dependence, but different kinetic parameters and thermostability. IMA1-A exhibited the highest binding affinity for α-MG (α-methylglucoside), turnover number, catalytic efficiency, and thermostability. Structure and sequence analysis revealed that even variation in two remote amino acids from the active residues and the substrate binding site could also lead to significantly different kinetic behavior and thermostability of isomaltases IMA1. [Conclusion] Our results will be useful for further investigation into the structure-function relationship of isomaltases IMA1.

Abstract:[Objective] To study the effect of banana pseudostem biochar on soil microorganisms in banana rhizosphere, we added the biochar into soil. [Methods] We mixed banana pseudostem biochar (BPB) with soil uniformly at the mass ratio of 0, 1%, 2% and 3%. Banana rhizosphere soil was separated after 3 months of pot culture. The bacterial community structure and abundance in rhizosphere soil were characterized by 16S rRNA high throughput sequencing. [Results] Increasing BPB application could increase soil organic matter, available potassium, available phosphorus contents and increase soil pH value, but decrease available nitrogen concentration. In total 2278 OTUs were obtained from 1% BPB samples, showing the greatest diversity of bacterial communities. When 3% BPB was applied to the soil, the relative abundance of Bacteroidates, Verrucomicrobia and Firmicutes increased significantly, whereas that of Actinobacteria, Acidobacteria and Gemmatimonade decreased significantly. Principal component analysis showed that soil bacterial communities were similar between 1% BPB and 2% BPB treated samples. [Conclusion] We changed the bacterial abundance and community structure in rhizosphere soil by applying different proportion of BPB, and the change in high proportion of BPB was more obvious.

Abstract:[Objective] Purpose of this work was to specifically screen the Gram positive (G⁺) bacteria of Bacillus that could encode the aconitate isomerase (AI) enzyme, to enrich our understanding of the distribution of AI and to provide theoretical and material basis for further research. [Methods] Through heat pretreatment of soil sample, plate cultivation by using ACO solid medium containing trans-aconitic acid (TAA) as the sole carbon source and the 16S rDNA sequences homologous analysis, the Bacillus target strains that encode AI can be isolated. [Results] We totally isolated 22 bacterial strains that could utilize TAA carbon from the ACO plate, and 16 of which were successfully classified as:2 strains of Bacillus megaterium, 7 strains of Bacillus aryabhattai, one Bacillus pumilus strain, and 6 undetermined Bacillus sp. strains; besides, we found that the AI-coding genes of these 16 Bacillus isolates were different from the already cloned ones in DNA sequence. [Conclusion] The species diversity of Bacillus bacteria encoding AI enzyme is rich, indicating the encoding ability in more G⁺ bacteria and updating the old opinion that considered AI distribution mainly in G^- hosts. Our research provided available microbial resource for further studies on the identification of AI gene as well as its biological significance.

Abstract:[Objective] It has been reported that phnolics contents could be changed by the inoculation with arbuscular mycorrhizal (AM) fungi. In the present study, the differentially expressed genes were screened, and the genes function was analyzed with the exogenous phenolic addition. [Methods] Poncitrus trifoliata (L.) inoculated with AM fungi was treated by exogenous phenolic. We obtained the differentially expressed bands by differential-display RT-PCR (DDRT-PCR), and then cloned the related genes and analyzed the bioninformation. [Results] A total of 154 differentially accumulated transcript-derived fragments were identified. After the sequencing and blast of 16 fragments, 9 genes were found related to putative functions which participated into the recognition of environment factors and internal signals, and the adjust of mycorrhizal formation. The bioinformatic analysis showed that the cDNA sequences of DD-11 was 1382 bp in length, containing a complete 1080 bp open reading frame and 454 amino acids were encoded, which was only expressed under mycorrhizal inoculation and exogenous phenolic. The molecular weight, theoretical pI of the DD-11 gene deduced protein were 49950.03 Da, 7.81, C₂₂₁₀H₃₄₂₇N₆₀₃O₆₅₇S₃₁. The deduced DD-11 protein included 22 Ser, 14 Thr, 7 Tyr, which became the phosphorylation sites of protein kinase. The secondary structure of the deduced DD-11 protein mainly included Alpha helix, Random coil, Extended strand, Beta turn, which were 21.81%, 57.71%, 20.48% and have no Beta turn. [Conclusion] The genes related to phenolic expressed in mycorrhizal formation were obtained in our study, which were related closely to the plants signal recognition. Our study provide a new perspective of molecular mechanism of phnolics occurring in mycorrhizal formation.

Abstract:[Objective] To investigate the role and its mechanism of the sporulation-related protein Srp1 in sexual reproduction and virulence in Cryptococcus neoformans. [Methods] Biolistic transformation method was conducted to construct the srp1Δ mutants and its complemented strains. Fungal virulence assay and bilateral mating assay were used to test the role of Srp1 in fungal development in C. neoformans. [Results] In comparison with the wild-type strain H99, the srp1Δ mutants had no significant difference in fungal virulence. Although the srp1Δ mutants can mate and form the dikaryotic hyphae in bilateral mating, the srp1Δ mutants have lost their ability to produce basidiospores. Fungal nuclei development assay showed that the nuclei in the bilateral mating of srp1Δ mutants failed to undergo meiosis after fusion, which can lead to failure of basidiospore formation in srp1Δ mutants. [Conclusion] The sporulation related protein Srp1 is not required for fungal virulence, but control sexual reproduction by regulating meiosis during mating in C. neoformans.