Abstract:Helicobacter pylori (H. pylori) is a strong risk factor for gastric disease ranging from chronic gastritis to gastric cancer. But the mechanisms underlying the pathogenesis of H. pylori are still not completely understood.The cytotoxin-associated gene A (CagA) of H. pylori, an important virulence factor and the only bacterial oncoprotein, is extensively studied. CagA is delivered into gastric epithelial cells via type IV secretion of H. pylori. Upon delivery, CagA perturbs multiple host signaling pathways by interacting with the host signaling molecules, resulting in cytopathic effects and subsequent cell transformation. Some animal experiments also provide in vivo evidence for the oncogenic capacity of CagA. In this review, recent advances in the structural property, delivery manner and pathogenesis of CagA are summarized, which we hope could better explain the CagA-mediated pathogenesis of Helicobacter pylori and provide directions for the future approach.

Abstract:DNA phosphorothioate modification was the first reported physiological modification in the DNA backbone. Five putative proteins encoded by the five-member dndABCDE gene cluster replaced the non-bridging oxygen in the sugar-phosphate backbone with a sulfur. Phosphorothioate modification occurrs in sequence-selective and Rp stereo-specific manner in diverse bacterial stains. In recent years, researchers have made systemic achievements in this area. To have a comprehensive understanding of this unusual modification, we reviewed the discovery and research progress in DNA phosphorothioate modification and also discussed opportunities and challenges in the future.

Abstract:Tuberculosis is still a global infectious disease. New drugs to shorten the course of treatment and new vaccines are the key points to control tuberculosis. The physiological study of Mycobacteria will contribute to the above-mentioned purposes. Polyphosphate plays an important role in the stress adaptation in bacteria. And there are two classes of enzymes:polyphosphate kinase and exopolyphosphatase involved in the polyphosphate metabolism to control the dynamic equilibrium of polyphosphate level in Mycobacteria. Present paper summarized the progress in metabolism and physiological roles of polyphosphate in Mycobacteria, to provide useful information for studying the physiological function of polyphosphate in Mycobacteria.

Abstract:[Objective] PDZ[Post-synaptic density-95 (PSD-95), Drosophilia tumor suppressor protein diskslarge-1 (DLG), the tight junction protein zonula occludentes 1 (ZO-1)] signal protein was encoded by Rv3194c gene from Mycobacterium tuberculosis, and its ability to adhere M. tuberculosis was studied.[Methods] Rv3194c protein was expressed in prokaryotic system. Rv3194c protein was separately incubated with hyaluronic acid, chondroitin sulfate and collagen Ι overnight at different temperature (37, 38, 39, 40℃). Then component changes of culture supernatant were tested by Western blot and ELISA.[Result] Western blot showed that Rv3194c protein expressed in prokaryotic system, with a molecular weight of about 35 kDa, was mainly in soluble form. Western blot showed that His-Rv3194c protein in supernatant of 39℃ experimental group was significantly less than that of other experimental groups (37, 38, 40℃)(***P<0.001). ELISA showed that hyaluronic acid, chondroitin sulfate and collagen Ι in supernatant of 39℃ experimental group was significantly less than that of other experimental groups (37, 38, 40℃)(***P<0.001).[Conclusion] For the first time it was affirmed that Rv3194c protein with detected activity of adhesions in this study will be targeted to the development of the new anti-M. tuberculosis drug.

Abstract:[Objective] To study the relationship between microbial community and degradation rate of rice straw, we compared and analyzed cellulose-decomposing ability, microbial community structures and shifts of microbial consortia F1 and F2.[Methods] We determined exoglucanase activity by 3, 5-dinitrosalicylic acid colorimetry. We determined content of cellulose, hemicellulose and lignin in rice straw by Van Soest method, and calculated degradation rates of rice straw by the weight changes before and after a 10-day incubation. We analyzed and compared the microbial communities and functional microbiology shifts by clone libraries, Miseq analysis and real time-PCR based on the 16S rRNA gene and cel48 genes.[Results] Total degradation rate, cellulose, and hemicellulose degradation rate of microbial consortia F1 were significantly higher than that of F2. The variation trend of exoglucanase activity in both microbial consortia F1 and F2 was consistent with that of cel48 gene copies. Microbial diversity of F1 was complex with aerobic bacteria as dominant species, whereas that of F2 was simple with a high proportion of anaerobic cellulose decomposing bacteria in the later stage of incubation. In the first 4 days, unclassified Bacillales and Bacillus were dominant in both F1 and F2. The dominant species and abundance became different after 4-day incubation, Bacteroidetes and Firmicutes were dominant phyla of F1 and F2, respectively. Although Petrimonas and Pusillimonas were common dominant species in F1 and F2, abundance of Petrimonas in F2 (38.30%) was significantly higher than that in F1 (9.47%), and the abundance of Clostridiales OPB54 in F2 increased to 14.85% after 8-day incubation.[Conclusion] The abundance of cel48 gene related with cellulose degradation rate and exoglucanase activity, and cel48 gene has the potential as a molecular marker to monitor the process of cellulose degradation. Microbial community structure has a remarkable impact on the degradation efficiency of straw cellulose, and Petrimonas, Paenibacillus, Bacillales, Clostridiales were vital species for microbial consortia F1 and F2 decomposing rice straw.

Abstract:[Objective] To isolate the fungus with phytotoxic activity from the gut of Pantala flavescens larvae. [Methods] Strain QTYC-51 was identified by morphological observation and 5.8S rDNA-ITS sequence analysis. Petri dish bioassay was used to test the phytotoxic activity of fermentation broth and monomer compounds of strain QTYC-51 on Echinochloa crusgalli and Amaranthus retroflexus. Bioactive components were isolated from ethyl acetate extracts via chromatographic methods, and the structures were determined by mass spectrum and nuclear magnetic resonance analyses.[Results] QTYC-51 was identified as Paraconiothyrium sp.. The fermentation broth had good phytotoxic activity on radical growth of E. crusgalli and A. retroflexus with the inhibition rates of 76.9% and 56.5%, respectively. Five monomer compounds were purified from the fermentation products, including 1,8-dihydroxyanthraquinone, 1-hydroxy-10-methoxy-dibenz[b,e]oxepin-6,11-dione, hydroxyvertixanthone, globosuxanthone and 1,3,6,8-tetrahydroxyanthraquinone. At the concentration of 100 μg/mL, compound globosuxanthone was found to possess obvious phytotoxic effects on radical growth of E. crusgalli and A. retroflexus with the inhibition rates of 94.1% and 79.0%, respectively, which were comparable to that of positive control 2,4-dichlorophenoxyacetic acid. Compound 1-hydroxy-10-methoxy-dibenz[b,e] oxepin-6,11-dione showed potent phytotoxic activity against E. crusgalli and A. retroflexus with inhibition rates of 50.3% and 58.6%, respectively.[Conclusion] Strain QTYC-51 could be potentially developed as a microbial herbicide.

Abstract:[Objective] To study the function of an RND family efflux pump encoded by MCHK_0866 and MCHK_0867 in Mesorhizobium huakuii 7653R. [Methods] Genetic organization of target genes was analyzed in genome. The change of growth was observed by measuring OD₆₀₀. Drug sensitivity was detected by minimal inhibitory concentrations; relative transcription level of target genes was measured by RT-PCR. Transcript regulation of the efflux pump was validated by bacterial one-hybrid system. [Results] Proteins encoded by MCHK_0866 and MCHK_0867 formed an RND family efflux pump. The OD₆₀₀ of growth curve reduced and it showed more sensitivity to nalidixic acid, tetracycline and SDS after disrupting the efflux pump. Genes relative transcription level increased in response to nalidixic acid treatment. Meanwhile, the downstream gene MCHK_0869 belongs to TetR family transcription factor and its expression product can interact with the promoter region of MCHK_0867. [Conclusion] The efflux pump is possibly associated with the transportation of nalidixic acid and affects rhizobial free-living growth. The pump is putatively regulated by a downstream local transcription factor.

Abstract:[Objective] To disrupt spa7074, which encodes a member of the TetR family transcriptional factors, in biocontrol strain Act12 and characterize the secondary metabolites in the mutant strain. [Methods] We disrupted the gene spa7074 by homologous recombination. The secondary metabolites of the mutant strain Δspa7074 and Act12 were detected by HPLC. The structure was analyzed by MS and NMR. [Results] Compared to the wild-type strain, the production of some unknown compounds in the mutant strain Δspa7074 increased obviously. We purified one of the compounds and identified as oligomycin D by MS and NMR analysis. [Conclusion] An oligomycin D-producing strain Δspa7074 was derived via genetic engineering.

Abstract:[Objective] Marine microorganisms have a great potential in producing biologically active secondary metabolites. In order to study the diversity and antimicrobial activity, we explored 9 sediment samples in different observation sites of Jiaozhou bay.[Methods] We used YPD and Z2216E culture medium to isolate bacteria from the sediments; 16S rRNA was sequenced for classification and identification of the isolates. Then, we used Oxford cup method to detect antimicrobial activities of the isolated bacteria against 7 test strains. Lastly, we selected 16 representatives to detect secondary-metabolite biosynthesis genes:PKSI, NRPS, CYP, PhzE, dTGD by PCR specific amplification.[Results] A total of 76 bacterial strains were isolated from Jiaozhou bay; according to the 16S rRNA gene sequence analysis. These strains could be sorted into 11 genera belonging to 8 different families:Aneurinibacillus, Brevibacillus, Microbacterium, Oceanisphae, Bacillus, Marinomonas, Staphylococcus, Kocuria, Arthrobacters, Micrococcus and Pseudoalteromonas. Of them 34 strains showed antimicrobial activity against at least one of the tested strains. All 16 strains had at least one function genes, 5 strains possessed more than three function genes.[Conclusion] Jiaozhou bay area is rich in microbial resources with potential in providing useful secondary metabolites.

Abstract:[Objective] Lipid transfer protein superfamily is involved in lipid transport and metabolism. This study aimed to construct mutants of three lipid transfer protein encoding genes in Mesorhizobium huakuii 7653R, and to study the phenotypes and function of mutations during symbiosis with Astragalus sinicus.[Methods] We used bioinformatics to predict structure characteristics and biological functions of lipid transfer proteins, and conducted semi-quantitative and fluorescent quantitative real-time PCR to analyze the expression levels of target genes in free-living and symbiotic conditions. Using pK19mob insertion mutagenesis to construct mutants, we carried out pot plant experiments to observe symbiotic phenotypes.[Results] MCHK-5577, MCHK-2172 and MCHK-2779 genes encoding proteins belonged to START/RHO alpha_C/PITP/Bet_v1/CoxG/CalC (SRPBCC) superfamily, involved in lipid transport or metabolism, and were identical to M. loti at 95% level. Gene relative transcription level of the three genes all increased compared to free-living condition. We obtained three mutants. Compared with wild-type 7653R, above-ground biomass of plants and nodulenitrogenase activity induced by the three mutants significantly decreased.[Conclusion] Results indicated that lipid transfer protein encoding genes of Mesorhizobium huakuii 7653R may play important roles in symbiotic nitrogen fixation, and the mutations significantly affected the symbiotic phenotypes. The present work provided a basis to study further symbiotic function mechanism associated with lipid transfer proteins from rhizobia.

Abstract:[Objective] To analyze the relationship between CRISPR/Cas system and drug-resistance, virulence. To investigate the effect of IS600 on the expression of CRISPR associated gene cse2 in Shigella.[Methods] CRISPR loci, CRISPR associated gene cse2, drug-resistant genes and virulent genes were detected by PCR in 33 Shigella strains; Trypan Blue counting test was used to detect bacterial virulence; Real-time PCR was used to detect relative mRNA expression of cse2; susceptibilities of Shigella strains were tested by agar diffusion method. Furthermore, we analyzed the relationship between CRISPR loci and drug-resistant genes, virulent genes. The effect of the IS600 on the expression of CRISPR associated gene cse2 was investigated.[Results] The mortality of Hela cells infected by Shigella with CRISPR1 loci was significantly lower (P<0.05) than those infected by Shigella without CRISPR1. The mRNA expression level of cse2 in group with IS600 was significantly (P<0.05) lower than that in group without IS600.[Conclusions] CRISPR loci were widely present in Shigella. Shigella without CRISPR1 has a higher pathogenicity. Due to the insertion of IS600, the mRNA expression level of cse2 was decreased in Shigella.

Abstract:[Objective] To understand the impact of genetic polymorphism of FnBPA-A on the immune biological characteristics of Staphylococcus aureus.[Methods] Sequence of FnBPA-A of Staphylococcus aureus isolated from bovine in Xinjiang was analyzed and 8 different genetic polymorphism eukaryotic recombinants of FnBPA-A were constructed. C57BL/6 mice were immunized with these recombinant plasmids and mice sera were collected. Level of the immune protection of immunized mice was compared.[Results] GS801, GS819 and GS856 were on the same branch; GW10-1, GW20-2, GY288 and GY309 belong to the same branch; GY278 was on a different branch. For the challenge experiment, GW20-2, GS801, GS819, GS856 and GY288 showed better protection.[Conclusion] The genetic polymorphism of FnBPA-A could significantly affect the immune protection of immunized mice.

Abstract:[Objective] A thermostable esterase EstZ1 from Bacillus sp. HJ14 able to degrade diethyl-phthalate (DEP) was heterologously expressed in Escherichia coli BL21(DE3) and characterized.[Methods] Full-length EstZ1 was obtained based on specific amplification and genome sequencing, and amino acid sequence of EstZ1 was analyzed. EstZ1 was expressed in Escherichia coli BL21(DE3) using the pEASY-E2 expression system. EstZ1 was purified to electrophoretic homogeneity by Ni²⁺-NTA metal chelating affinity chromatography, and the enzyme was characterized. The degradation products from DEP were detected by high-pressure liquid chromatography and electrospray ionization mass spectrometry.[Results] The 903 bp full-length EstZ1 encoded 300 amino acid residues (EstZ1:33.84 kDa). EstZ1 showed the highest identity of 98% with hormone-sensitive lipase (HSL)-like family in NCBI databases. The optimal temperature and pH was 50℃ and 9.0, respectively, with p-NP butyrate as the best substrate. Meanwhile, it was stable between 40 and 70℃, pH 7.0 to 9.5. Most of metal ions, chemical agents had little impact. DEP could partially be degraded by EstZ1 to its corresponding monoalkyl and alcohol.[Conclusion] Our findings may serve as reference for phthalate esters degradation.