Abstract:[Background] Methanogenic microbial consortia from coal geological environment can metabolize coal matrix to produce methane, which is of great significance for realizing the reutilization of coalbed methane (CBM) resources. [Objective] In order to prove the feasibility of the methods of culture preservation, community dynamics of methanogens was analyzed and the yeild of methane was tested during the preservation process. Meanwhile, the results would give theoretic basis for microbial enhanced CBM. [Methods] Three culture preservation methods involving glycerol/L-cysteine, eutrophication, and coal-basic salt method were compared at different temperatures. Microbial methanogenic activity in different preservations was tested by gas production. In addition, the compositions of microbial community in coal and basic salt preservation at 25 °C were studied by 454 high-throughput sequencing technology for 16S rRNA genes of bacteria and archaea. [Results] The preservation methods of 9 groups were compared, and the best culture preservation was the coal and basic salt preservation at 25 °C. Under this condition, the microbial methanogenic activity and the methane production were the highest. The yeild of methane was 12% to 25% and 24% to 73% using anthracite and lignite as carbon sources, respectively. In the coal and basic salt preservation test, the dominant bacteria and archaea in the early period were Pseudomonas and Methanosarcina at 25 °C, respectively. The structure of bacterial community changed dramatically with preservation time. The dominant bacteria changed to fermentative bacteria and acetogenic bacteria. The composition of dominant archaea was relatively stable. [Conclusion] The best preservation was coal and basic salt preservation at 25 °C, in which microbial activity of methanogens could be sustained at a better status and methanogens had good methane production ability.

Abstract:[Background] The unicellular green microalga Haematococcus pluvialis can synthesize astaxanthin which possess super antioxidant properties. H. pluvialis is an excellent microalga for commercial production of high-value natural astaxanthin. [Objective] The current study is to investigate effects of different nitrogen (N) sources on growth of H. pluvialis, with a view to develop the optimized N nutrition technology system for increasing biomass and astaxanthin yield of H. pluvialis. [Methods] Therefore, NaNO3 (NO3–-N) and NH4Cl (NH4+-N) were selected as two different N sources for the culture of H. pluvialis strain 797, respectively. At the same time, Hepes pH buffer was added into the media to keep pH stability. The measurements included pH changes of alga suspension culture media, cell growth rate at vegetative growth stage (“green stage”), chlorophyll content in cells and algal biomass. [Results] The results presented here demonstrated that the specific growth rate, biomass, chlorophyll a and b content in algal cells were higher in the NO3–-N media than in the NH4+-N media. Different N sources showed a significantly effect on pH in H. pluvialis suspension culture media. NH4Cl led to pH reduction in the culture media, whereas NaNO3 resulted in pH increase in the media. The addition of Hepes pH buffer successfully stabilized the pH in the culture media and thus, greatly promoted growth of H. pluvialis, with NH4Cl+Hepes showing much significant in this regard. It is the pH change in the algal suspension culture media caused by two different N sources that resulted in the obvious difference of H. pluvialis’s cell growth, biomass, and other features at “green stage” between the two N source treatments. [Conclusion] Addition of Hepes buffer into the media of either NO3–-N or NH4+-N could effectively stabilize the pH in microalga suspension culture media, and consequently, significantly promoted H. pluvialis’s cell growth and biomass accumulation at vegetative growth stage.

Abstract:[Background] Misgurin is an important component of nonspecific immune defense system in Misgurnus anguillicaudatus. It has broad spectrum and strong antimicrobial ability. Therefore, it is necessary to obtain large quantities of antimicrobial peptides. [Objective] To achieve high-efficient expression of misgurin. [Methods] Misgurin gene was cloned into expression vector pPIC9K in order to construct recombinant expression plasmid pPIC9K-misgurin. Using Sal I restriction endonuclease digest linearization scheme, the recombinant expression vector was integrated into the chromosome of Pichia pastoris SMD1168 by electric shock method. The positive single colonies was screened through minimal dextrose (MD) solid medium, which was inoculated MD liquid medium at 30 °C, 200 r/min shake flask for 96 h before transfer to BMMY liquid medium to induce expression, adding 5% methanol every 24 h. By comparing the diameter of bacteriostasis circle of Escherichia coli and Staphylococcus aureus, strain pPIC9K-misgurin-22 with the better antimicrobial activity was screened out and inoculated to a 100 L of fermentor induction for 48 h and then detected of activity. [Results] This active substance of pPIC9K-misgurin-22 strain was identified as misgurin by Tricine-SDS-PAGE protein gel detection and mass spectrometry analysis. In comparison with shaking flask fermentation, the bioavailability of Escherichia coli and Staphylococcus aureus in fermentor fermentation increased by 1.47 fold, 1.43 fold, respectively; misgurin has weak antimicrobial activity to Acinetobacter baumannii and Salmonella. It has no antimicrobial activity to Clostridium perfringens and probiotics (Bacillus subtilis, Enterococcus faecalis, Lactobacillus), and has no obvious hemolytic activity; When the temperature reaches 90 °C, the antimicrobial activity of fermentation broth is obviously weakened, the pH value of fermentation broth at 1.0?12.0, everyone has antimicrobial activity; The antimicrobial activity is weakened after adding trypsin, pepsin and protease K. [Conclusion] A strain producing misgurin is obtained, which has high expression and industrialized production potential in Pichia pastoris.

Abstract:[Background] A strain of microalgae was isolated from the tropical sea of Hainan province, and it has a high growth rate and strong adaptability. The microalga was identified as Chlorella vulgaris. [Objective] To improve the growth rate of C. vulgaris. [Methods] Based on the formulation of “Ningbo University 3#” medium, organic carbon of C6H12O6 and CH3COONa were added for the autotrophic culture, mixed culture and heterotrophic culture, respectively, and also, the mixotrophy medium of C. vulgaris was further optimized. [Results] The mixed culture method with 6 g/L CH3COONa showed the highest growth rate of C. vulgaris, comparing with heterotrophic and autotrophic culture method. The optimal formulation of mixotrophy medium was as followed as 6 g/L CH3COONa, 20 mg/L (NH4)2SO4-N, 5 mg/L NaH2PO4-P, 3 mg/L FeSO4-Fe, 1 mg/L Vitamin B1 and 0.000 5 mg/L Vitamin B12. The biomass (cell density) of the “mixotrophy medium” was up to 4.20×107 cells/mL by the end of the sixth day, being 2.30 times higher than the cell density of “Ningbo University 3#” medium. [Conclusion] The mixotrophic mode is optimal to the cultivation of C. vulgaris as compared with other two cultivation modes, and the optimized mixotrophic medium significantly enhanced the biomass (P<0.01).

Abstract:[Background] 2-Picolinic acid is highly toxic, carcinogenic, and long-term persisted in water bodies, thereby further endangers the environment. [Objective] to develop a technique that can efficiently and economically treat 2-picolinic acid-loaded wastewater. [Methods] We isolated a strain that could use 2-picolinic acid as the sole source of carbon, nitrogen and energy under aerobic conditions, and characterized its degradation. [Results] Based on the 16S rRNA gene sequence analysis, this strain was identified as a Chryseobacterium sp. and named as ZD2. When the initial concentration of 2-picolinic acid was 100, 200, 400, 600 and 800 mg/L, ZD2 completely degraded 2-picolinic acid within 10, 18, 22, 78 and 114 h, respectively. Zero-order kinetic model expressed the degradation behavior of 2-picolinic acid by ZD2 well. Between the concentration of 100 and 400 mg/L, the degradation rate constant increased with the increase in concentration and reached the maximum at 400 mg/L. Between the concentration of 600 and 800 mg/L, the degradation rate constant began to decrease, indicating an inhibitory effect. [Conclusion] The degradation efficiency of 2-picolinic acid by ZD2 demonstrates its potential in decontaminating wastewater containing 2-picolinic acid.

Abstract:[Background] Phaeodactylum tricornutum has emerged as a potential producer for biofuel feedstocks. Under stress conditions, the glycerolipids of P. tricornutum could be reconstructed to adapt to alterations of external environmental factors, concomitant with accumulation of triacylglycerols (TAGs), which can be converted to biofuels. Thus, mechanistic studies of glycerolipid remodeling in P. tricornutum under nitrogen stress are essential for deciphering mechanism of TAG biosynthesis and accumulation. [Objective] In order to confirm the origin of the accumulated acyl groups of TAG and the fate of the decreased acyl groups of multiple polar lipids, the alterations of fatty acids and glycerolipid components were studied in detail for P. tricornutum under nitrogen repletion and depletion conditions, which could provide insights into the response mechanism of glycerolipid of this alga subjected to nitrogen stress. [Methods] The fatty acids and glycerolipid components were determined qualitatively and quantitatively for P. tricornutum using high performance thin layer chromatography coupled with gas chromatography. [Results] Although the total content of glycerolipids remained unaltered under both nitrogen repletion and depletion conditions in P. tricornutum, the content of the individual glycerolipid class and the respective fatty acyl compositions varied notably. A prominent decrease in content of each polar glycerolipid was observed, accompanied by an increase in TAG amount, up to 57.8 mg/g, in stress-induced P. tricornutum. Based on variations of the contents of the fatty acyl groups comprising those glycerolipids, it was concluded that the lipid remodeling took place in multiple pathways. First, the saturated and monounsaturated fatty acids of TAG were significantly produced. In particular, a majority of 16:0 was concentrated into TAG through the de novo synthesis pathway and most of 16:1n7 was assembled into TAG through turnover of polar glycerolipids. In addition, a portion of EPA derived from polar glycerolipids were recycled into TAG as an acyl donor. Second, the polyunsaturated fatty acids degraded at expense of notable reductions of polar lipid components, i.e., 16:2n4, 16:3n4 and EPA. [Conclusion] When the TAG of nitrogen-deprived P. tricornutum accumulated up to 57.8 mg/g, the accumulated acyl groups of TAG derived from both the de novo synthesis pathway and the polar lipid turnover pathway and the contributions of each pathway were 48% and 52%, respectively. In addition, there were 54% of the decreased acyl groups of polar lipids to be incorporated into TAG and the other 46% of those entered into degradation reactions in P. tricornutum under nitrogen stress.

Abstract:[Background] Virioplankton is one of the most important components of microbial communities. The study on the temporal and spatial distribution of the virioplankton contributes to the protection and development of local microbial resources. [Objective] Virioplankton and planktonic bacteria of water samples were counted, aiming to investigate the virioplankton distribution in the Napahai plateau wetland. [Methods] Water samples from seven sites were obtained in December 2013 and September 2014. Virioplankton and planktonic bacteria abundance were determined using flow cytometry. Here we reported on virioplankton relationships with environmental factors, such as bacterial abundance, chlorophyll a concentration and other environmental factors. [Results] Virioplankton and planktonic bacteria abundance in rainy season was higher than that of in dry season at the seasonal distribution. Virioplankton abundance in dry season was influenced by bacteria abundance and chlorophyll a concentration; whereas virioplankton abundance in the rainy season was influenced by pH and temperature. [Conclusion] We found high virioplankton and planktonic bacteria abundance values. Virioplankton abundance was correlated with bacterioplankton abundance and chlorophyll a concentration in different seasons and sampling points. The phage, rather than the algal or phytoplankton viruses, was the dominant population in dry season of the Napahai plateau wetland.

Abstract:[Background] There are very rare researches on the biocontrol genes of Streptornyces sampsonii until now. Only two chitinase gene fragments were cloned, and there is no relevant study in the complete gene sequence of the chitinase. [Objective] To clone and prokaryotic expression of the chitinase gene ChiKJ40 of S. sampsonii KJ40, then purify recombinant protein and investigating antibacterial characters. [Methods] Firstly we cloned the chitinase gene ChiKJ40 from S. sampsonii KJ40 by PCR amplification, then ligated it into vector pET-32a and expressed in Escherichia coli BL21(DE3). We purified the recombinant chitinase using the His-tagged protein microscopy kit, determined the concentration of the crude enzyme solution and the purified enzyme solution by the Bradford protein concentration assay kit, measured the chitinase activity of the crude enzyme solution and the purified enzyme solution using the chitinase kit. Finally also checked antibacterial characters of the recombinant chitinase on the Cylindrocladium scoparium, Cryphonectria parasitica, Alternaria alternata and Rhizoctonia violacea. [Results] We induced expression of the ChiKJ40 gene (accession number: MF434484) through IPTG in E. coli, the recombinant chitinase size is 42 kD. There are no significant differences in production of protein using different concentrations of IPTG at 37 °C inducing 3 h. 0.2 mmol/L IPTG inducing 16 °C overnight, recombinant chitinase mainly existed in the form of soluble supernatant, small existed in inclusions precipitation. The chitinase activities of crude protein and purified protein were 0.080 U/mL and 0.046 U/mL, respectively. The specific activity of crude protein and purified protein were 0.041 U/mg and 0.115 U/mg, the purification ratio was 2.8, the rate of 57.5%. After treating with the purified protein, mycelium cells of C. scoparium, C. parasitica, A. alternata were segmented with inflating the mycelia, and myceliums of R. violacea were broken. [Conclusion] This study of ChiKJ40 provides biocontrol background of S. sampsonii and finds a new source for the chitinase genes, and lays a theoretical foundation for its application.

Abstract:[Background] Chitin is the second most abundant natural resource next to cellulose. Chitinase can catalyze chitin into chitosan oligosaccharide which achieved high value utilization of waste. [Objective] The chitinase gene of Vibrio sp. GR52 was cloned and expressed in Escherichia coli, and the properties of recombinase were characterized. [Methods] The chitinase gene chiGR52-1 was cloned from the genomic DNA of Vibrio sp. GR52 and the recombinant strain BL21 (DE3)/pET22b-chiGR52-1 was constructed. The recombinase rchiGR52-1 was purified by Ni-NTA affinity chromatography and the enzymatic properties were studied. [Results] The optimum catalytic pH of rchiGR52-1 was 6.0. It was stabled in the pH range of 5.0 to 10.0 and could maintain more than 85% of its relative activity after incubation at 37 °C for 1 hour. The optimum catalytic temperature of the enzyme was 50 °C and 60% of enzyme activity was remained after incubation at 50 °C for 1 hour. At the concentration of 1 mmol/L, Cu2+ and Ca2+ had stimulation on rchiGR52-1, while Hg+ had significant inhibition. At the concentration of 5 mmol/L, Ni+ stimulated the chitinase, while Mn2+, Co2+, Li+, Fe2+, Hg+ and SDS inhibited the enzyme. The kinetic parameters Km, Vmax and kcat of rchiGR52-1 were 0.85 mg/mL, 0.19 μmol/(mL·min) and 7.02 s?1, respectively. Substrate specificity analysis showed that rchiGR52-1 could catalyze chitin specifically. [Conclusion] Recombinant chitinase has good enzymatic properties which provide a theoretical foundation for chitinase applications.

Abstract:[Background] Root-associated fungi (RAF) are essential for seedling survival, establishment and growth. However, little is known about the identity and ecological characteristics of RAFnaturally established on seedling roots. [Objective] To reveal phylogenetic diversities and ecological types of RAF naturally established on nursery grown seedling of Aquilaria sinensis and Dalbergia odorifera and to evaluate the host effects on RAF community structuring. [Methods] root samples were collected to extract DNA, from which fungal ITS region was amplified, cloned and sequenced by using both universal and specific fungal primer pairs. Phylogenetic placements of fungi were inferred by ITS sequence analysis. The putative trophic modes and guilds of RAF were assigned by functional analysis with the FUNGuild software. Effects of plant species, height, basal diameter and leaf area on root-associated fungal species composition were determined by non-metric multidimensional scaling (NMDS) analysis. [Results] A. sinensis and D. odorifera seedlings grown in nurseries were naturally colonized by a highly diverse suite of RAF, including Mucoromycota (51%), Ascomycota (43%) and Basidiomycota (6%). These RAF can be assigned to multiple trophic modes and guilds including symbiotroph (29 species) representing by Glomeromycetes sp. 2, Rhizophagus irregularis, and saprotroph (5 species) representing by Talaromyces pinophilus and Rhizopycnis vagum. In addition, two fungal species, Mycoleptodiscus sp. and Fusarium phaseoli were assigned to pathotroph, while ecological characteristics of 15 species are unclear. NMDS analysis indicated that the effects of host plant species, plant height, basal diameter and leaf area of seedling on root-associated fungi were not significant. However weak effect of plant height was found in Arbuscular mycorrhiza fungi fungal community. [Conclusion] The RAF inoculum is high in culture media of this nursery that seedlings were associated with phylogenetically distant and multiple trophic modes and guilds of fungi. Diversity of AMF might be underrepresented when universal primers such as ITS1F/ITS4 were applied to investigate the diversity of RAF community.

Abstract:[Background] Strawberry is an important horticultural crops in our country and in the world, but mainly cultivated with annual cropping system in the strawberry area so as to cause continuous cropping phenomenon. Soil-borne disease caused by fungal disease is a major problem in strawberry cultivation. [Objective] Strawberry rhizosphere soils were used as material to explore effects of dazomet fumigation and biological fertilizer on strawberry soil fungal diversity under replant conditions, so as to provide theoretical basis for control of strawberry continuous cropping obstacle. [Methods] Replant soils before (A) or after dazomet fumigation (B) were collected, genomic DNA was extracted, and PCR amplification was made to establish libraries, as the same treatments with the soil samples at stage of flowering with dazomet fumigation (C1) or dazomet fumigation and biological fertilizer (C2). In this study, the fungal ITS1 region was sequenced by Illumina high-throughput sequencing technology on Hiseq 2500 platform, and related biological analysis was conducted to explore the changes of soil fungal abundances, diversities and structures. [Results] A total of 723 fungal OTUs were obtained from 4 strawberry rhizosphere soil samples, among them, Ascomycota and Basidiomycota were the dominant fungi. The fungal abundances and diversities of replant soils were decreased with dazomet fumigation, while the fungal abundances were increased and the fungal diversities were decreased with dazomet fumigation and biological fertilizer. At phylum level, the proportion of Basidiomycota was decreased and Ascomycota was increased with dazomet fumigation, while both of them were increased with dazomet fumigation and biological fertilizer. The analysis of dominant fungal community shows that the fungal proportions were decreased in terms of Acremonium, Aspergillus, Funneliformis, Fusarium, Talaromyces, Alternaria and increased in terms of Malassezia, Ophiocordyceps, Pleurotus with dazomet fumigation, while which were decreased in terms of Funneliformis, Fusarium, Mortierella, Lecanicillium, Dactylella and increased in terms of Aspergillus, Penicillium, Talaromyces, Simplicillium. [Conclusion] In conclusion, fungal diversity was decreased, pathogenic fungals were limited and benificial fungals were boosted with dazomet fumigation and biological fertilizer in strawberry cropping soil. Our results indicated a combination of dazomet fumigation and biological fertilizer could more effectively reduce the strawberry disease.

Abstract:[Background] Traditional fermented sweet paste has a delicious taste and unique flavor but little is known about microbial diversity during the fermentation by using high-throughput sequencing. [Objective] To analyze the microbial community structure and dynamic succession during natural fermentation of traditional sweet paste. [Methods] High-throughput sequencing was used to study the microbial diversity during natural fermentation of traditional sweet paste. [Results] In this study, 100 genera fungi and 432 genera bacteria were identified. Aspergillus is the dominant fungal species (≥76.96%) during the first 90 days of fermentation. Zygosaccharomyces is the dominant fungal species (≥85.42%) during 120 to 180 days of fermentation. Moreover, Bacillus is the primary bacterial genus (98.07%) during the first 43 h of fermentation and thereafter. In addition, the predominant bacteria in the koji and mash included Staphylococcus, Ralstonia, Pantoea, Burkholderia, Pediococcus, Sphingomonas, Bifidobacterium, Faecalibacterium, Kocuria and Lactobacillus. [Conclusion] The dominant florae of traditional sweet paste at different stages were determined. The results provide a theoretical basis for the study of the influence of different microorganisms on the flavor formation of the sweet paste.

Abstract:[Background] Campylobacter is a Gram-negative microaerobic bacterium. It is a major foodborne pathogen and causes human gastroenteritis worldwide. [Objective] This study was to isolate, identify and molecular type Campylobacter from animal in six different areas of Anhui province. [Methods] The strains were identified by morphological and culture characteristics, biochemical tests and PCR. Seven Campylobacter housekeeping genes including aspA, glnA, gltA, glyA, pgm, tkt and uncA were amplified by PCR and sequenced, and then the sequences of genes were analyzed using multilocus sequence typing (MLST) database and evolutionary tree. [Results] There were a total of 42 strains of Campylobacter were isolated from six areas. The isolates had more consistent morphological characteristics and similar biochemical properties. MLST results show that 32 sequence types, 9 new sequence types (including 8190, 8222, 8223, 8831, 8833, 8841, 8832, 8834 and 8843) and 6 new allelic profiles (including glnA606, glnA607, gltA518, glyA680, pgm863 and uncA541) were found. Evolutionary tree results show that C. jejuni and C. coli genetic relationship is very different and belong to two groups, which have five branches and three branches, respectively. [Conclusion] C. jejuni and C. coli have rich genotypes in Anhui province, and there is no obvious dominant genotype. From the perspective of genetic variation, C. jejuni is more complex and diverse, while C. coli is relatively conservative.

Abstract:[Background] Enterotoxigenic Escherichia coli K99 is one of the main pathogens causing diarrhea in piglets. So far the treatment of this disease mainly depends on antibiotics, but the long-term use of antibiotics has caused the increase of drug resistance, therefore, new product is urgently needed to solve this problem. [Objective] A lytic phage against Enterotoxigenic Escherichia coli K99 was isolated from the manure of livestock and identified, and its biological properties were assayed as well. The preventive and therapeutic effects of the bacteriophage on Enterotoxigenic Escherichia coli K99 infection were examined in mice. [Methods] The lytic phage was isolated by the double-layer agar culture method and the phage plaque was checked. Morphology of the phage was observed by transmission electron microscopy and the type of nucleic acids was also identified after the phage purification. The biological parameters including the thermal stability, pH stability and optimal multiplicity of infection (MOI) were assayed separately. The inhibitory effects of the bacteriophage on Escherichia coli infection in mice was tested by in vivo experiments with mice. [Results] A lytic phage (designated as ФK99-1) against Enterotoxigenic Escherichia coli K99 was successfully isolated from livestock manure. ФK99-1 belongs to family Myoviridae according to the electron microscopy and type identification of nucleic acid. The phage could withstand the temperature up to 50 °C and maintain stable titer under pH 3.0–10.0. The optimal MOI of ФK99-1 was 0.000 01, and the burst size was 108 333 PFU/cell. Application of ФK99-1 in mice which were infected with Enterotoxigenic Escherichia coli K99 could alleviate the related symptoms and viscera lesions. [Conclusion] The isolated phage, ФK99-1, belongs to lytic phage of Myoviridae, which shows strong adapting capacity to various temperature and acid-base environments. At the same time the isolated phage shows inhibitory effect on E. coli infection in mice.

Abstract:[Background] Adenosine triphosphatase (ATPase) controls DNA replication initiation and host response switch to pathogenic microbes. Recent genomics studies on aquatic animal iridoviruses have suggested that they share a gene encoding ATPase. Here, the function of the core gene in iridovirus will be first elucidated. [Objective] Andrias davidianus ranavirus (ADRV), which belong to the family Iridoviridae, is the viral pathogen of mass mortality in Chinese giant salamander—the world’s largest living amphibians. In order to identify virus gene and test the gene expression effects on virus replication and host cell, an ADRV gene, ADRV-96L was cloned, expressed and function analyzed. [Methods] Sequence data were analyzed using PredicProtein software. pET32a/His ADRV-96L recombinant prokaryotic expression plasmid was constructed, and the protein was expressed in Escherichia coli DE3 using IPTG induction, purified by Nickel Resin and eluted with imidazole. ATPase activity of the purified ADRV-96L recombinant protein was measured by the production of inorganic phosphorus (Pi) using molybdenum blue spectrophotometry method. Stable transfected cells were constructed and characterized. Then, the effects of ADRV-96L were analyzed by one-step multiplication curve for virus replication and estimating the growth rates of transferred cell, respectively. [Results] The multiple sequence alignments provided that ADRV-96L was characterized by the presence of the conserved AAA-ATPase (The ATPase associated with a variety of cellular activities) domains (20–159 residues) that contain Walker A and Walker B motifs, and two highly conserved arginines. The recombinant prokaryotic plasmid expressed a 52 kD fusion protein containing ADRV-96L, and the protein has ATPase activity 4.68 U/mg. The results show that ADRV-96L has no measurable impact on the ADRV replication, but it has been shown to promote cell growth. [Conclusion] Andrias davidianus ranavirus 96L gene (ADRV-96L) encodes an ATPase that is involved in cell proliferation and growth.

Abstract:[Background] People tend to study endophytic actinomycetes to discover new compounds due to the increasing difficulty to obtain new compounds from soil actinomycetes and antibiotic abuse causing increased resistance of pathogens. [Objective] We explored the diversity of endophytic actinomycetes isolated from toxic plants in Xishuangbanna tropical rainforest, and isolated strains with antibacterial potential to develop new drugs. [Methods] We analyzed communities of endophytic actinomycetes from three toxic plants, Antiaris toxicaria Lesch., Alangiumchinense (Lour.) Harms and Lantana camara L. by Illumina HiSeq high-throughput sequencing and culture-dependent approach. Antimicrobial activity was screened by disk diffusion test and seven synthetic genes were detected by PCR amplifications. [Results] Strains from 3 toxic plants were identified at the phylum level with 2 phyla of the Archaea, 18 phyla and the tentative RsaHF231, WD272 of the bacteria. And 30 genera of actinomycetes were detected at the genus level. Both the microbial community structure of Alangiumchinense (Lour.) Harms and Lantana camara L. were more abundant than Antiaris toxicaria Lesch. In total 34 strains of 11 genera were isolated from culture-dependent approach. Most of the genus detected by high-throughput could not be obtained through pure culture. The antibacterial activity results showed that Streptomyces had broad spectrum antimicrobial activity. The detection rates of NRPS and PKS in Streptomyces were significantly higher than other compounds synthetic gene. [Conclusion] The diversity of endophytic actinomycetes in toxic plants is very rich. Actinomycetes in the toxic plants secondary metabolites synthesis have enormous potential to provide an abundant microbial resource for the development of biological pesticides and antibiotics.

Abstract:[Background] Microorganisms have a role in spore germination, hyphal growth, mycorrhiza formation and basidiocarp development of mycorrhizal fungi. [Objective] Microorganisms of eight basidiocarps of Thelephora ganbajun which were collected from Songming County, Kunming and Lufeng County, Chuxiong Yi autonomous Prefecture, Yunnan Province, SW China was studied, which lay a foundation for follow-up studies on the interaction between microorganisms and T. ganbajun. [Methods] The dilution plate method was carried out to obtain the microbial isolates from basidiocarps of T. ganbajun. The difference of microbial quantity of T. ganbajun which were collected from different sites was analyzed by t-test. Diversity of microorganisms was analyzed by 16S rRNA gene and ITS amplification, sequencing and establishment of phylogenetic tree. [Results] A total of 282 culturable bacterial isolates of eight basidiocarps from Songming County and Lufeng County belonged to 15 species and 12 genera in 2 phyla. 80% of bacteria were affiliated to the Proteobacteria and dominated by γ-Proteobacteria and Pseudomonas. The other 20% were affiliated to the Bacteroidetes. A total of 114 culturable fungal isolates belonged to 10 species and 10 genera in 2 phyla. 62% of fungi were affiliated to the Ascomycota, dominated by Lophiostoma which was only isolated from Lufeng County. 38% of fungi were affiliated to the Basidiomycota, dominated by Asterotremella. [Conclusion] There was no significant difference in the number of CFU of bacteria (p=0.22) and fungi (p=0.65) among basidiocarps of T. ganbajun which were collected from different sites. All bacteria isolates were dominated by γ-Proteobacteria and Psedomonas. Fungi were dominated by Basidiomycota and Asterotremella in Songming, whereas fungi were dominated by Ascomycota and Lophiostoma in Lufeng.

Abstract:[Background] The Stromaticum clade of Trichoderma was defined by Samuels et al. in 2012, including 9 species. However, only 3 species, i.e. T. stromaticum, T. vermipilum and T. floccosum were reported in China. [Objective] Two Trichoderma species, i.e. T. ivoriense and T. barbatum, are newly recorded in China. [Methods] They were collected from Beijing and Shandong by the selective medium THSM, and identified by the translation elongation factor 1 alpha (TEF1-α), RNA polymeraseⅡsubunit 2 (RPB2) and observation of morphological characteristics. [Results] By phylogenetic analyses of TEF1-α and RPB2, two Trichoderma strains were close to T. ivoriense and T. barbatum respectively; however, morphologically they are markedly different from typical species of T. ivoriense or T. barbatum. Therefore these two strains were identified as T. ivoriense and T. barbatum, or relative species of them. [Conclusion] Two species of the genus Trichoderma were newly discovered in China and were described, they belong to the Stromaticum clade. The numbers of species in that clade add up to five.

Abstract:Carboxysome is an organelle-like structure that can concentrate CO2 from the environment. It exists in some autotrophic microorganisms and can promote cell growth. Among nitrogen-removal microorganisms, nitrifying bacteria, anaerobic ammonia oxidation (Anammox) bacteria and some denitrifying bacteria are autotrophic microorganisms. Understanding the composition, structure and function of carboxysome is helpful to reveal the growth of autotrophic microorganisms and to optimize the process of biological nitrogen-removal. Based on the literatures and our researches, the composition, structure, function and detection of carboxysome in autotrophic bacteria are summarized in this paper.

Abstract:Lincomycin, produced by Streptomyces lincolnensis, is a lincosamide antibiotic, which inhibits the protein biosynthesis of bacteria, and widely used to treat the infectious diseases caused by the Gram positive bacteria in clinic. Lincomycin biosynthetic gene cluster (lmb) was cloned and sequenced. Biosynthesis and regulation of lincosamide and propylproline were extensively elusive with the breakthrough of S-functionalization of lincomycin. This article reviewed the recent proceedings and achievements on the process of lincomycin biosynthesis.

Abstract:Endophytic fungi from desert and grassland areas are an ecologically important, though poorly studied, class of fungi in terms of chemistry with potentially and vitally ecological or biological roles. Though limited secondary metabolites isolated up to date compared with those of other fungi, there still are many secondary metabolites with novel structure features and potentially biological activities purified from this member of special fungi. In this paper, the different types of secondary metabolites isolated from endophytic fungi inhabiting in grassland and desert plants were reviewed from the perspectives of structural types (alkaloids, polyketides, phenolic acids, terpenoids and cyclic peptides), pharmacological activities (antiviral, immunoregulation, promotion of bone marrow proliferation, cytotoxicity and Hsp90 inhibitor activity) and ecological roles (insecticidal, antifeedant and animal neurotoxic activity). This review might provide a solid foundation for further chemical and biological investigation of this special family of endophytic fungi. Finally, the existing problems in this field are analyzed and discussed, and the prospect is also put forward.