Production of the Quorum-Sensing Molecules N-Acylhomoserine Lactones by Endobacteria Associated with Mortierella alpina A-178

Gram‐negative bacteria communicate with one another using N‐acylhomoserine lactones (AHLs) as signaling molecules. This mechanism, known as quorum sensing (QS), is needed to develop pathogenicity, as well as symbiotic interactions with eukaryotic hosts, such as animals and plants. Increasing evidence indicates that certain bacteria, namely endobacteria, also inhabit fungal cells and establish symbiotic relationships with their hosts. However, it has not been clear whether bacterial QS acts in developing the relationships. Here we describe the isolation and identification of N‐heptanoylhomoserine lactone and N‐octanoylhomoserine lactone from the culture broth of the zygomycete fungus Mortierella alpina A‐178. This suggested the presence of endobacteria in the fungus, as was confirmed by PCR, fluorescence in situ hybridization, and transmission electron microscopy. Two major bands obtained by PCR‐denaturing gradient gel electrophoresis showed sequence identity to genes in the β‐proteobacterium Castellaniella defragrans (100 %) and the Gram‐positive bacterium Cryobacterium sp. (99.8 %). The production of AHLs depended on the presence of endobacteria and was induced in response to the increase in the concentration of AHLs, suggesting that the bacterium conducts AHL‐mediated QS in the fungus. This paper is the first to report the production of AHLs by endofungal bacteria and raises the possibility that QS plays roles in the development of fungus–endobacterium symbiosis.     

Isolation and Molecular Characterization of Biofouling Bacteria and Profiling of Quorum Sensing Signal Molecules from Membrane Bioreactor Activated Sludge

The formation of biofilm in a membrane bioreactor depends on the production of various signaling molecules like N-acyl homoserine lactones (AHLs). In the present study, a total of 200 bacterial strains were isolated from membrane bioreactor activated sludge and screened for AHLs production using two biosensor systems, Chromobacterium violaceum CV026 and Agrobacterium tumefaciens A136. A correlation between AHLs production and biofilm formation has been made among screened AHLs producing strains. The 16S rRNA gene sequence analysis revealed the dominance of Aeromonas and Enterobacter sp. in AHLs production; however few a species of Serratia, Leclercia, Pseudomonas, Klebsiella, Raoultella and Citrobacter were also identified. The chromatographic characterization of sludge extract showed the presence of a broad range of quorum sensing signal molecules. Further identification of sludge AHLs by thin layer chromatography bioassay and high performance liquid chromatography confirms the presence of C4-HSL, C6-HSL, C8-HSL, 3-oxo-C8-HSL, C10-HSL, C12-HSL, 3-oxo-C12-HSL and C14-HSL. The occurrence of AHLs in sludge extract and dominance of Aeromonas and Enterobacter sp. in activated sludge suggests the key role of these bacterial strains in AHLs production and thereby membrane fouling.     

海洋放线菌JWH-09代谢产物中农用抗菌组分的分离纯化及结构鉴定

从134株海洋微生物中筛选得到放线菌JWH-09,其发酵代谢产物具有较好的抗植物病原真菌活性.根据生物活性跟踪测试,采用硅胶柱层析、薄层层析以及制备高效液相色谱等技术对JWH-09发酵产物中的抗菌活性组分进行分离纯化,得到了一种抗真菌活性化合物JWH-09-A,经ESI-MS、EI-MS、1H NMR、13C NMR、2D NMR等谱图分析,确定该活性物质的结构为1-菲羧,1,2,3,4,4a,9,10,10a-八氢-1,4a-二甲基-7-(1-甲基乙基),俗名脱氢松香酸(dehydroabietic acid).经盆栽试验证明该化合物对水稻纹枯病、黄瓜白粉病和灰霉病具有较好的防治效果.     

Production,Biochemical Characterization and Detergents Application of Keratinase from the Marine Actinobacterium Actinoalloteichus sp. MA-32

The present study is the first report on poultry feathers as a novel, inexpensive substrate for the production of a thermo‐ and detergent stable keratinase from a marine actinobacterium belonging to the genus Actinoalloteichus. Medium composition and culture conditions for the keratinase production by Actinoalloteichus sp. MA‐32 were optimized using two statistical methods: Plackett–Burman design was applied to find the key ingredients and conditions for the best yield of enzyme production and central composite design used to optimize the concentration of the five significant variables: whole chicken feather, soy flour, MgSO₄·7H₂O, KH₂PO₄ and NaCl. The medium optimization resulted in a 19.30‐fold increase with a 31.99 % yield with a specific activity of 3842.57 U mg^?1 and the molecular weight was estimated as 66 kDa. The enzyme was optimally active at pH 8–10 and temperature 50–60 °C and it was most stable up to pH 12 and 10–14 % of NaCl concentration. The enzyme activity was reduced when treated with Hg²⁺, Pb²⁺, Tween‐80, 1,10‐phenanthroline and EDTA and stimulated by Fe²⁺, Mg²⁺, Cu²⁺, Ca²⁺, Ni²⁺, Mn²⁺, SDS, ethoxylated (9.5EO) octylphenol, DMSO, sodium sulfite and β‐mercaptoethanol. The keratinase exhibited a significant stability and compatibility with most of the tested commercial laundry detergents, demonstrating its feasibility for inclusion in laundry detergent formulation. These results suggest that this extracellular keratinase may be a useful alternative and eco‐friendly route for handling the abundant amount of waste feathers or for applications in detergent formulation and other industrial processes.     

Characterization by pyrolysis-gas chromatography-mass spectrometry of the insoluble organic residues derived from the hydrogenation of Tasmanites sp. oil shale

The insoluble organic residues from the hydrogenation of Tasmanites sp. oil shale have been characterized by Curie point pyrolysis combined with gas chromatography and computerized gas chromatography-mass spectrometry. The resulting pyrograms show that very little, if any, hydrogenation of the residues occurs. Rather, the initial step in the sequence is pyrolysis of the residue followed by hydrogenation of the liquid products. Major compounds identified from the pyrograms of the residues include alkene/alkane doublets, mixtures of diterpenoids and mono- di- and triaromatic compounds. Changes in the distribution of these compounds with changes in reaction conditions are discussed.     

大庆乙烯副产C9馏分的GC及GC／MS分析

本文通过对大庆乙烯副产的C9馏份油进行GC及GC／MS分析确定了该C9的组成。共检出38种物质，其中不饱和烃约为46．7％。其结果与C9馏份的实沸点切割、溴值测定及^1H-NMR分析结果相吻。     

Characterization of linear low‐density polyethylene/poly(vinyl alcohol) blends and their biodegradability by Vibrio sp. isolated from marine benthic environment

Increasing amounts of plastic waste in the environment have become a problem of gigantic proportions. The case of linear low‐density polyethylene (LLDPE) is especially significant as it is widely used for packaging and other applications. This synthetic polymer is normally not biodegradable until it is degraded into low molecular mass fragments that can be assimilated by microorganisms. Blends of nonbiodegradable polymers and biodegradable commercial polymers such as poly (vinyl alcohol) (PVA) can facilitate a reduction in the volume of plastic waste when they undergo partial degradation. Further, the remaining fragments stand a greater chance of undergoing biodegradation in a much shorter span of time. In this investigation, LLDPE was blended with different proportions of PVA (5–30%) in a torque rheometer. Mechanical, thermal, and biodegradation studies were carried out on the blends. The biodegradability of LLDPE/PVA blends has been studied in two environments: (1) in a culture medium containing Vibrio sp. and (2) soil environment, both over a period of 15 weeks. Blends exposed to culture medium degraded more than that exposed to soil environment. Changes in various properties of LLDPE/PVA blends before and after degradation were monitored using Fourier transform infrared spectroscopy, a differential scanning calorimeter (DSC) for crystallinity, and scanning electron microscope (SEM) for surface morphology among other things. Percentage crystallinity decreased as the PVA content increased and biodegradation resulted in an increase of crystallinity in LLDPE/PVA blends. The results prove that partial biodegradation of the blends has occurred holding promise for an eventual biodegradable product. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2012     

Biosorption of mercury and lead by aqueous Streptomyces VITSVK9 sp. isolated from marine sediments from the bay of Bengal,India

Toxic heavy metals are increasingly accumulating in the environment worldwide and are considered to be life threatening contaminants. The biosorption of mercury and lead by marine actinomycetes isolated from marine sediment collected from the Bay of Bengal coast of Puducherry, India, was evaluated. The maximum tolerance concentration (MTC) of Streptomyces sp. was determined by a well diffusion method and a broth dilution method. The effects of the initial metal ion concentration, the pH and the biomass dosage on the biosorption of mercury and lead ions were investigated. The MTC of the isolate to metals was 200 mg·L^-1 for mercury and 1800 mg·L^-1 for lead. At neutral pH, the isolate had a maximum biosorption of metal ions of 200 mg·L^-1 and 150 mg·L^-1 for mercury and lead respectively. Fourier transform infrared (FTIR) absorption spectra showed the chemical interactions between the functional groups in the biomass such as hydroxyl (-OH), amine (-NH₂), carboxyl (-COOH) and the metal ions. The isolate was further characterized by molecular taxonomy and identified as a member of the genus Streptomyces. Based on the phenotypic and phylogenetic analysis, the strain was classified as a new species of the genus Streptomyces and designated as Streptomyces VITSVK9 sp. (HM137310). A blast search of the 16S rDNA sequence of the strain showed the most similarity (95%) with Streptomyces sp. A515 Ydz-FQ (EU384279). Based on the results, it can be concluded that this marine Streptomyces could be used as a biosorbent for the removal of heavy metal ions from aqueous environments.     

Dual Bioactivities of Essential Oil Extracted from the Leaves of Artemisia argyi as an Antimelanogenic versus Antioxidant Agent and Chemical Composition Analysis by GC/MS

The study was aimed at investigating the antimelanogenic and antioxidant properties of essential oil when extracted from the leaves of Artemisia argyi, then analyzing the chemical composition of the essential oil. The inhibitory effect of the essential oil on melanogenesis was evaluated by a mushroom tyrosinase activity assay and B16F10 melanoma cell model. The antioxidant capacity of the essential oil was assayed by spectrophotometric analysis, and the volatile chemical composition of the essential oil was analyzed with gas chromatography-mass spectrometry (GC/MS). The results revealed that the essential oil significantly inhibits mushroom tyrosinase activity (IC₅₀ = 19.16 mg/mL), down-regulates B16F10 intracellular tyrosinase activity and decreases the amount of melanin content in a dose-dependent pattern. Furthermore, the essential oil significantly scavenged 2,2-diphenyl-1-picryl-hydrazyl (DPPH) and 2,2′-azino-bis (3-ethylbenzthiazoline- 6-sulphonic acid) ABTS radicals, showed an apparent reduction power as compared with metal-ion chelating activities. The chemicals constituents in the essential oil are ether (23.66%), alcohols (16.72%), sesquiterpenes (15.21%), esters (11.78%), monoterpenes (11.63%), ketones (6.09%), aromatic compounds (5.01%), and account for a 90.10% analysis of its chemical composition. It is predicted that eucalyptol and the other constituents, except for alcohols, in the essential oil may contribute to its antioxidant activities. The results indicated that essential oil extracted from A. argyi leaves decreased melanin production in B16F10 cells and showed potent antioxidant activity. The essential oil can thereby be applied as an inhibitor of melanogenesis and could also act as a natural antioxidant in skin care products.     

LC-ESI-QTOF-MS/MS Analysis,Cytotoxic, Antiviral,Antioxidant, and Enzyme Inhibitory Properties of Four Extracts of Geranium pyrenaicum Burm. f.: A Good Gift from the Natural Treasure

This study focused on the biological evaluation and chemical characterization of Geranium pyrenaicum Burm. f. Different solvent extracts (hexane, ethyl acetate, methanol, and water extracts) were prepared. The phytochemical profile, antioxidant, and enzyme inhibitory activity were investigated. Cytotoxicity was assessed using VERO, FaDu, HeLa and RKO cells. The antiviral activity was carried out against HSV-1 (Herpes simplex virus 1) propagated in VERO cell line. The aqueous extract, possessing high phenolic content (170.50 mg gallic acid equivalent/g extract), showed the highest reducing capacity (613.27 and 364.10 mg Trolox equivalent/g extract, for cupric reducing antioxidant capacity and ferric reducing antioxidant power, respectively), radical scavenging potential (469.82 mg Trolox equivalent/g extract, against 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)), metal chelating ability (52.39 mg ethylenediaminetetraacetic acid equivalent/g extract) and total antioxidant capacity (3.15 mmol Trolox equivalent/g extract). Liquid chromatography-electrospray ionization-quadrupole time-of-flight-mass spectrometry (LC-ESI-QTOF-MS/MS) alloved to tentatively identify a total of 56 compounds in the extracts, including ellagitannins, gallic acid and galloyl derivatives amongst others. The ethyl acetate extracts substantially depressed cholinesterase enzymes (4.49 and 12.26 mg galantamine equivalent/g extract against AChE and BChE, respectively) and α-amylase enzyme (1.04 mmol acarbose equivalent/g extract). On the other hand, the methanolic extract inhibited tyrosinase (121.42 mg kojic acid equivalent/g extract) and α-glucosidase (2.39 mmol acarbose equivalent/g extract) activities. The highest selectivity towards all cancer cell lines (SI 4.5–10.8) was observed with aqueous extract with the FaDu cells being the most sensitive (CC₅₀ 40.22 µg/mL). It can be concluded that the presence of certain bioactive antiviral molecules may be related to the high anti HSV-1 activity of the methanolic extract. This work has generated vital scientific data on this medicinal plant, which is a prospective candidate for the creation of innovative phyto-pharmaceuticals.     

Unravelling the Specificity of Laminaribiose Phosphorylase from Paenibacillus sp. YM-1 towards Donor Substrates Glucose/Mannose 1-Phosphate by Using X-ray Crystallography and Saturation Transfer Difference NMR Spectroscopy

Glycoside phosphorylases (GPs) carry out a reversible phosphorolysis of carbohydrates into oligosaccharide acceptors and the corresponding sugar 1-phosphates. The reversibility of the reaction enables the use of GPs as biocatalysts for carbohydrate synthesis. Glycosyl hydrolase family 94 (GH94), which only comprises GPs, is one of the most studied GP families that have been used as biocatalysts for carbohydrate synthesis, in academic research and in industrial production. Understanding the mechanism of GH94 enzymes is a crucial step towards enzyme engineering to improve and expand the applications of these enzymes in synthesis. In this work with a GH94 laminaribiose phosphorylase from Paenibacillus sp. YM-1 (PsLBP), we have demonstrated an enzymatic synthesis of disaccharide 1 (β-d -mannopyranosyl-(1→3)-d -glucopyranose) by using a natural acceptor glucose and noncognate donor substrate α-mannose 1-phosphate (Man1P). To investigate how the enzyme recognises different sugar 1-phosphates, the X-ray crystal structures of PsLBP in complex with Glc1P and Man1P have been solved, providing the first molecular detail of the recognition of a noncognate donor substrate by GPs, which revealed the importance of hydrogen bonding between the active site residues and hydroxy groups at C2, C4, and C6 of sugar 1-phosphates. Furthermore, we used saturation transfer difference NMR spectroscopy to support crystallographic studies on the sugar 1-phosphates, as well as to provide further insights into the PsLBP recognition of the acceptors and disaccharide products.     

Chemical deactivation of V2O5/WO3–TiO2 SCR catalysts by additives and impurities from fuels, lubrication oils and urea solution: Part II. Characterization study of the effect of alkali and alkaline earth metals

A characterization study on a practice-oriented V₂O₅/WO₃–TiO₂ SCR catalyst deactivated by Ca and K, respectively, was carried out using NH₃-TPD, DRIFT spectroscopy, and XPS as well as theoretical DFT calculations. It was found from NH₃-TPD experiments that strongly basic elements like K or Ca drastically affect the acidity of the catalysts. Detailed DRIFT spectroscopy experiments revealed that these poisoning agents mostly interact with the Brønsted acid sites of the V₂O₅ active phase, thus affecting the NH₃ adsorption. Moreover, these experiments also indicated that the V⁵⁺ = O sites are much less reactive on the poisoned catalysts. XPS investigations of the O 1s binding energies showed that the oxygen atoms of the V⁵⁺ = O sites are affected by the presence of the poisoning agents. Based on these results and on DFT calculations with model clusters of the vanadia surface, the poisoning mechanism is explained by the stabilization of the non atomic holes of the (0 1 0) V₂O₅ phase as a result of the deactivation element. Consequently, V–OH Brønsted acid sites and V⁵⁺ = O sites are inhibited, which are both of crucial importance in the SCR process. The deactivation model also gives an explanation to the very low concentrations of potassium needed to deactivate the SCR catalyst, since one metal atom sitting on such a non-atomic hole site deactivates up to four active vanadium centers.