Showing papers in "Journal of Basic Microbiology in 2009"
TL;DR: Cell wall‐degrading enzymes may contribute to pathogenesis by degrading wax, cuticle and cell walls, thus aiding tissue invasion and pathogen dissemination, and can act as elicitors of host defense reaction.
Abstract: Plant-pathogenic fungi produce an array of extracellular hydrolytic enzymes that enable them to penetrate and infect the host tissue; these enzymes are collectively called cell wall-degrading enzymes (CWDE). They may contribute to pathogenesis by degrading wax, cuticle and cell walls, thus aiding tissue invasion and pathogen dissemination. Furthermore, they can act as elicitors of host defense reaction.Fusarium head blight (FHB) is a disease caused principally by Fusarium graminearum on crops, occurring all over the world. Important economic losses on wheat-growing areas have been registered by altering quality parameters of grains. Significant progress has been made in understanding the infection process from F. graminearum on wheat, based on genomic technologies. The virulence degree of this phytopathogen on crops could arise from differences in the production of extracellular enzymes, factors controlling the establishment of infection.Fusarium graminearum isolates from different geographical areas have been examined, and a combination of morphological and molecular data allowed the division of fungi in diverse groups, which have been related to the variation in pathogenicity. In most studied cases there is a correlation between the presence of pectic enzymes, disease symptom and virulence, being also their production decisive in the infection process.
202 citations
TL;DR: From 16 samples of traditional fermented koumiss collected in Inner Mongolia Autonomous Region of China, forty‐eight lactobacilli strains were isolated and phenotypically characterized by their abilities to ferment different carbohydrates and by additional biochemical tests to select potentially probiotic strains.
Abstract: From 16 samples of traditional fermented koumiss collected in Inner Mongolia Autonomous Region of China, forty-eight lactobacilli strains were isolated and phenotypically characterized by their abilities to ferment different carbohydrates and by additional biochemical tests. The dominant lactobacilli species were identified as L. casei (17 strains), L. helveticus (10 strains) and L. plantarum (8 strains), with a lower frequency of isolation for L. coryniformis subsp. coryniformis (5 strains), L. paracasei (3 strains), L. kefiranofaciens (2 strains), L. curvatus (1 strain), L. fermentum (1 strain) and W. kandleri (1 strain). The pH values of all these samples were ranging from 3.37 to 3.94. In isolates, L. casei Zhang, L. helveticus ZL12-1, and L. plantarum BX6-6 were selected as potentially probiotic strains through the preliminary tests including resistance to low acid, abilities to grow in MRS with bile salts, antimicrobial activities and the viabilities during prolonged cold storage in fermented milk. Moreover 16S rDNA was conducted to confirm the identification.
117 citations
TL;DR: Five strains of microbes selected after testing their potential as plant growth promoters are suggested to enhance plant growth in nickel‐spiked land and remediate nickel from contaminated sites.
Abstract: Phytoremediation i.e. the use of plants to adsorb, accumulate or detoxify contaminants is an emerging area of interest. A viable technology needs optimum biomass production in metal contaminated soil. Five strains of microbes were selected after testing their potential as plant growth promoters, on the basis of their phosphate solubilization ability, IAA, siderophore and HCN production and biocontrol potentials. They were examined for growth in synthetic medium supplemented with nickel and their MIC (2 mM) was determined. These isolates were also able to grow and produce siderophores in presence of heavy metals like Ni, Zn and Cd. A positive response of bacterial inoculants was observed in chickpea plants towards toxic effect of nickel present in soil at different concentration (0, 1 and 2 mM). Bacterial inoculants enhanced fresh and dry weight of plants even at 2 mM nickel concentration. Pot experiments indicated that presence of nickel at upto 1 mM enhanced plant growth compared to uninoculated nickel free plants. The accumulation of nickel/plant was just 50% in Pseudomonas inoculated plants as compared to uninoculated plants with 2 mM nickel concentration along with increased biomass. The results suggest the use of these PGPR to enhance plant growth in nickel-spiked land and remediate nickel from contaminated sites. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
115 citations
TL;DR: Wild boars could be a reservoir of antimicrobial resistance genes in E. coli isolates, as evidenced by this study.
Abstract: ESBL-producing E. coli isolates have been isolated from eight of seventy seven faecal samples (10.4%) of wild boars in Portugal. The ESBL types identified by PCR and sequencing were bla(CTX-M-1) (6 isolates) and bla(CTX-M-1) + bla(TEM1-b) (2 isolates). Further resistance genes detected included tet (A) or tet (B) (in three tetracycline-resistant isolates), aad A (in three streptomycin-resistant isolates), cml A (in one chloramphenicol-resistant isolate), sul 1 and/or sul 2 and/or sul 3 (in all sulfonamide-resistant isolates). The intI 1 gene encoding class 1 integrase was detected in all ESBL-producing E. coli isolates. One isolate also carried the intI 2 gene, encoding class 2 integrase. The ESBL-producing E. coli isolates could be assigned to phylogenetic groups B1 (3 isolates), B2 (3 isolates) or A (2 isolates). Amino acid change in GyrA protein (Ser83Leu or Asp87Tyr) was detected in three nalidixic acid-resistant and ciprofloxacin-susceptible isolates. Two amino acid changes in GyrA (Ser83Leu + Asp87Asn) and one in ParC (Ser80Ile) were identified in two nalidixic acid- and ciprofloxacin-resistant isolates. As evidenced by this study wild boars could be a reservoir of antimicrobial resistance genes.
105 citations
TL;DR: This work provides quantitative information on Cr(VI) reduction in soil samples by an indigenous actinomycete and bioremediation ability of Streptomyces sp.
Abstract: This work provides quantitative information on Cr(VI) reduction in soil samples by an indigenous actinomycete. Streptomyces sp. MC1, previously isolated from sugarcane, has shown ability to reduce Cr(VI) in liquid minimal medium. A reduction of 100 and 75% was obtained at initial Cr(VI) concentrations of 5 and 50 mg l(-1), respectively, after 48 h of incubation. Bioremediation ability of Streptomyces sp. MC1 was assayed in soil extracts and soil samples. Relative growth of Streptomyces sp. MC1 was 77 and 38% when grown in soil extract with 10 and 50 mg l(-1) of Cr(VI), respectively. MC1 was able to reduce 30% of Cr(VI) after 96 h of incubation with 10 mg l(-1) of Cr(VI), and reduction coincided with the exponential growth phase at pH 7 and 30 degrees C.In soil samples, Streptomyces sp. MC1 was able to reduce up to 94% of the Cr(VI) bioavailability (50 mg kg(-1)) after 7 d. These results were compared with non-inoculated soil samples with Cr(VI). Bioremediation activity of Streptomyces sp. MC1 was not inhibited by natural soil microbial flora. Besides, Streptomyces sp. MC1 growth was not inhibited by 50 mg kg(-1) of Cr(VI). In contrast to findings obtained by other authors, our results showed almost complete Cr(VI) removal from soil without any previous treatment, and without addition of any substrate and with a normal soil humidity level. These results confirm the Cr(VI)-contaminated soil bioremediation potential of Streptomyces sp. MC1.
93 citations
TL;DR: This review focuses on the remodeling of the cell wall during transition from the vegetative cell to a heterocyst, including the formation of theheterocyst‐specific glycolipid layer and the heterocySt envelope polysaccharide layer.
Abstract: Filamentous cyanobacteria like Anabaena sp. PCC 7120 are able to develop a specialized cell type named heterocyst from vegetative cells in times of nitrogen starvation. Heterocyst development is controlled by the function of two master-regulators, NtcA and HetR. This review focuses on the remodeling of the cell wall during transition from the vegetative cell to a heterocyst, including the formation of the heterocyst-specific glycolipid layer and the heterocyst envelope polysaccharide layer. The functional assignment of genes involved therein, their genomic organization and their regulation are highlighted. Communication pathways and exchange routes for metabolites between heterocysts and vegetative cells are discussed. Further on, an overview of the heterocyst outer membrane proteome is given, together with possible functions of the identified proteins in the metabolism of heterocysts.
82 citations
TL;DR: This review focuses on the recent advances in identifying and characterizing the components of the CCM by transcriptome analyses of the Chlamydomonas cells that are transferred to CO2‐limiting stress conditions in light.
Abstract: Aquatic photosynthetic organisms can acclimate to the variable and limiting availability of CO(2) by operation of carbon-concentrating mechanism (CCM) that allows them to optimize carbon acquisition for photosynthesis. The CCMs of both eukaryotic alga and cyanobacteria function to facilitate CO(2) assimilation, when inorganic carbon (Ci; CO(2) and/or HCO(3)(-)) is limited. By active Ci uptake systems, internal Ci levels are increased and then carbonic anhydrase supplies sufficient CO(2) to ribulose 1,5-bisphosphate carboxylase/oxygenase by the dehydration of accumulated bicarbonate. Although the molecular components of CCM have been intensively studied in cyanobacteria, significant advances in understanding of the eukaryotic CCM have been achieved especially in a model green alga, Chlamydomonas reinhardtii. Recent accumulation of genomic sequence data of algae leads to start comparative genomic analyses of functional components of eukaryotic CCM. This review focuses on the recent advances in identifying and characterizing the components of the CCM by transcriptome analyses of the Chlamydomonas cells that are transferred to CO(2)-limiting stress conditions in light.
71 citations
TL;DR: A high sample throughput protocol exploiting first and second retention index for spectral library search and subsequent construction of a high dimensional data matrix useful for statistical analysis is presented.
Abstract: Two dimensional gas chromatography coupled to time-of-flight mass spectrometry (GCxGC-TOF-MS) is a promising technique to overcome limits of complex metabolome analysis using one dimensional GC-TOF-MS. Especially at the stage of data export and data mining, however, convenient procedures to cope with the complexity of GCxGC-TOF-MS data are still in development. Here, we present a high sample throughput protocol exploiting first and second retention index for spectral library search and subsequent construction of a high dimensional data matrix useful for statistical analysis. The method was applied to the analysis of 13C-labelling experiments in the unicellular green alga Chlamydomonas reinhardtii. We developed a rapid sampling and extraction procedure for Chlamydomonas reinhardtii laboratory strain (CC503), a cell wall deficient mutant. By testing all published quenching protocols we observed dramatic metabolite leakage rates for certain metabolites. To circumvent metabolite leakage, samples were directly quenched and analyzed without separation of the medium. The growth medium was adapted to this rapid sampling protocol to avoid interference with GCxGC-TOF-MS analysis. To analyse batches of samples a new software tool, MetMax, was implemented which extracts the isotopomer matrix from stable isotope labelling experiments together with the first and second retention index (RI1 and RI2). To exploit RI1 and RI2 for metabolite identification we used the Golm metabolome database (GMD [1] with RI1/RI2-reference spectra and new search algorithms. Using those techniques we analysed the dynamics of 13CO2 and 13C-acetate uptake in Chlamydomonas reinhardtii cells in two different steady states namely photoautotroph and mixotroph growth conditions. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
65 citations
TL;DR: 13 morphologically distinct strains of thermophilic bacteria isolated from a hot spring site in Garhwal region of Indian Himalaya have been characterized and identified using phenotypic and genotypic characters.
Abstract: 13 morphologically distinct strains of thermophilic bacteria isolated from a hot spring site in Garhwal region of Indian Himalaya have been characterized and identified using phenotypic and genotypic characters. All the strains developed circular to irregular colonies between 2-3 mm on Tryptone Yeast extract (TY) agar plates at 65 degrees C following 24-36 h incubation. In TY broth, facultative bacterial growth was observed within 12-16 h of incubation at 65 degrees C. The bacterial strains could tolerate a temperature range between 40-45 degrees C to 85-90 degrees C (optimum 65-70 degrees C) and pH between 4-11 (optimum 6-8). The cell morphology varied from short to long rods arranged in single, diplobacilli (in V or L shape) or short or long spiral chains with coiling. The bacterial strains varied in respect of their biochemical tests conducted for various enzymes, fermentation of sugars, tolerance to antibiotics and salt. Based on the 16S rRNA analysis, 11 strains showed maximum similarity with Geobacillus stereothermophilus, one strain with G. kaustophilus and one with Geobacillus sp.
56 citations
TL;DR: The results support the postulate that a combination of conventional biochemical and genotyping methods allows a thorough characterization and identification of isolates and propose that genotypic characterization could be complemented by biochemical characterization to discriminate L. plantarum strains.
Abstract: Ten Lactobacillus strains originally isolated from Thai fruits and vegetables fermentation were characterized by various phenotypic and genotypic methods. The phenotypic analysis using the method of carbohydrate fermentation patterns (API50CHL) revealed that the isolates belonged to the L. plantarum species. This was further confirmed by 16S rRNA gene sequencing. Multilocus sequence typing (MLST) revealed a strongly clonal population structure and a low genotypic diversity in this collection. However, the analyzed L. plantarum population demonstrated a higher level of diversification after API50CHL that reflects the role of available carbohydrate sources in bacterial evolution. Our results support the postulate that a combination of conventional biochemical and genotyping methods allows a thorough characterization and identification of isolates. We propose that genotypic characterization could be complemented by biochemical characterization to discriminate L. plantarum strains. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
52 citations
TL;DR: Consortial treatments effected very good growth promotion in Lycopersicon esculentum Mill and the treated plants also developed resistance against wilt pathogen, Fusarium oxysporum f.
Abstract: Biological systems are getting more relevance than chemical control of plant pathogens as they are not only eco-friendly and economic in approach but are also involved in improving the soil consistency and maintenance of natural soil flora. Plant growth promoting rhizosphere microorganisms were isolated from three different tree rhizospheres using selective culture media. Five microorganisms were selected from each rhizosphere soil based on their efficiency and screened for their ability to promote plant growth as a consortium. Each of the developed consortium has a phosphate solubilizer, nitrogen fixer, growth hormone producer, heterotrophic member and an antagonist. The plant growth promoting ability of the microbial members present in the consortium was observed by estimating the IAA production level and also by the nitrogenase activity of the nitrogen fixers. The biocontrol potentiality of the consortium and the antagonist present in the consortium were checked by both dual plate assay and cross-streaking technique. Consortial treatments effected very good growth promotion in Lycopersicon esculentum Mill and the treated plants also developed resistance against wilt pathogen, Fusarium oxysporum f. sp. lycopersici though the effect was well pronounced with consortium developed from Santalum album. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: The high acid‐resistance, bile resistance and antagonism against pathogens, suggest that the four lactic acid bacteria isolated from pig feces could prove useful as piglet probiotics.
Abstract: This study examined four lactobacilli isolated from pig feces. Two hundred lactic acid bacteria strains were obtained from pig feces using selective culture media (with vancomycin and bromocresol green; termed LAMVAB agar). Microscopy, the catalase test, Gram-staining, and RAPD-PCR analysis were used to group the bacteria into 20 related clusters. Phenotypic analysis using the API 50 CH test and genotypic analysis of 16S rDNA sequences identified these clusters as representing single strains of each of Lactobacillus fermentum, Lactobacillus salivarius, Lactobacillus plantarum, and Lactobacillus reuteri. Bacterial survival under the conditions of low pH (2.0) and high concentration (5.0%, w/v) of bile salt was much better than that of the reference strain (Lactobacillus acidophilus ATCC 33199). The isolated bacteria were quite capable of inhibiting the growth of two pathogens, Escherichia coli K88 and Salmonella typhimurium. The high acid-resistance, bile resistance and antagonism against pathogens, suggest that the four lactic acid bacteria isolated from pig feces could prove useful as piglet probiotics.
TL;DR: All of the respiratory inhibitors tested namely rotenone, azide, cyanide and antimycin A did not show any inhibition to the molybdenum‐reducing activity suggesting components of the electron transport system are not responsible for the reducing activity.
Abstract: Extensive use of metals in various industrial applications has caused substantial environmental pollution. Molybdenum-reducing bacteria isolated from soils can be used to remove molybdenum from contaminated environments. In this work we have isolated a local bacterium with the capability to reduce soluble molybdate to the insoluble molybdenum blue. We studied several factors that would optimize molybdate reduction. Electron donor sources such as glucose, sucrose, lactose, maltose and fructose (in decreasing efficiency) supported molybdate reduction after 24 h of incubation with optimum glucose concentration for molybdate reduction at 1.5% (w/v). The optimum pH, phosphate and molybdate concentrations, and temperature for molybdate reduction were pH 6.5, 5.0, 25 to 50 mM and 37 degrees C, respectively. The Mo-blue produced by cellular reduction exhibited a unique absorption spectrum with a maximum peak at 865 nm and a shoulder at 700 nm. Metal ions such as chromium, cadmium, copper, silver and mercury caused approximately 73, 71, 81, 77 and 78% inhibition of the molybdenum-reducing activity, respectively. All of the respiratory inhibitors tested namely rotenone, azide, cyanide and antimycin A did not show any inhibition to the molybdenum-reducing activity suggesting components of the electron transport system are not responsible for the reducing activity. The isolate was tentatively identified as Enterobacter sp. strain Dr.Y13 based on carbon utilization profiles using Biolog GN plates and partial 16S rDNA molecular phylogeny.
TL;DR: The phylogenetic diversity of a collection of 96 Bacilli, isolated from 17 distinct stations of 5 oceanographic campaigns, points out the need of more extensive studies to understand their distribution and ecology in deep‐sea environments.
Abstract: Members of the genus Bacillus and related genera are ubiquitous in nature. However, Bacillus species isolated from marine sediments have attracted less interest respect to their terrestrial relatives. Here, we report the phylogenetic diversity of a collection of 96 Bacilli, isolated from 17 distinct stations of 5 oceanographic campaigns. The diversity was analysed by phenotypic and molecular approaches based on the amplified rDNA restriction analysis (ARDRA), amplification of the internal transcribed spacers (ITS-PCR) and on 16S rRNA sequencing. Intra-specific polymorphism was efficiently detected by biochemical analysis and ARDRA while results of ITS-PCR were in agreement with 16S rRNA sequencing. The identification results assigned 68% of the isolates to the species B. subtilis, B. licheniformis, B. pumilus and B. cereus. Phylogenetic analysis allowed the separation of 9 isolates in a clade that may represent a group of obligate marine Bacillus since they clustered with B. firmus, B. foraminis and marine isolates with metal oxidation and bioaccumulation capabilities. The remaining isolates showed a close affiliation to the genera Virgibacillus, Gracilibacillus and Paenibacillus. The widespread of Bacilli and their high diversity level observed in this work point out the need of more extensive studies to understand their distribution and ecology in deep-sea environments.
TL;DR: The potential capacity of Streptomyces sp.
Abstract: 46 actinomycetes were isolated from two polluted sites and one unpolluted site. One strain, F4, was selected through primary qualitative screening assays because of its cadmium resistance, and physiologically and taxonomically characterized. F4 was able to grow at 7.5% NaCl and 100 microg/ml lysozyme and at a pH between 6 and 10. 16S rDNA sequence analysis showed that F4 was closely related to Streptomyces tendae. Growth of Streptomyces sp. F4 on culture medium with 8 mg/l Cd(2+) for 8 days showed 80% inhibition. Maximum specific biosorption was 41.7 mg Cd(2+)/g dry weight after 7 days of growth and highest Cd(2+ )concentration was found in the cell wall (41.2%). The exopolysaccharide layer only contained 7.4%, whereas 39.4% of Cd(2+) was found in the cytosolic fraction. Twelve % was found in the ribosomes and membrane fraction. This was verified with TEM, showing Streptomyces sp. F4 cytoplasm with dark granulate appearance. This study could present the potential capacity of Streptomyces sp. F4 for Cd(2+) bioremediation.
TL;DR: The ethyl acetate fractions of Nigrospora sphaerica and Phoma betae liquid fermentations contained the synergistic compounds 8‐hydroxy‐6‐methoxy‐3‐methylisocoumarin and (22E,24R)‐ergosta‐4,6,8(14),22‐tetraen‐3-one which are potential compounds for drug discovery.
Abstract: Smallanthus sonchifolius is a traditional Andean plant which has been cultured mainly in Brazil, Japan and New Zealand due to its medicinal properties. A study of the endophytic fungi associated to the plant was carried out in order to characterize new cytotoxic agents. Thirty two fungal strains were isolated and submitted to cultivation and extraction producing 186 extracts. Of these, 12% displayed moderate to high cytotoxic activities and were considered promising anticancer compound sources. The ethyl acetate fractions of Nigrospora sphaerica and Phoma betae liquid fermentations contained the synergistic compounds 8-hydroxy-6-methoxy-3-methylisocoumarin and (22E,24R)-ergosta-4,6,8(14),22-tetraen-3-one which are potential compounds for drug discovery. Another isolated compound, pimara-7,15-dien-3-beta-ol diterpene is being characterized for the first time through a detailed spectroscopic analysis including GC/MS, homo- and hetero-nuclear correlated NMR experiments (HMQC, HMBC, COSY and NOEdiff) along with its optical rotation.
TL;DR: Rhizobium radiobacter MTCC 8161 completely decolorized methyl violet within 8 h both at static and shaking conditions and significant increase in the activities of lignin peroxidase and aminopyrine N‐demethylase indicated involvement of these enzymes in the decolorization process.
Abstract: Rhizobium radiobacter MTCC 8161 completely decolorized methyl violet (10 mg l⁻¹) within 8 h both at static and shaking conditions. The decolorization time increased with increasing dye concentration. The effect of different carbon and nitrogen sources on the decolorization of methyl violet was studied. The maximum decolorization was observed in the presence of sucrose (1%) and urea (1%). UV-Visible, HPLC and FTIR analysis of extracted products confirmed biodegradation of methyl violet. The significant increase in the activities of lignin peroxidase and aminopyrine N-demethylase in the cells obtained after decolorization indicated involvement of these enzymes in the decolorization process. In addition to methyl violet, this strain also shows an ability to decolorize various industrial dyes, (red HE7B, yellow 4G, blue 2B, navy blue HE22, red M5B and red HE3B). (© 2009 WILEY-VCH Verlag GmbH '' Co. KGaA, Weinheim)
TL;DR: Endophytes are microorganisms that colonize plant tissues internally without causing harm to the host, and yeast could assist in the establishment of this pathogen in its host plant and, therefore, contribute to the development of disease symptoms.
Abstract: Endophytes are microorganisms that colonize plant tissues internally without causing harm to the host. Despite the increasing number of studies on sweet orange pathogens and endophytes, yeast has not been described as a sweet orange endophyte. In the present study, endophytic yeasts were isolated from sweet orange plants and identified by sequencing of internal transcribed spacer (ITS) rRNA. Plants sampled from four different sites in the state of Sao Paulo, Brazil exhibited different levels of CVC (citrus variegated chlorosis) development. Three citrus endophytic yeasts (CEYs), chosen as representative examples of the isolates observed, were identified as Rhodotorula mucilaginosa, Pichia guilliermondii and Cryptococcus flavescens. These strains were inoculated into axenic Citrus sinensis seedlings. After 45 days, endophytes were re-isolated in populations ranging from 106 to 109 CFU/g of plant tissue, but, in spite of the high concentrations of yeast cells, no disease symptoms were observed. Colonized plant material was examined by scanning electron microscopy (SEM), and yeast cells were found mainly in the stomata and xylem of plants, reinforcing their endophytic nature. P. guilliermondii was isolated primarily from plants colonized by the causal agent of CVC, Xylella fastidiosa. The supernatant from a culture of P. guilliermondii increased the in vitro growth of X. fastidiosa, suggesting that the yeast could assist in the establishment of this pathogen in its host plant and, therefore, contribute to the development of disease symptoms. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: Pycnoporus sanguineus is a white‐rot fungus that produces ligninolytic enzymes such as laccases that are useful for pulp bleaching, dye decolorization and phenolic degradation.
Abstract: Pycnoporus sanguineus is a white-rot fungus that produces ligninolytic enzymes such as laccases. These enzymes can endure temperatures as high as 60 degrees C and are useful for pulp bleaching, dye decolorization and phenolic degradation.Laccase production by fungi depends not only on the carbon and nitrogen sources but also on the nitrogen concentration of the culture medium. In this work, we examined the effect of four carbon sources (maltose, glucose, fructose and sucrose) and four nitrogen sources (ammonium tartrate, sodium nitrate, asparagine and yeast extract) on the activity of laccase from Pycnoporus sanguineus. All carbon and nitrogen sources exhibited a strong influence on laccase activity, a sucrose-asparagine medium providing the best results (320 mU/ml). Moreover, using an asparagine concentration 5 times higher than the reference level increased laccase activity to 820 mU/ml. Higher asparagine concentrations, however, resulted in no further increase in activity.Consistent with previous results, the carbon and nitrogen sources, and the nitrogen concentration, had a strong impact on laccase activity, the optimum conditions depending on the particular fungus. The conditions of the culture medium had a marked effect on laccase activity, which increased up to 820 mU/ml.
TL;DR: The partially purified enzyme was found to be glyco‐metalloproteinic in nature as concluded from the analytical and spectroscopic profiles of the enzyme preparation.
Abstract: Solid-state fermentation was carried out for the production of extra-cellular L-methioninase by Aspergillus flavipes (Bain and Sart.) using nine agro-industrial residues, namely wheat bran, rice bran, wheat flour, coconut seeds, cotton seeds, ground nut cake, lentil hulls, soya beans and chicken feathers. Chicken feathers were selected as solid substrate for L-methioninase production by A. flavipes. The maximum L-methioninase productivity (71.0 U/mg protein) and growth (11 mg protein/ml) of A. flavipes was obtained using alkali pretreated chicken feathers of 50% initial moisture content as substrate supplemented with D-glucose (1.0% w/v) and L-methionine (0.2% w/v). External supplementation of the fermentation medium with various vitamin sources has no overinductive effect on L-methioninase biosynthesis. The partially purified A. flavipes L-methioninase preparation showed highest activity (181 U/ml) at pH 8.0 with stability over a pH range (pH 6-8) for 2 h. L-methioninase activity was increased by preincubation of the enzyme for 2 h with Co(2+), Mn(2+), Cu(2+) and Mg(2+) and strongly inhibited by the presence of EDTA, NaN(3), Li(2+), Cd(2+), DMSO and 2-mercaptoethanol. The enzyme preparation has a broad substrate spectrum showing a higher affinity to deaminate L-glycine, N -acetylglucosamine and glutamic acid, in addition to their proteolytic activity against bovine serum albumin, casein, gelatin and keratin. The partially purified enzyme was found to be glyco-metalloproteinic in nature as concluded from the analytical and spectroscopic profiles of the enzyme preparation. The demethiolating activity of the enzyme was also visualized chromogenially.
TL;DR: Annotation of genes encoding isozymes of the reductive and oxidative pentose phosphate pathways in the genomes of the centric diatom Thalassiosira pseudonana and the pennate diatom Phaeodactylum tricornutum and bioinformatically inferred their intracellular distribution indicated that this compartment might be involved in metabolic processes in diatoms.
Abstract: Diatoms contribute a large proportion to the worldwide primary production and are particularly effective in fixing carbon dioxide. Possibly because diatom plastids originate from a secondary endocytobiosis, their cellular structure is more complex and metabolic pathways are rearranged within diatom cells compared to cells containing primary plastids. We annotated genes encoding isozymes of the reductive and oxidative pentose phosphate pathways in the genomes of the centric diatom Thalassiosira pseudonana and the pennate diatom Phaeodactylum tricornutum and bioinformatically inferred their intracellular distribution. Prediction results were confirmed by fusion of selected presequences to Green Fluorescent Protein and expression of these constructs in P. tricornutum. Calvin cycle enzymes for the carbon fixation and reduction of 3-phosphoglycerate are present in single isoforms, while we found multiple isoenzymes involved in the regeneration of ribulose-1,5-bisphosphate. We only identified one cytosolic sedoheptulose-1,7-bisphosphatase in both investigated diatoms. The oxidative pentose phosphate pathway seems to be restricted to the cytosol in diatoms, since we did not find stromal glucose-6-phosphate dehydrogenase and 6-phosphogluconolactone dehydrogenase isoforms. However, the two species apparently possess a plastidic phosphogluconolactonase. A 6-phosphogluconolactone dehydrogenase is apparently plastid associated in P. tricornutum and might be active in the periplastidic compartment, suggesting that this compartment might be involved in metabolic processes in diatoms.
TL;DR: In vitro analysis of extracellular enzymes like protease, amylase, caseinase, chitinase and lipase was undertaken in an attempt to understand their relevance to virulence of the isolates, finding a 50% relationship associated with original insect host, pathogenicity and enzyme production.
Abstract: Extracellular enzymes produced by Beauveria bassiana, are believed to play a key role in cuticle hydrolysis. Enzyme production and pathogenicity has been found to be positively correlated. Twenty-eight isolates of B. bassiana, collected from different geographical regions and host ranges were characterized by in vitro extracellular enzyme production and SDS–PAGE techniques for discerning biochemical basis for virulence among the different isolates. In vitro analysis of extracellular enzymes like protease, amylase, caseinase, chitinase and lipase was undertaken in an attempt to understand their relevance to virulence of the isolates. The different isolates of B. bassiana were evaluated for virulence to the second instar larvae of Helicoverpa armigera in laboratory bioassays. SDS–PAGE of total intracellular soluble proteins was also studied in order to understand affinities among the different isolates of B. bassiana. The relationship between enzyme production and pathogenicity and vice versa was nearly 50%. There was a 50% relationship associated with original insect host, pathogenicity and enzyme production. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: The yeast strain CLOA 72 isolated from the effluent of a dairy industry in Brazil and identified as Trichosporon montevideense, was able to grow and produce a glycolipid biosurfactant when cultured on a mineral medium with sunflower oil as the carbon source.
Abstract: The yeast strain CLOA 72 isolated from the effluent of a dairy industry in Brazil and identified as Trichosporon montevideense, was able to grow and produce a glycolipid biosurfactant when cultured on a mineral medium (MM) with sunflower oil as the carbon source. Biosurfactant production was partially growth-associated and maximal emulsification activity was observed at 144 h of cultivation (78.92%). The biosurfactant purified by precipitation with ethanol showed 78.66% emulsifying activity when used in concentrations above 4.5 mg/ml and was able to reduce the surface tension of water to values below 44.9 mN/m. The critical micellar concentration (CMC) was found to be 2.2 mg/ml. The highest emulsifying activity (E24) has been observed with vegetable oils, toluene, kerosene, isooctane, cyclohexane, hexane, diesel oil and hexadecane as compared to mineral oil and oleic acid. The biosurfactant also showed good stability during exposure to 100 °C for different periods of time (10 to 60 min), to high salinity (30% of NaCl, KCl and NaHCO3), and to a wide range of pH values (1–10). The biosurfactant purified by gel filtration chromatography is a glycolipid, with lipid portion containing 16.03% (9Z)-octadec-9-enoic acid, 14.92% hexadecanoic acid, and 9.63% (E) octadec-9-enoic acid and the carbohydrate portion containing mannose (35.29%), xylose (41.99%), arabinose (17.47%), and glucose (5.25%). (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: This survey showed that faecal bacteria such as E. coli and enterococci of healthy growing children's could be a reservoir of antimicrobial resistance genes.
Abstract: From stool specimens of 118 healthy children's (1–14 years) in Portugal 92 E. coli and 101 Enterococcu s spp. strains have been isolated. Almost half (40.2%) of the E. coli isolates were resistant to ampicillin, 25.0% were resistant to tetracycline and 26.1% were resistant to streptomycin. Resistance genes detected by specific PCR included blaTEM and/or blaSHV and/or blaCTX-M (33 of 37 ampicillin and/or cefotaxime resistant isolates), tet (A) and/or tet (B) (16 of 23 tetracycline-resistant isolates), aad A (19 of 24 streptomycin-resistant isolates), cml A (in the two chloramphenicol-resistant isolates), aac (3)-II with/without aac (3)-IV (in the four gentamicin-resistant isolates), sul 1 and/or sul 2 and/or sul 3 (in all trimethoprim/sulfamethoxazole resistant isolates). The majority of the resistant E. coli isolates (69.1%) belonged to phylogenetic group B2. Of the enterococci isolates E. faecium (n = 53), E. faecalis (n = 41), E. hirae (n = 4) and E. durans (n = 3) more than one-fourth (28.7%) of the isolates were resistant to tetracycline; 21.8% were resistant to erythromycin and 8.9% were resistant to kanamycin. Resistance genes detected by PCR in enterococci included aph (3)′-IIIa (in all kanamycin-resistant isolates), aac (6′) (in all gentamicin-resistant isolates), tet (M) and/or tet (L) (26 of 29 tetracycline-resistant isolates), erm (B) (17 of 22 erythromycin-resistant isolates). This survey showed that faecal bacteria such as E. coli and enterococci of healthy growing children's could be a reservoir of antimicrobial resistance genes. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: Doubts are raised about the utility of indicator VOC for the detection of hidden mold growth in indoor environments because most of the microbial‐produced VOC concentrations were below the analytical detection limit for conventional indoor air analysis.
Abstract: Microorganisms such as bacteria and molds produce an enormous variety of volatile metabolites. To determine whether typical microbial volatile metabolites can be used as indicator compounds for the detection of hidden mold in indoor environments, we examined 14 typical indoor fungal strains for their growth rates and their capability to produce volatile organic compounds (VOC) on standard clinical media and on agar medium made from building materials. Air samples from Headspace Chambers (HSC) were adsorbed daily on Tenax TA tubes and analyzed by thermal desorption gas chromatography and mass spectrometry. In parallel, metabolic activity was measured by determining oxygen demand, the microbial biomass was assessed by dry weighing. Profiling of the volatile metabolites showed that VOC production depended greatly on fungal strain, culture medium, biological activity, and time. The laboratory-derived maximum emission rates were extrapolated to approximate indoor air concentrations in a hypothetical mold-infested room. The extrapolated indoor air data suggest that most of the microbial-produced VOC concentrations were below the analytical detection limit for conventional indoor air analysis. Additionally, conducted indoor air analysis in mold homes confirmed these findings for the most part. The present findings raise doubts about the utility of indicator VOC for the detection of hidden mold growth in indoor environments. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: First evidence for putative regulatory nucleotide sequences which will be useful in future studies on nickel uptake and incorporation are presented, in order to address regulation by nickel availability and incorporation into the mature protein.
Abstract: Superoxide dismutases are essential enzymes involved in detoxification of reactive oxygen by dismutation of the superoxide radical anion. A class of nickel containing superoxide dismutases has been described for streptomycetes and cyanobacteria. In silico analysis was used to study the distribution of genes coding for NiSOD in other taxa and to elucidate signals linked to nickel incorporation and maturation of NiSOD. Data mining revealed homologous proteins from actinobacteria, proteobacteria, chlamydiae, and eukarya (green algae) thus allowing a comparison of protein structural elements. Nickel ligands and maturation signals for N-terminal proteolysis were highly conserved. Genomic sequences surrounding genes encoding NiSOD homologs were compared in order to detect putative accessory enzymes involved in maturation. An endopeptidase gene linked to sodN coding for NiSOD was found in actinobacteria and cyanobacteria, but not in other taxa. The distribution of NiSOD encoding sequences showed four clusters which are not consistent with the phylogeny of the species. In addition, the different genomic context argues for heterologous gene transfer, most likely from actinobacteria to other taxa. In order to address regulation by nickel availability and incorporation into the mature protein, we present first evidence for putative regulatory nucleotide sequences which will be useful in future studies on nickel uptake and incorporation. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: Many genes encoding cytokines, such as IL2, IL‐1B, CXCL2, CxCL5, CCL20, CD14 and TNFRSF13B, were up‐regulated during the infection, confirming the report that they are important mediators to activate host responses to invading pathogens.
Abstract: Despite the importance of pneumonic plague caused by Yersinia pestis, a few is known about the interaction between Y. pestis and its host at the molecular level during the pneumonic plague development. In this study, we employed an intranasally challenged plague model in mice for investigating the kinetics of the disease progression by transcriptional profiling of Y. pestis and mice using qRT-PCR and microarray, respectively. The increasing transcription of important virulence genes of Y. pestis and of mice genes involving in immune and inflammatory defensive responses, and responses to stimuli, presents an overview of interaction between Y. pestis and mice during development of pneumonic plague. The early and persisting up-regulation of caf 1, psa A and lcr V in vivo indicated their role in resisting the host innate immune responses. The up-regulation of fur, ybt A and hms H in vivo reflected the ability of Y. pestis for acquiring iron. The transcription regulators, including pho P, oxy R and omp R, were up-regulated during plague development, suggesting their roles in interaction between Y. pestis and mice. Many genes encoding cytokines, such as IL2, IL-1B, CXCL2, CXCL5, CCL20, CD14 and TNFRSF13B, were up-regulated during the infection, confirming the report that they are important mediators to activate host responses to invading pathogens. The up-regulation of some genes encoding important virulent factors of Y. pestis and expression alterations of some genes encoding cytokines in the host reflect the interaction between the pathogen and the host, which will help us better understand plague pathogenesis. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
TL;DR: According to the results, the exopolymers studied play an important role not only on structure and biofilm biomass but also on cellular activity, and it is suggested that extracellular protein production it is not a determinative factor for biofilm total biomass.
Abstract: Staphylococcus epidermidis is now well established as a major nosocomial pathogen, associated with indwelling medical devices. Its major virulence factor is related with the ability to adhere to indwelling medical devices and form biofilms. In this study, the biofilm matrix of four S. epidermidis clinical isolates was extracted and the polysaccharides and proteins content was quantified. The results were correlated with the total biofilm biomass (determined by crystal violet assay) and cellular metabolic activity (evaluated with XTT reduction assay). According to the results, the exopolymers studied play an important role not only on structure and biofilm biomass but also on cellular activity. Thus, the strain forming biofilms with the highest level of polysaccharides (S. epidermidis 1457) also formed thicker biofilms but with the lowest metabolic activity. The protein concentration also varied among strains, with the biofilm matrix of S. epidermidis 9142 presenting a higher concentration of proteins comparing to the remaining strains. This fact indicates the different levels of importance that matrix proteins can hold on biofilm composition among strains albeit overall, it is suggested that extracellular protein production it is not a determinative factor for biofilm total biomass.
TL;DR: Sixteen Bacillus thuringiensis strains screened for their anti‐insect, antibacterial and antifungal determinants by phenotypic tests and PCR targeting major insecticidal proteins and complements could be very promising in field application.
Abstract: Sixteen Bacillus thuringiensis (Bt) strains were screened for their anti-insect, antibacterial and antifungal determinants by phenotypic tests and PCR targeting major insecticidal proteins and complements, chitinases, lactonases, beta-1,3-glucanases and zwittermicinA. Six strains had genes of at least two major insecticidal toxins and of insecticidal complements. With regard to fungal biocontrol, all the strains inhibited Fusarium oxysporum and Aspergillus flavus growth and four strains had all or most of the antifungal determinants examined, with strain Bt HD932 showing the widest antifungal activity spectrum. Autolysins, bacteriocin and AHL-lactonases were produced by all or most of the tested strains with different activity spectra including pathogens like Listeria monocytogenes. Safety evaluation was carried out via PCR by screening the B. cereus psychrotolerance-related genes, toxin genes and the virulence pleiotropic regulator plcR. Diarrheal enterotoxins and other toxin genes were widespread among the collection with strains Bt HD9 and H45 lacking psychrotolerance-related genes, while five strains were positive. Only three strains (BMG1.7, H172, H156) resulted positive with primer sets targeting partial or complete plcR gene. By Vero Cell Assays, Bt HD868 followed by Bt HD9 were shown to be the safest strains. These polyvalent and safe Bt strains could be very promising in field application.
TL;DR: This study evaluated the contamination rate of VTEC in slaughterhouses and butchers' shops in southern Jordan and determined that the isolated VTEC was highly sensitive to gentamicin and co‐trimoxazole and highly resistant to tetracycline and ampicillin.
Abstract: This study was conducted to determine the prevalence rate of VTEC in slaughtered sheep and goats and to evaluate the contamination rate of VTEC in slaughterhouses and butchers' shops in southern Jordan. 201 E. coli isolates from animals' faecal samples and 33 E. coli isolates from slaughterhouse/butcher shop samples were characterized by multiplex PCR (mPCR) reaction for detection of stx1, stx2, eae A and E-hly A virulent genes. Twenty-six virulent E. coli isolates were characterized by mPCR to seven different virulent patterns: stx1, stx1+stx2, stx1+eae A, stx1+E-hly A, stx1+eae A+E-hly A, eae A and E-hly A. It was found that VTEC comprised 6.4% and 21% of the total E. coli isolates from slaughtered small ruminants and slaughterhouses/ butchers' shops, respectively. The VTEC comprised 76.2% of the virulent isolates. The proportion of stx1:stx1+stx2 patterns was 19:1. It was found that the characterized complex VTEC (containing eae A and/or E-hly A) possessed three virulence patterns, including (VTEC) stx1 +eae A, (VTEC/EHEC) stx1 +E-hly A and (VTEC/EHEC) stx1 +eae A +E-hly A in percentages of 30%, 25% and 10%, respectively, in relation to the total VTEC isolates. Only two VTEC isolates were characterized as E. coli O157 and O26 serotypes, as highly pathogenic strains. Each of the O157 and O26 VTEC isolates was in a percentage of 0.4% in relation to the total E. coli isolates with virulent patterns stx1, eae A and E-hly A. The rest of the VTEC isolates were non-O157 VTEC. The antibiotic sensitivity test showed that the isolated VTEC was highly sensitive to gentamicin and co-trimoxazole and highly resistant to tetracycline and ampicillin. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)