Brazilian Journal of Microbiology, Volume: 40, Issue: 4, Published: 2009
  • Production of Poly-γ-Glutamate (PGA) Biopolymer by Batch and Semicontinuous Cultures of Immobilized Bacilluslicheniformis strain-R Industrial Microbiology

    Berekaa, Mahmoud M.; El Aassar, Samy A.; El-Sayed, Samia M.; EL Borai, Aliaa M.

    Abstract in English:

    Production of Polyglutamate (PGA) biopolymer by immobilized Bacilluslicheniformis strain-R was intensively investigated. Preliminary experiments were carried out to address the most suitable immobilization methodology. Entrapment of Bacillus cells in alginate-agar led optimal PGA production (36.75 g/l), with 1.32- and 2.18-fold increase in comparison with alginate- or K-carrageenan-immobilized cells, respectively. During semicontinuous cultivation of agar-alginate gel-cell mixture, production of PGA by 10 ml mixture was increased from 2nd to 3rd run whereas, increased till the 4th run using 15ml mixture. Adsorption was the most suitable immobilization technique for production of PGA and the sponge cubes was the preferred matrix recording 43.2 g/l of PGA with the highest cell adsorption. Furthermore, no PGA was detected when B. licheniformis cells were adsorbed on wood and pumice. Although luffa pulp-adsorbed cells recorded the highest PGA production (50.4 g/l), cell adsorption was the lowest. Semicontinuous cultivation of B. licheniformis cells adsorbed on sponge led to increase of PGA production till the 3rd run and reached 55.5 g/l then slightly decreased in the 4th run. The successful use of fixed-bed bioreactor for semicontinuous cultivation of B. licheniformis cells held on sponge cubes (3 runs, 96 hours/run) provides insight for the potential biotechnological production of PGA by immobilized cells.
  • Chitinase Production by Streptomyces sp. ANU 6277 Industrial Microbiology

    Narayana, Kolla J.P.; Vijayalakshmi, Muvva

    Abstract in English:

    Chitinase production by a terrestrial Streptomyces sp. ANU 6277 was studied under sub-merged fermentation. Chitinase production started after 24 h of incubation and reached maximum levels after 60 h of cultivation. A high level of chitinase activity was observed in the culture medium with pH 6 at 35ºC. Culture medium amended with 1% chitin was found to be suitable for maximum production of chitinase. An optimum concentration of colloidal chitin for chitinase production was determined. Studies on the influence of additional carbon and nitrogen sources on chitinase production revealed that starch and yeast extract served as good carbon and nitrogen sources to enhance chitinase yield.Chitinase was purified from crude enzyme extract by single step gel filtration by Sephadex G-100. Purified chitinase of the strain exhibited a distinct protein band near 45 kDa by means of SDS-PAGE.
  • Effect of branched-chain amino acids, valine, isoleucine and leucine on the biosythesis of bitespiramycin 4"-O-acylspiramycins Industrial Microbiology

    Li, Zhen-Lin; Wang, Yong-Hong; Chu, Ju; Zhuang, Ying-Ping; Zhang, Si-Liang

    Abstract in English:

    Bitespiramycin, a group of 4"-O-acylated spiramycins with 4"-O-isovalerylspiramycins as the major components, was produced by recombinantspiramycin-producing strain Streptomyces spiramyceticus harboring a 4"-O-acyltransferase gene. The experiment was initially performed in synthetic medium with 0.5 g l-1 Valine, Isoleucine or Leucine feeding at 36 h cultivation. When valine was fed, the biological titer of bitespiramycin was 45.3% higher than that of the control group, but the relative content of total isovalerylspiramycin components decreased by 22.5%. In the case of ilecine, the biological titer of bitespiramycin and the total isovalerylspiramycins alone were 85% and 72.1% of the control group, respectively. In contrast, the relative content of other acylated spiramycins increased by 54.41%. However, leucine feeding increased the relative content of total isovalerylspiramycins by 41.9% while the biological titer of bitespiramycin was nearly equal to that of the control group. The improvement effect of leucine on the biosynthesis of isovalerylspiramycins was further confirmed by feeding of 2.0 g l-1 leucine to the culture with complex medium. After batch feeding with a total amount of 2.0 g l-1 leucine to the culture from 70 h to 90 h, the biological titer of bitespiramycin was almost unreduced, and the final relative content of total isovalerylspiramycins increased from 31.1% to 46.9%.
  • Screening of microbial lipases and evalutaion of their potential to produce glycerides with high gamma linolenic acid concentration Industrial Microbiology

    Fregolente, Patricia B.L.; Fregolente, Leonardo V.; Maciel, Maria R.W.; Carvalho, Patricia O.

    Abstract in English:

    Gamma-linolenic acid (GLA, 18:3, cis- 6,9,12-octadecatrienoic acid), an important compound in n-6 eicosanoid family biosynthesis, occurs in the lipids of a few plant and microbial sources. This study focused on the screening of microbial strains with suitable lipase activity for enrichment of GLA by selective hydrolysis of the borage oil (21.6 % of GLA/total fatty acids). Firstly, 352 microrganisms were tested for their lipolytic capacity using screening techniques on agar plates containing borage oil, strains were then selected and screened for their activity (U/mg) using both submerged fermentation (SmF) and solid state fermentation (SSF). The rate of hydrolysis and the selective preference of these hydrolytic enzymes towards fatty acids, with a special focus on enrichment of GLA were studied and compared with those obtained by two commercially-available lipases. Only one of the lipases tested during this study displayed selectivity, discriminating the GLA during the hydrolysis reaction. Using the enzymatic extract from Geotrichum candidum as a biocatalyst of the reaction, it was possible to obtain a percentage of 41.7% of GLA in acylglycerols fraction when the borage oil was treated in a fixed-bed reactor for 24 hours at 30ºC.
  • Antibacterial and Antioxidant Activities of Acid and Bile Resistant Strains of Lactobacillus fermentum Isolated from Miang Industrial Microbiology

    Klayraung, Srikanjana; Okonogi, Siriporn

    Abstract in English:

    Miang is a kind of traditional fermented tea leaves, widely consumed in northern Thailand as a snack. It contains several kinds of Lactobacilli spp. The aim of this study was to isolate strains of Lactobacillus fermentum from miang and to investigate their antibacterial and antioxidant activities. The agar spot and well assays were used for determination of antibacterial power. The antibacterial mechanism was investigated by cell morphologic change under scanning electron microscope (SEM). Antioxidant activity was studied by means of free radical scavenging and ferric reducing power assays. The acid and bile screening tests indicated that L. fermentum FTL2311 and L. fermentum FTL10BR presented antibacterial activity against several pathogenic bacteria: Listeria monocytogenes DMST 17303, Salmonella Typhi DMST 5784, Shigella sonnei DMST 561 (ATCC 11060)and Staphylococcus aureus subsp. aureus DMST 6512 (ATCC 6538Ptm). The results from SEM suggested that the antibacterial action was due to the destruction of cell membrane which consequently caused the pathogenic cell shrinking or cracking. The antioxidant study suggested that both L. fermentum FTL2311 and L. fermentum FTL10BR strains could liberate certain substances that possessed antioxidant activity expressed as trolox equivalent antioxidant capacity (TEAC) and equivalent concentration (EC) values for free radical scavenging and reducing mechanisms, respectively. The supernatant of L. fermentum FTL2311 broth revealed TEAC and EC values of 22.54±0.12 and 20.63±0.17 µM.mg-1 respectively, whereas that of L. fermentum FTL10BR yielded TEAC and EC values of 24.09±0.12 and 21.26±0.17 µM.mg-1 respectively. These two strains isolated from miang present high potential as promising health-promoting probiotics.
  • Effect of lysozyme or nisin on survival of some bacteria treated with high pressure at subzero temperature Industrial Microbiology

    Malinowska-Pańczyk, Edyta; Kołodziejska, Ilona

    Abstract in English:

    The aim of this work was to examine the inactivation of some Gram-positive and Gram-negative bacteria exposed to the pressure of 193 MPa at -20 °C in the presence of lysozyme or nisin at concentration of 400 mg/ml. The highest effect of pressure at subzero temperature and lysozyme was found with pressure sensitive Pseudomonas fluorescens; viable cells of this strain were not detected in 1 ml of sample after combined treatment. The action of pressure at subzero temperature and lysozyme or nisin against Escherichia coli led to synergistic reduction by 0.7 or 1.6 log cycles, respectively, while it was practically insignificant for two Staphylococcus aureus strains. Viability loss of E. coli and S. aureus occurred during storage for 20 h of the samples at 37 and 5 °C, which were previously pressurized with lysozyme or nisin. The synergistic effect of pressure and nisin at pH 5 against E. coli cells just after the pressure treatment was lower than that at pH 7, however, the extent of the lethal effect after storage was higher.
  • New vectors derives from pUC 18 for clonig and thermal-induced expression in Escherichia coli Industrial Microbiology

    Xavier, Mauro Aparecido Souza; Kipnis, André; Torres, Fernando Araripe Gonçalves; Astofi-Filho, Spartaco

    Abstract in English:

    We report the construction of two vectors for Escherichia coli: pUC72, for molecular cloning, and pPLT7, for thermal-induced expression. The main feature of pUC72 is a novel polylinker region that includes restriction sites for Nde I and Nco I which provide an ATG codon for proper translation initiation of expressed genes. Vector pPLT7 is ideal for thermo-inducible expression in host cells that carry the cI857 repressor gene. The use of pPLT7 was validated by the successful expression of the genes encoding carp and porcine growth hormones. These vectors provide novel cloning possibilities in addition to simple, non-expensive, high level expression of recombinant proteins in E. coli.
  • Characterization And Application Of Tannase Produced By Aspergillus Niger ITCC 6514.07 On Pomegranate Rind Industrial Microbiology

    Srivastava, Anita; Kar, Rita

    Abstract in English:

    Extracellular tannase and gallic acid were produced optimally under submerged fermentation at 37 0C, 72 h, pH 5.0, 10 %(v/v) inoculum and 4 %(w/v) of the agroresidue pomegranate rind (PR) powder by an Aspergillus niger isolate. Tannic acid (1 %) stimulated the enzyme production by 245.9 % while with 0.5 % glucose, increase was marginal. Tannase production was inhibited by gallic acid and nitrogen sources such as NH4NO3, NH4Cl, KNO3, asparatic acid, urea and EDTA. The partially purified enzyme showed temperature and pH optima of 35 0C and 6.2 respectively which shifted to 40 0C and 5.8 on immobilization in alginate beads. Activity of the enzyme was inhibited by Zn+2, Ca+, Mn+2, Mg+2, Ba+2and Ag+. The immobilized enzyme removed 68.8 % tannin from juice of aonla/myrobalan (Phyllanthus emblica), a tropical fruit, rich in vitamin C and other essential nutrients. The enzymatic treatment of the juice with minimum reduction in vitamin C is encouraging as non enzymatic treatments of myrobalan juice results in vitamin C removal.
  • Production of laccase by Pynoporus sanguineus using 2,5 - Xylidine and ethanol Industrial Microbiology

    Valeriano, Viviane S.; Silva, Anna Maria F.; Santiago, Mariângela F.; Bara, Maria T. F.; Garcia, Telma A.

    Abstract in English:

    Enzyme application in biotechnological and environmental processes has had increasing interest due to its efficiency, selectivity and mainly for being environmentally healthful, but these applications require a great volume of enzymes. In this work the effect of different concentrations of ethanol and 2,5 - xylidine on growth and production of laccase by Pycnoporus sanguineus was investigated. In a medium containing 200 mg.L-1 of 2,5 - xylidine or 50 g.L-1 of ethanol, the maximum activity of laccase was 2019 U.L-1 and 1035 U.L-1, respectively. No direct correlation between biomass and activity of laccase was observed for any of the inducers used during the tests. Ethanol concentrations, larger than or equal to 20 g.L-1, inhibited the radial growth of P. sanguineus. This study showed that ethanol, which has less toxicity and cost than the majority of the studied inducers, presents promising perspectives for laccase production by P. sanguineus.
  • Purification and characterisation of an extracellular phytase from Aspergillus niger 11T53A9 Industrial Microbiology

    Greiner, Ralf; Silva, Lucineia Gomes da; Couri, Sonia

    Abstract in English:

    An extracellular phytase from Aspergillus niger 11T53A9 was purified about 51-fold to apparent homogeneity with a recovery of 20.3% referred to the phytase activity in the crude extract. Purification was achieved by ammonium sulphate precipitation, ion chromataography and gel filtration. The purified enzyme behaved as a monomeric protein with a molecular mass of about 85 kDa and exhibited maximal phytate-degrading activity at pH 5.0. Optimum temperature for the degradation of phytate was 55°C. The kinetic parameters for the hydrolysis of sodium phytate were determined to be K M = 54 µmol l-1 and k cat = 190 sec-1 at pH 5.0 and 37°C. The purified enzyme was rather specific for phytate dephosphorylation. It was shown that the phytase preferably dephosphorylates myo-inositol hexakisphosphate in a stereospecific way by sequential removal of phosphate groups via D-Ins(1,2,4,5,6)P5, D-Ins(1,2,5,6)P4, D-Ins(1,2,6)P3, D-Ins(1,2)P2 to finally Ins(2)P.
  • Decolorization of different textile dyes by Penicillium simplicissimum and toxicity evaluation after fungal treatment Industrial Microbiology

    Bergsten-Torralba, L.R.; Nishikawa, M.M.; Baptista, D.F.; Magalhães, D.P.; Silva, M. da

    Abstract in English:

    The objective of this study was to investigate the capacity of decolorization and detoxification of the textile dyes Reactive Red 198 (RR198), Reactive Blue 214 (RB214), Reactive Blue 21 (RB21) and the mixture of the three dyes (MXD) by Penicillium simplicissimum INCQS 40211. The dye RB21, a phthalocyanine, was totally decolorized in 2 days, and the others, the monoazo RR198, the diazo RB214 and MXD were decolorized after 7 days by P. simplicissimum. Initially the dye decolorization involved dye adsorption by the biomass followed by degradation. The acute toxicity after fungal treatment was monitored with the microcrustacean Daphnia pulex and measured through Effective Concentration 50% (EC50). P. simplicissimum reduced efficiently the toxicity of RB21 from moderately acutely toxic to minor acutely toxic and it also reduced the toxicity of RB214 and MXD, which remained minor acutely toxic. Nevertheless, the fungus increased the toxicity of RR198 despite of the reduction of MXD toxicity, which included this dye. Thus, P. simplicissimum INCQS 40211 was efficient to decolorize different textile dyes and the mixture of them with a significant reduction of their toxicity. In addition this investigation also demonstrated the need of toxicological assays associated to decolorization experiments.
  • Purification and Partial characterization of manganese peroxidase from Bacillus pumilus AND Paenibacillus sp. Industrial Microbiology

    Oliveira, Patrícia Lopes de; Duarte, Marta Cristina Teixeira; Ponezi, Alexandre Nunes; Durrant, Lúcia Regina

    Abstract in English:

    The production of manganese peroxidase (MnP) from Bacillus pumilus and Paenibacillus sp. was studied under absence and presence of the inducers indulin AT, guayacol, veratryl alcohol, lignosulfonic acid and lignosulfonic acid desulfonated. Indulin AT increased the activity of B. pumilus MnP up to 31.66 U/L after 8 h, but no improve was observed for Paenibacillus sp., which reached maximum activity (12.22 U/L) after 20 h. Both MnPs produced by these microorganisms were purified in phenyl sepharose resin and the proteins from crude extracts were eluted in two fractions. However, only the first fraction of each extract exhibited MnP activities. Tests in different pH and temperature values, from pH 5.0 to pH 10.0 and 30 ºC to 60 ºC, respectively, were carried out with the purified MnP. The maximum activity reached for B. pumilus and Paenibacillus sp. MnPs were 4.3 U/L at pH 8.0 and 25 ºC and 11.74 U/L at pH 9.0 and 35 ºC, respectively. The molar masses determined by SDS-PAGE gel eletrophoresis were 25 kDa and 40 kDa, respectively, for the purified enzyme from B. pumilus and Paenibacillus sp.
  • Phylogenetic analysis on the soil bacteria distributed in karst forest Environmental Microbiology

    Zhou, JunPei; Huang, Ying; Mo, MingHe

    Abstract in English:

    Phylogenetic composition of bacterial community in soil of a karst forest was analyzed by culture-independent molecular approach. The bacterial 16S rRNA gene was amplified directly from soil DNA and cloned to generate a library. After screening the clone library by RFLP, 16S rRNA genes of representative clones were sequenced and the bacterial community was analyzed phylogenetically. The 16S rRNA gene inserts of 190 clones randomly selected were analyzed by RFLP and generated 126 different RFLP types. After sequencing, 126 non-chimeric sequences were obtained, generating 113 phylotypes. Phylogenetic analysis revealed that the bacteria distributed in soil of the karst forest included the members assigning into Proteobacteria, Acidobacteria, Planctomycetes, Chloroflexi (Green nonsulfur bacteria), Bacteroidetes, Verrucomicrobia, Nitrospirae, Actinobacteria (High G+C Gram-positive bacteria), Firmicutes (Low G+C Gram-positive bacteria) and candidate divisions (including the SPAM and GN08).
  • Adaptive and cross-protective responses against cadmium and zinc toxicity in cadmium-resistant bacterium isolated from a zinc mine Environmental Microbiology

    Prapagdee, Benjaphorn; Watcharamusik, Anchulee

    Abstract in English:

    Cadmium (Cd) is a major environmental hazard, which usually is detected in its ionic form of Cd2+. It also causes adverse toxic effects on human health and other living organisms. Cd-resistant bacteria were isolated from Cd-contaminated soils. One isolate, TAK1, was highly resistance level to Cd toxicity. TAK1 was isolated from soil contaminated with a high Cd concentration (204.1 mg.kg-1). The result of 16S rDNA sequence analysis found that the TAK1 showed the similarity to Ralstonia sp. Physiological adaptive and cross-protective responses to Cd and Zn killing were investigated in Ralstonia sp.TAK1. Exposure to a low concentration of Cd induced adaptive resistance to higher concentrations of Cd. In addition, pretreatment of Ralstonia sp.TAK1 with an inducing concentration of Cd conferred cross-protective response against subsequent exposure to the lethal concentrations of Zn. The induced adaptive and cross-protective response Ralstonia sp.TAK1 required newly synthesized protein(s). Cd-induced adaptive and cross-protective responses against Cd and Zn toxicity are the important mechanisms used by Ralstonia sp.TAK1 to survive in the heavy metal contaminated environments. These findings might lead to the use of Ralstonia sp.TAK1 for microbial based remediation in Cd and Zn-contaminated soils.
  • Agaricales Fungi from atlantic rain forest fragments in Minas Gerais, Brazil Environmental Microbiology

    Rosa, Luiz Henrique; Capelari, Marina

    Abstract in English:

    Two Atlantic Rain Forest fragments in Minas Gerais state were studied to access their Agaricales fungal richness. A total of 187 specimens were collected and 109 species, 39 genera, and eight families were identified. Thirty-three species were cited for the first time in Brazil.
  • Genetic diversity of indigenous common bean (Phaseolus vulgaris L.) rhizobia from the state of Minas Gerais, Brazil Environmental Microbiology

    Torres, Adalgisa Ribeiro; Cursino, Luciana; Muro-Abad, Júpiter Israel; Gomes, Eliane Aparecida; Araújo, Elza Fernandes de; Hungria, Mariangela; Cassini, Sérvio Túlio Alves

    Abstract in English:

    We characterized indigenous common bean rhizobia from five districts of the state of Minas Gerais, Brazil. The isolates were trapped by two common bean varieties, the Mineiro Precoce (Andean origin) and Ouro Negro (Mesoamerican origin). Analysis by BOX-PCR of selected isolates detected a high level of genetic diversity.
  • Virioplankton Abundance in Trophic Gradients of an Upwelling Field Environmental Microbiology

    Pereira, G.C.; Granato, A.; Figueiredo, A.R.; Ebecken, N.F.F.

    Abstract in English:

    This work correlates time series of biological and physical variables to the marine viruses across trophic gradients within Arraial do Cabo upwelling system, Southeast of Brazil. The objective is to investigate the major controlling factors of virioplankton dynamics among different water masses. It was used an in situ and ex situ flow cytometry for accessing the plankton community. Viruses were highly correlated to bacteria and phytoplankton, but although the lack of direct correlation with physicals, upwelling turned out to be the main contributing factor to the highest values of viral abundance and virus:bacterial ratio. Our data suggest that the lowest temperature of upwelled South Atlantic Central Waters would help to maintain a high viral abundance and higher temperatures of Coastal and Tropical Waters might be another ecological niche allowing the co-existence.
  • Quantification of natural populations of Gluconacetobacter diazotrophicus and Herbaspirillum spp. In sugar cane (Saccharum spp.) Using differente polyclonal antibodies Environmental Microbiology

    Silva-Froufe, Lúcia Gracinda da; Boddey, Robert Michael; Reis, Veronica Massena

    Abstract in English:

    The species Gluconacetobacterdiazotrophicus, Herbaspirillum seropedicae and H. rubrisubalbicans are endophytic N2-fixing [diazotrophic] bacteria which colonise not only roots, but also the aerial tissue of sugar cane. However, the technique most commonly used to quantify the populations of these microbes in plants is by culturing serial dilutions of macerates of plant tissues in N free semi-solid media which are only semi-selective for the species/genera [the Most Probable Number (MPN) Technique] and each culture must be further subjected to several tests to identify the isolates at the species level. The use of species-specific polyclonal antibodies with the indirect ELISA (enzyme-linked immunosorbent assay) can be an alternative which is rapid and specific to quantify these populations of bacteria. This study was performed to investigate the viability of adapting the indirect ELISA technique to quantify individually the populations of these three species of diazotroph within the root and shoot tissues of sugarcane. The results showed that species-specific polyclonal antibodies could be obtained by purifying sera in protein-A columns which removed non-specific immuno-globulins. It was possible to quantify the three bacterial species in the Brazilian sugarcane variety SP 70-1143 in numbers above 10(5) cells per g fresh weight in roots, rhizomes and leaves. The numbers of the different bacterial species evaluated using the ELISA technique were found to be higher than when the same populations were evaluated using the MPN technique, reaching 1400 times greater for G. diazotrophicus and 225 times greater for Herbaspirillum spp. These results constitute the first quantification of Herbaspirillum using immunological techniques.
  • In vitro degradation of linamarin by microorganisms isolated from cassava wastewater treatment lagoons Environmental Microbiology

    Vasconcellos, S. P; Cereda, M. P.; Cagnon, J. R.; Foglio, M.A.; Rodrigues, R.A.; Manfio, G. P.; Oliveira, V. M.

    Abstract in English:

    This study aimed at isolating and characterizing of microorganisms able to use linamarin as sole carbon source. Thirty one microbial strains were isolated from manipueira, a liquid effluent of cassava processing factories. Among these strains, Bacillus licheniformis (isolate 2_2) and Rhodotorulla glutinis (isolate L1) were able to degrade 71% and 95% of added linamarin, respectively, within 7 days, showing high biodegradation activity and great potential for detoxification of cassava processing wastewaters.
  • Degradation of Benzo [a] Pyrene by a novel strain Bacillus subtilis BMT4i (MTCC 9447) Environmental Microbiology

    Lily, Madhuri Kaushish; Bahuguna, Ashutosh; Dangwal, Koushalya; Garg, Veena

    Abstract in English:

    Benzo [a] Pyrene (BaP) is a highly recalcitrant, polycyclic aromatic hydrocarbon (PAH) with high genotoxicity and carcinogenicity. It is formed and released into the environment due to incomplete combustion of fossil fuel and various anthropogenic activities including cigarette smoke and automobile exhausts. The aim of present study is to isolate bacteria which can degrade BaP as a sole source of carbon and energy. We have isolated a novel strain BMT4i (MTCC 9447) of Bacillus subtilis from automobile contaminated soil using BaP (50 μg /ml) as the sole source of carbon and energy in basal salt mineral (BSM) medium. The growth kinetics of BMT4i was studied using CFU method which revealed that BMT4i is able to survive in BaP-BSM medium up to 40 days attaining its peak growth (10(29) fold increase in cell number) on 7 days of incubation. The BaP degradation kinetics of BMT4i was studied using High Performance Liquid Chromatography (HPLC) analysis of BaP biodegradation products. BMT4i started degrading BaP after 24 hours and continued up to 28 days achieving maximum degradation of approximately 84.66 %. The above findings inferred that BMT4i is a very efficient degrader of BaP. To our best of knowledge, this is the first report showing utilization of BaP as a sole source of carbon and energy by bacteria. In addition, BMT4i can degrade a wide range of PAHs including naphthalene, anthracene, and dibenzothiophene therefore, it could serve as a better candidate for bioremediation of PAHs contaminated sites.
  • Isolation and identification of Profenofos degrading bacteria Environmental Microbiology

    Malghani, Saadatullah; Chatterjee, Nivedita; Yu, Hu Xue; Luo, Zejiao

    Abstract in English:

    An enrichment culture technique was used to isolate bacterial strains responsible for the biodegradation of profenofos in a soil from Hubei province of central China. Two pure bacterial cultures, named W and Y, were isolated and subsequently characterized by sequencing of 16S rRNA genes and biochemical tests. Isolate W showed 96% similarity to the 16S rRNA gene of a Pseudomonas putida unlike Y which showed 99% similarity to the 16S rRNA gene of Burkholderia gladioli. Both strains grew well at pH 5.5-7.2 with a broad temperature profile ranging from 28º to 36 ºC. Bioremediation of profenofos-contaminated soil was examined using soil treated with 200 ug g-1; profenofos resulted in a higher degradation rate than control soils without inoculation. In a mineral salt medium (FTW) reduction in profenofos concentration was 90% within 96 hours of incubation. A literature survey revealed that no data is available regarding the role of Burkholderia gladioli on pesticide biodegradation as well as on profenofos.
  • Antineoplasic activity of Agaricus brasiliensis basidiocarps on different maturation phases General Microbiology

    Mourão, Francielly; Linde, Giani Andrea; Messa, Valdeci; Cunha Júnior, Paulo Luiz da; Silva, Aristeu Vieira da; Eira, Augusto Ferreira da; Colauto, Nelson Barros

    Abstract in English:

    The fungus Agaricus brasiliensis is a Basidiomycete studied because of its immunomodulation and/or antitumor substances. The objective of this study was to verify the Agaricus brasiliensis antineoplasic activity in vivo on different basidiocarp maturation phases on Sarcoma 180 cells implanted in mice. Sarcoma cells were implanted in mice and after seven days mice were divided in three groups. The first group was treated with saline solution, the second group was treated with closed basidiocarp extract solution and the third group was treated with opened basidiocarp extract solution. After 30 days of being daily orally treated with these three solutions all animals suffered euthanasia, and the splenic index, tumor mass and volume were determined. No significant differences of the tumor growth inhibition in function of the different basidiocarp maturation phases for the Agaricus brasiliensis strain were observed. The in vivo basidiocarp antineoplasic average activity was 89.22%.
  • Isolation, Purification and Evaluation of Antibacterial Agents from Aloe vera General Microbiology

    Lawrence, Rubina; Tripathi, Priyanka; Jeyakumar, Ebenezer

    Abstract in English:

    The ethanol, methanol and acetone extracts of Aloe vera gel were studied for their antimicrobial activity against four Gram-positive and Gram-negative bacteria using agar well diffusion method. The extracts showed varied levels of antimicrobial activity against the tested pathogens. The ethanol and methanol extracts showed higher activity while acetone extract, showed least or no activity against most of the tested pathogens. Fractions obtained from the extracts by Thin Layer and Column Chromatography were studied for their antagonistic properties using Spot Assay Technique. Compounds with maximum antibacterial activity isolated from the ethanol and methanol extracts were identified as p - coumaric acid (Mol. wt.165), ascorbic acid (Mol. wt.177 ), pyrocatechol (Mol. wt.110 ) and cinnamic acid (Mol. wt.148), on the basis of Gas Chromatography Mass Spectrometry. The study suggests the antimicrobial activity of the A. vera gel extract to be dependant on the synergistic effect of different compounds. With the broad spectral antimicrobial effect of A. vera gel, it could be further recommended in the treatment of various bacterial diseases.
  • Antimicrobial and toxicological studies of some metal complexes of 4-methylpiperazine-1-carbodithioate and phenanthroline mixed ligands General Microbiology

    Kalia, S.B.; Kaushal, G.; Kumar, M.; Cameotra, S.S.; Sharma, A.; Verma, M.L.; Kanwar, S.S.

    Abstract in English:

    A few mixed ligand transition metal carbodithioate complexes of the general formula [M(4-MPipzcdt)x(phen)y]Y (M = Mn(II), Co(II), Zn(II); 4-MPipzcdt = 4-methylpiperazine-1-carbodithioate; phen = 1,10-phenanthroline; x = 1 and y = 2 when Y = Cl; x = 2 and y = 1 when Y = nil) were synthesized and screened for their antimicrobial activity against Candida albicans, Escherichia coli, Pseudomonas aeruginosa,Staphylococcus aureus andEnterococcusfaecalis by disk diffusion method. All the complexes exhibited prominent antimicrobial activity against tested pathogenic strains with the MIC values in the range <8-512 μgmL-1. The complexes [Mn(4-MPipzcdt)2(phen)] and [Co(4-MPipzcdt)(phen)2]Cl inhibited the growth of Candida albicans at a concentration as low as 8 µgmL-1.The complexes were also evaluated for their toxicity towards human transformed rhabdomyosarcoma cells (RD cells). Moderate cell viability of the RD cells was exhibited against the metal complexes.
  • Glass bead transformation method for gram-positive bacteria General Microbiology

    Rattanachaikunsopon, Pongsak; Phumkhachorn, Parichat

    Abstract in English:

    A simple, inexpensive and reproducible transformation method was developed for Gram-positive bacteria. It was based on agitation of bacterial protoplasts with glass beads in the presence of DNA and polyethylene glycol. By using this method, introduction of pGK12 into protoplasts of several strains of Gram-positive bacteria was achieved.
  • Molecular characterization of Verocytotoxigenic Escherichia coli O157:H7 isolates from Argentina by Multiple-Loci VNTR Analysis (MLVA) General Microbiology

    Bustamante, Ana V.; Lucchesi, Paula M.A.; Parma, Alberto E.

    Abstract in English:

    The aim of this work was to adapt described MLVA protocols to the molecular typing and characterization of VTEC O157:H7 isolates from Argentina. Nine VNTR loci were amplified by PCR showing diversity values from 0.49 to 0.73. Nine MLVA profiles were observed and the cluster analysis indicated both unrelated and closely related VTEC O157:H7 strains. In spite of the limited number of isolates studied, the panel of VNTR used made it possible to perform a first approach of the high genetic diversity of native strains of O157:H7 by MLVA.
  • Mechanism of Sterigmatocystin Biosynthesis Regulation by pH in Aspergillus nidulans General Microbiology

    Delgado-Virgen, Francisco; Guzman-de-Peña, Doralinda

    Abstract in English:

    External pH constitutes one of the most important environmental factors that control growth, metabolism and differentiation in microorganisms, including fungi. We have analyzed the effect of external pH on sterigmatocystin biosynthesis in Aspergillus nidulans. It was observed in repeated experiments that alkaline pH, in opposition to acid pH, increased sterigmatocystin production and the transcript levels of aflR, the master gene that regulates expression of the sterigmatocystin cluster in A. nidulans. It is known that pH effects in fungi operate mostly through the Pal/Pac signaling pathway, originally described in Aspergillus nidulans. Accordingly, we studied the role of this signaling pathway in ST biosynthesis. It was observed that aflR transcript levels were increased in the "alkalinity mimicking" mutant pacCc14 and were minimal in the "acidity mimicking" mutant palA1. No sterigmatocystin was produced by palA1 or pacC- mutants at neither acid or alkaline pH of incubation. Finally, fluG and flbA, genes known to regulate both conidiation and sterigmatocystin synthesis upstream in the regulatory cascade, were up-regulated at alkaline pH.
  • Mutants with heteroresistance to amphotericin B and fluconazole in Candida Medical Microbiology

    Claudino, A.L.R.; Peixoto Junior, R.F.; Melhem, M.S.C.; Szeszs, M.W.; Lyon, J.P.; Chavasco, J.K.; Franco, M.C.

    Abstract in English:

    Several studies have reported the occurrence of infections caused by Candida yeasts as well as the increasing prevalence of non albicans species. The aim of the present work is focused on the obtaining of heteroresistance to amphotericin B and fluconazole in Candida species using two distinct methodologies: selection and induction. Resistant samples were obtained by selective pressure using a medium with fluconazole for growth, followed by growth in a medium with amphotericin B. The selective pressure was also created beginning with growth in amphotericin B medium followed by growth in fluconazole medium. Concomitantly, samples were submitted to the induction of resistance through cultivation in increasing concentrations of fluconazole, followed by cultivation in increasing concentrations of amphotericin B. Subsequently, the induction began with amphotericin B followed by fluconazole. Three samples resistant to fluconazole and amphotericin B were obtained, two by induction (C. glabrata and C. tropicalis) and one by selection (C. tropicalis). Both C. tropicalis originated from the same wild sample. After successive transfers for drug free medium, only the sample obtained by selection was able to maintain the resistance phenotype. These results suggest that the phenotype of heteroresitance to fluconazole and amphotericin B can be produced by two methodologies: selection and induction.
  • Use of pcr-rflp of thefla a gene for detection and subtyping of Campylobacter jejuni strains Potentially related to Guillain-barré syndrome, isolated from humans and animals Medical Microbiology

    Scarcelli, E.; Piatti, R.M.; Harakava, R.; Miyashiro, S.; Campos, F.R.; Souza, M.C.A.; Cardoso, M.V.; Teixeira, S.R.; Genovez, M.E.

    Abstract in English:

    The objectives of the present study were the subtyping of Campylobacter jejuni subsp. jejuni strains obtained from humans and different animal species using PCR-RFLP, and the detection, by means of the same technique, of strains related to serotype PEN O19:LIO 7, the main C. jejuni serotype linked to Guillain-Barré Syndrome (GBS). Seventy C. jejuni strains isolated from human feces (n=33), primates (n=15), dogs (n=5), swine (n=2), bovines (n=1), abortion material from goats (n=2) and poultry carcasses (n=12), all collected in the state of São Paulo, were subtyped by means of PCR-RFLP of fla A gene, using restriction endonucleases Hae III, Afa I and Mbo I. Seven subtypes were observed when using the enzyme Hae III; eight when using Mbo I; and seven when using Afa I. The combination of the three endonucleases led to 16 fla-RFLP subtypes, from which ten subtypes shared strains of human and animal origin. From these, seven subtypes were observed in human and broiler strains. In eight subtypes, the other animal species shared patterns with human strains. It was inferred that, besides broilers, swine, goats, dogs and primates may be sources of infection for human in São Paulo. PCR-RFLP is a highly discriminatory technique that may be applied to molecular epidemiology studies of samples from different origins. Besides, the study also enabled the detection of two human strains and two primate strains related to serotype PEN O19: LIO 7.
  • Influence of probiotics on Candida presence and IgA anti-Candida in the oral cavity Medical Microbiology

    Santos, Agda Lima dos; Jorge, Antônio Olavo Cardoso; Santos, Silvana Soléo Ferreira dos; Silva, Célia Regina Gonçalves e; Leão, Mariella Vieira Pereira

    Abstract in English:

    Probiotics are defined as microorganisms that promote benefits to host health, mainly by regulating resident microbiota. Disequilibrium in microbiota can favor the growth of opportunist microorganisms and the development of pathologies, like candidosis caused by yeasts of the Candida genus. This work evaluated whether probiotics consumption was able to influence a specific immunological response to Candida and the presence of these yeasts in the oral cavity. Saliva samples were collected from healthy individuals and plated in Dextrose Saboraud Agar with chloramphenicol. Individuals presenting Candida in the oral cavity used the probiotic Yakult LBâ for 20 days, after which new collections and identifications were performed. Anti-Candida IgA analysis was conducted using the ELISA technique. Analysis of the results showed a significant reduction in Candida prevalence (46%) and mean Candida CFU/mL counts (65%). The Candida species identified were C. albicans (98%) and C.tropicalis (2%), before and after probiotics consumption. Immunological analysis demonstrated a significant reduction in anti-Candida IgA levels after probiotics use, probably due to less antigenic stimulation. In conclusion, in the individuals studied, probiotics use significantly reduced the amount of Candida in the oral cavity, possibly due to competition between the yeasts rather than by specific secretory immune response stimulation.
  • Prevalence of the precore G1896A mutation in Chinese patients with e antigen negative hepatitis B virus infection and its relationship to pre-S1 antigen Medical Microbiology

    Zhang, Jing; Xu, Wei-Jia; Wang, Qing; Zhang, Yong; Shi, Ming

    Abstract in English:

    This study investigated the prevalence of the precore G1896A mutation in Chinese patients with hepatitis B e antigen (HBeAg) negative HBV infection and its relation to serum HBV pre-S1 antigen. The overall prevalence of the precore G1896A mutation was 72.6% in HBeAg-negative Chinese patients with detectable serum HBV DNA. The prevalence of the precore G1896A is significantly higher in Chinese HBeAg-negative patients with chronic hepatitis B than that in inactive HBV carriers with detectable serum HBV DNA. Serum pre-S1 and the precore G1896A mutation were simultaneously detected in most of Chinese HBeAg-negative patients.
  • Characterization of recombinant MSP5 Anaplasma marginale Havana isolate Veterinarian Microbiology

    Corona, B.; Machado, H.; Rodríguez, M.; Martínez, S.

    Abstract in English:

    Anaplasma marginale is the causative agent of bovine anaplasmosis, a disease of worldwide economic importance. Major surface proteins (MSPs) are involved in host-pathogen and tick-pathogen interactions and they have been used as markers for the genetic characterization of A. marginale strains and phylogenetic studies. The major surface protein 5 (MSP5) is highly conserved in the genus Anaplasma and in all isolates of A. marginale. The aim of the present work was to carry out the cloning, sequencing and characterization of the recombinant MSP5 Anaplasma marginale Havana isolate. The sequence of the msp5 gene of Anaplasma marginale Havana isolate with a size of 633 pb was determined (Acc. No. AY527217). This gene was cloned into pRSETB vector and expressed in Escherichia coli. The MSP5 protein was recognized by the monoclonal antibody ANAF16C1 and it showed a high similitude percent with the gene sequence described for other Anaplasma marginale isolates. These data are very important for the development of a diagnostic test for A. marginale using the MSP5 recombinant protein.
  • Postantibiotic effects and postantibiotic sub-MIC effects of tilmicosin, erythromycin and tiamulin on erythromycin-resistant Streptococcus suis Veterinarian Microbiology

    Wang, Liping; Zhang, Yuanshu

    Abstract in English:

    The postantibiotic effects (PAEs) and postantibiotic sub-MIC effects (PA SMEs) of tilmicosin, erythromycin and tiamulin on erythromycin-susceptible and erythromycin-resistant strains of Streptococcus suis (M phenotype) were investigated in vitro. Tilmicosin and tiamulin induced significantly longer PAE and PA SME against both erythromycin-susceptible and erythromycin-resistant strains than did erythromycin. The durations of PAE and PA SMEs were proportional to the concentrations of drugs used for exposure. The PA SMEs were substantially longer than PAEs on S. suis (P<0.05) regardless of the antimicrobial used for exposure. The results indicated that the PAE and PA SME could help in the design of efficient control strategies for infection especially caused by erythromycin-resistant S. suis and that they may provide additional valuable information for the rational drug use in clinical practice.
  • A RT-PCR assay for the detection of encephalomycarditis virus infections in pigs Veterinarian Microbiology

    Pérez, Lester J.; Díaz de Arce, Heidy

    Abstract in English:

    Encephalomyocarditis virus (EMCV) infections can cause losses in pig farms all over the world. Rapid, sensitive and unequivocal detection of this virus is therefore essential for the diagnosis and control of the disease. An RT-PCR assay was developed, optimized and evaluated for encephalomyocarditis virus detection in organ based on a pair of primers that amplifies a 165 bp DNA fragment from a highly conserved nucleotide region of the viral 3D glycoprotein. PCR products of the expected size were obtained from Cuban EMCV 744/03 strain. Non-specific reactions were not observed when other porcine RNA genome viruses and uninfected cells were used. The analytical sensitivity of the test was estimated to be 2 TCID50/50 mL. The analysis of tissue homogenate samples from naturally infected animals proved the potential usefulness of the method for a rapid disease diagnosis from field cases.
  • The contribution of genes required for anaerobic respiration to the virulence of Salmonella enterica serovar Gallinarum for chickens Veterinarian Microbiology

    Paiva, J.B.; Penha Filho, R.A.C.; Pereira, E.A.; Lemos, M.V.F.; Barrow, P.A.; Lovell, M.A.; Berchieri Jr, A.

    Abstract in English:

    Salmonella enterica serovar Gallinarum (SG) is an intracellular pathogen of chickens. To survive, to invade and to multiply in the intestinal tract and intracellularly it depends on its ability to produce energy in anaerobic conditions. The fumarate reductase (frdABCD), dimethyl sulfoxide (DMSO)-trimethylamine N-oxide (TMAO) reductase (dmsABC), and nitrate reductase (narGHIJ) operons in Salmonella Typhimurium (STM) encode enzymes involved in anaerobic respiration to the electron acceptors fumarate, DMSO, TMAO, and nitrate, respectively. They are regulated in response to nitrate and oxygen availability and changes in cell growth rate. In this study mortality rates of chickens challenged with mutants of Salmonella Gallinarum, which were defective in utilising anaerobic electron acceptors, were assessed in comparison to group of bird challenged with wild strain. The greatest degree of attenuation was observed with mutations affecting nitrate reductase (napA, narG) with additional attenuations induced by a mutation affecting fumarate reductase (frdA) and a double mutant (dmsA torC) affecting DMSO and TMAO reductase.
  • Increased presevation of sliced mozzarella cheese by antimibrobial sachet incorporated with allyl isothiocyanate Food Microbiology

    Pires, Ana Clarissa dos Santos; Soares, Nilda de Fátima Ferreira; Andrade, Nélio José de; Silva, Luis Henrique Mendes da; Camilloto, Geany Peruch; Bernardes, Patrícia Campos

    Abstract in English:

    There is an increasing tendency to add natural antimicrobials of plant origin into food. The objective of this work was to develop a microbial sachet incorporated with allyl isothiocyanate (AIT), a volatile compound of plant origin, and to test its efficiency against growth of yeasts and molds, Staphylococcus sp. and psychrotrophic bacteria on sliced mozzarella cheese. Another objective was to quantify the concentration of AIT in the headspace of cheese packaging. A reduction of 3.6 log cycles was observed in yeasts and molds counts in the mozzarella packed with the antimicrobial sachet over 15-day storage time. The sachet also showed an antibacterial effect on Staphylococcus sp., reducing 2.4 log cycles after 12-day storage. Psychrotrophic bacteria species were the most resistant to the antimicrobial action. The highest concentration of AIT (0.08µg.mL-1) inside the active packaging system was observed at the 6-day of storage at 12 ºC ± 2 ºC. At the end of the storage time, AIT concentration decreased to only 10% of the initial concentration. Active packaging containing antimicrobial sachet has a potential use for sliced mozzarella, with molds and yeasts being the most sensitive to the antimicrobial effects.
Sociedade Brasileira de Microbiologia USP - ICB III - Dep. de Microbiologia, Sociedade Brasileira de Microbiologia, Av. Prof. Lineu Prestes, 2415, Cidade Universitária, 05508-900 São Paulo, SP - Brasil, Ramal USP 7979, Tel. / Fax: (55 11) 3813-9647 ou 3037-7095 - São Paulo - SP - Brazil
E-mail: bjm@sbmicrobiologia.org.br