摘要:

Health effects associated with poor indoor air quality have created a need for accurate, reproducible methods of monitoring the microbiological content of indoor air. Improved methods of detection may allow researchers to clarify the effect of individual species present in the indoor environment on human health. This review discusses the shortcomings of current methods of identification and detection and focuses on the potential for molecular techniques in this emerging field. Probe techniques, restriction endonuclease analysis, karyotyping, and DNA and polymerase chain reaction fingerprinting methods available to detect and identify bacteria and fungi significant in the indoor air environment are discussed. Problems that may be encountered using these techniques are also considered. The authors have included a brief discussion on current air sampling techniques as well as adapting these techniques for use with molecular detection methods.Key words: indoor air microbiology, microbiological air sampling, molecular detection methods.

ISSN:0008-4166    

DOI:10.1139/m95-091

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

The bacterial community in the gut of the earthwormLumbricus terrestriswas analyzed by whole-cell hybridization with 16S rRNA targeted oligonucleotide probes. Whole-cell hybridization protocols using fluorescence-, peroxidase-, or digoxigenin-labeled oligonucleotide probes facilitated detection of significant fractions of bacterial cells stained with 4′,6-diamidino-2-phenylindole (DAPI) in the fore-, mid-, and hind-gut and cast of the earthworm. The application of peroxidase- and digoxigenin-labeled probes, however, was hampered by several methodological drawbacks: the requirement of enzymatic permeabilization, the diffuse images of stained cells, and the incompatibility with DAPI staining used as control. Quantitative analysis of the bacterial community was also influenced by its considerable variability in different individual earthworms. Though the number of bacteria detected by DAPI staining as well as by whole-cell hybridization with the fluorescent eubacterial probe Eub338 generally showed a significant increase in the number of bacteria towards the end of the gut, a decrease in bacterial numbers could be found in some earthworms. In situ analysis of the bacterial community in the fore-, mid-, and hind-gut of one individual earthworm by whole-cell hybridization with the fluorescent eubacterial probe Eub338 recorded 15, 30, and 25% of DAPI-stained bacteria, respectively. In the cast 37% of the bacteria were detected. Similar to counts obtained by DAPI and by whole-cell hybridization with probe Eub338, the number of bacteria belonging to the α-, β-, and γ-subgroups of proteobacteria increased significantly towards the end of the gut and remained high in the cast. While the most significant difference in the counts of bacteria belonging to the α-subgroup was obtained between the hind-gut and cast, bacterial populations of the β- and γ- subgroups of proteobacteria increased most prominently between the fore- and hind-gut.Key words: digoxigenin, fluorescent probes, in situ detection,Lumbricus terrestris, rRNA, whole-cell hybridization.

ISSN:0008-4166    

DOI:10.1139/m95-092

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

A cosmid able to complement the Nif−and nitrate-dependent growth phenotypes of theAzospirillum brasilensemutant FP9 was isolated from a genomic library of the wild-type strain FP2. A 6-kb DNA region was sequenced and showed two open reading frames (ORFs) identified as thentrBandntrCgenes. An ORF1 located upstream from thentrBgene and coding for a 36-kDa polypeptide showed similarity to thenifR3gene ofRhodobacter capsulatusand the ORF1 ofRhizobium leguminosarum, both located upstream from thentrBgene in a complex operon. Two other unidentified ORFs (ORF5 and partial ORF4) coding for hydrophobic polypeptides were also observed. ΔORF1-ntrBC, ORF1,ntrB, andntrCmutants obtained by recombination of suicide plasmids containing an insertion of a promoterlesslacZkanamycin cassette showed decreased nitrogenase activities and were unable to grow on nitrate as the sole N source. These phenotypes were restored by complementation with plasmids containing thentrCgene. Analysis oflacZtranscriptional fusions suggested that the ORF1-ntrBCoperon inAzospirillum brasilenseis expressed from a promoter located upstream from the ORF1 and that it is negatively regulated by thentrCgene product.Key words:Azospirillum brasilense,ntrB,ntrC,nifR3-like, nitrogen fixation.

ISSN:0008-4166    

DOI:10.1139/m95-093

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

An increase in early injury to the roots and collar of sunflower plants caused bySclerotinia sclerotiorumhas been observed in France for several years. In vitro tests (inhibition of fungi and cyanide production by bacterial strains) and in situ tests (in nonsterile humus, in a growth chamber) were performed to screen the most efficientPseudomonas fluorescensandPseudomonas putidastrains effective against this form of injury caused byS.sclerotiorum. Although there was no correlation between in vitro and in situ tests, a positive correlation between in situ tests and field experiments was obtained. At least 1 × 106bacteria per seed were required for significant protection in in situ tests and field trials demonstrated that significant protection of sunflower was obtained by seed bacterization with selections ofP.fluorescensandP.putida.Key words: biological control,Pseudomonas fluorescens,Pseudomonas putida,Sclerotinia sclerotiorum, sunflower, bacterization.

ISSN:0008-4166    

DOI:10.1139/m95-094

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

Glucan synthesis was sensitive to several sulfhydryl reacting compounds: mercurials, reversible disulfides, and an alkylating sulfhydryl reagent (IC503–45 μM). Thiol groups associated with glucan synthesis were hydrophilic in nature, since both hydrophilic and hydrophobic reagents were active. Glucan synthase complex consists of at least two components: a peripheral GTP-binding protein that can be solubilized with detergents (supernatant) and the catalytic membrane-bound component (pellet). A rapid separation technique was developed to study sulfhydryl interactions with the complex. The GTP-binding protein was solubilized with 0.6% 3-((3-cholamidopropyl)dimethylammonio)-1-propane sulfonate from isolated microsomes ofCandida albicanscells grown at either 10 or 30 °C. The residual membranous fraction contained the core catalytic moiety of glucan synthase. Both fractions were devoid of glucan synthase activity until they were reconstituted by mixing the two fractions together. In reconstitution experiments, the pellet lost almost 50% activity when preincubated with 2.5 μMN-ethylmaleimide and combined with an untreated supernatant whereas only 10% activity was lost when the supernatant was treated withN-ethylmaleimide. The catalytic active site of glucan synthase was not protected with UDP-Glc when preincubated with 10 μMN-ethylmaleimide but the GTP-binding fraction was partially protected with GTPγS.Key words:Candida albicans, (1,3)-β-glucan synthase, GTP-binding proteins, solubilization, sulfhydryl reagents.

ISSN:0008-4166    

DOI:10.1139/m95-095

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

A new rod-shaped endospore-forming bacterium is described, which was isolated from the hindgut of the termiteReticulitermes santonensis(Feytaud). The isolate stains Gram negative and its DNA has a guanine-plus-cytosine content of 35 mol%. Despite the Gram-staining reaction, both biochemical and physiological features place the isolate in the genusBacillusand indicate a phenotypic resemblance to theBacillus firmus–lentusgroup of species. On the basis of comparative 16S rRNA analysis and some phenotypic features the isolate clearly represents a new species for which the nameBacillus oleroniusis proposed. The type strain isBacillus oleroniusRt 10 (DSM 9356).Key words:Bacillus, termites, hindgut flora.

ISSN:0008-4166    

DOI:10.1139/m95-096

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

Root colonization and in vitro carbon substrate utilization by two seedling growth-promotingBacillusstrains that originated from different root microsites were studied in greenhouse and growth chamber experiments. Strain L6, identified asBacillus polymyxa, was previously isolated from rhizosphere soil containing roots of pasture plants, and Pw-2, tentatively identified also asB.polymyxa, was isolated from within surface-sterilized lodgepole pine (Pinus contortavar.latifolia(Dougl.) Engelm.) roots. Rifamycin-resistant strains derived spontaneously from wild-type strains L6 and Pw-2, designated strain L6-16R and Pw-2R, respectively, were used to monitor lodgepole pine root colonization in a closed tube assay system. Three-week-old pine seedlings were inoculated with 105colony-forming units (cfu) of strain Pw-2R or 106 cfu of strain L6-16R, and external and internal root colonization was assessed 2 and 4 weeks later. Strains L6-16R and Pw-2R were both recovered from pine rhizosphere samples with > 5 × 107 cfu/g fresh root tissue 2 weeks after inoculation, but neither strain was detected in the root interior. When root colonization was assessed 4 weeks after inoculation, the rhizosphere populations of both strains had declined slightly to between 5 × 106and 5 × 107 cfu/g fresh root tissue, but strain Pw-2R was also detected within root tissues with 105 cfu/g fresh root tissue. Lateral root formation was abundant 4 weeks after inoculation and may have facilitated colonization of internal root tissues by strain Pw-2R. Both strains possessed pectolytic activity, although differences between the strains were detected in in vitro substrate utilization capabilities using BIOLOG assays. These differences may be related to their abilities to colonize internal root tissues. On the basis of our results, we hypothesize that internal root colonization byBacillusstrains is not a random event and that root-endophyticBacillusstrains possess specific physiological and (or) biochemical characteristics that facilitate colonization of internal root tissues.Key words:Bacillus, PGPR, rhizosphere, endophytes, colonization.

ISSN:0008-4166    

DOI:10.1139/m95-097

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

Culture conditions that lead to swelling and germination dramatically influence cell surface characteristics and properties ofAspergillus fumigatusconidia. Conidial adherence to polystyrene and agglutination markedly increased during swelling, in a time-dependent manner. Agglutination appeared to be sensitive to cycloheximide and calcium. Removal of cell wall polysaccharides by lyticase or sodium metaperiodate suppressed agglutination of conidia. Proteinase K weakly decreased it whereas dithiothreitol strongly dispersed the cells. These observations suggest that both cell surface carbohydrates and proteins are involved in the agglutination process. Electron microscopic observations demonstrated that the cell wall of conidia was subject to some rearrangements during swelling, involving degradation and loss of the external convoluted layer, and subsequent exposure of underlying ligands. This was confirmed using lectins labelled with gold or fluorescein isothiocyanate, which showed that some carbohydrates, particularly those acting as ligands for peanut agglutinin, are largely exposed during the process. Finally, SDS-PAGE revealed major protein changes between resting and swollen conidia. We conclude that the ability ofA.fumigatusconidia to aggregate correlates with an increase in adherence and biochemical reorganization of the cell wall.Key words:Aspergillus fumigatus, adherence, agglutination, cell wall rearrangements.

ISSN:0008-4166    

DOI:10.1139/m95-098

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

The desertomycin action uponSaccharomyces uvarumwall synthesis has been studied. Spheroblast regeneration was carried out in a liquid medium containing labeled glucose to monitor the synthesis of different wall components. In the presence of desertomycin, wall synthesis was affected; this was expressed as a net reduction of insoluble alkali constituents content, more precisely the insoluble acido-alkali fraction that, in yeasts, is constituted by chains of β(1,3)-glucans linked among themselves by β(1,6) bonds. Mannan formation was not inhibited; such polymers that cannot be fixed to the glucan matrix of the wall were liberated in the regeneration medium. Because of desertomycin action, the decrease in insoluble alkali content revealed an interference with the enzymatic systems catalyzing glucan synthesis. In vitro, however, this antifungal had little effect upon glucan synthetase activity: doses 5 times superior to the subinhibiting level used in vivo caused only 30% inhibition. This result can be explained by an indirect action of desertomycin. Parietal disorders were the result of membrane structure disturbance, notably the phospholipids and localized enzymatic systems. This antifungal presents an analogical structure with macrolides with recognized membrane action.Key words: desertomycin, wall, yeast.

ISSN:0008-4166    

DOI:10.1139/m95-099

出版商:NRC Research Press    

年代:1995

数据来源: NRC

摘要:

Seven bacterial strains capable of utilizing monochloroacetate (MCA) at a concentration of 50 mM as the sole carbon source were isolated from soil and displayed MCA dehalogenase activity. Three of them were identified asPseudomonasspp., and the remaining four strains asAlcaligenessp.,Agrobacteriumsp.,Arthrobactersp., andAzotobactersp. This latter is the first reported example of a bacterium fixing atmospheric nitrogen under aerobic conditions that also uses a chloro-organic compound as sole source of carbon and energy. MCA dehalogenase activity in these strains was found to be inducible under different growth conditions. Crude extracts from all seven isolated strains also displayed dehalogenating activity with a relatively wide range of halogenated organic compounds (aliphatic acids, ketones, alcohols, alkanes, and aromatics), which, depending on the strain, were dehalogenated to different extents. The estimatedKmvalues for MCA were used to classify the dehalogenase activities into three groups: high affinity (30–40 μM) inAlcaligenesandAgrobacteriumspecies, medium affinity (100–180 μM) inPseudomonasandAzotobacterspecies, and low affinity (100 mM) inArthrobactersp. Both the optimal pH range for MCA dehalogenase activity (between pH 8 and 10) and the pH profile of stability (in the neutral–basic range) were found to be similar in all strains, whereas the thermal stability profiles were variable.Key words: dehalogenase, halohydrolase, monochloroacetate, soil.

ISSN:0008-4166    

DOI:10.1139/m95-100

出版商:NRC Research Press    

年代:1995

数据来源: NRC