摘要:

To gain more information about the determination of cell polarity and its relationship to the organisation of cytoskeleton, we have examined the mycelial mutantsep1-1and the multinucleate multipolar syncytia of the triple mutantsep1-1 spl1-1 cdc4-8by indirect immunofluorescence techniques. We have found that polarity is predetermined by the shape of the cell. During transition from mitosis to interphase the microtubules of the arising cytoplasmic cytoskeleton gradually form a basket-like pattern that reflects the curvatures of the cell envelope. The presumable growing poles, where actin accumulates, usually correlates with the sites where the cell tapers and the microtubules converge. However, no growth can be launched at these sites if the cell surface has not been properly processed. Mitosis and meiosis are not affected significantly by changes in cell morphology and polarity, but larger cells are less effective during sporulation. The azygotic asci produced by multinucleate syncytia frequently contain over 20 ascospores.Key words: cell division cycle, cytokinesis, cytoskeleton, fission yeast.

ISSN:0008-4166    

DOI:10.1139/m97-143

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

In continuous flow biofilm cultures in medium resembling cystic fibrosis bronchial secretions,Pseudomonas aeruginosawas not eradicated from biofilms by 1 week of treatment with high concentrations of ceftazidime and gentamicin, to which the strains were sensitive on conventional testing. The addition of rifampicin, which has little activity against the strains as measured by the minimum inhibitory concentration, led to the apparent elimination of the bacteria from the biofilms. The effect was not strain specific.Key words:Pseudomonas aeruginosa, biofilm, rifampicin.

ISSN:0008-4166    

DOI:10.1139/m97-144

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

Glutamate dehydrogenase is an enzyme responsible for ammonium assimilation and glutamate catabolism in organisms. The tylosin producerStreptomyces fradiaepossesses both NADP- and NAD-dependent glutamate dehydrogenases. The latter enzyme was purified 498-fold with a 7.5% recovery by a six-step protocol. The enzyme is composed of two subunits, each ofMr47 000, and could form active aggregates of four or eight subunits. Its activity was inactivated by alkaline pH or temperatures of −20 °C or above 40 °C. Activities assayed in the direction of oxidative deamination and reductive amination were optimal at pH 9.2 and 8.8, respectively, and at temperatures of 30–35 °C. No activity was found when NAD(H) was replaced with NADP(H). TheKmvalues were 32.2 mM forL-glutamate, 0.3 mM for NAD+, 3.4 mM for 2-ketoglutarate, 14.2 mM for NH4+, and 0.05 mM for NADH. Deamination activity was partially inhibited by adenyl nucleotides and several divalent cations; amination activity was not affected by the nucleotides but significantly inhibited by Cu2+or Ni2+.Key words:Streptomyces fradiae, NAD-dependent glutamate dehydrogenase, purification, properties.

ISSN:0008-4166    

DOI:10.1139/m97-145

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

The causal agents of Dutch elm disease,Ophiostoma ulmi(isolate H200) andOphiostoma novo-ulmi(isolate CKT-11), secreted similar amounts of β-galactosidase in liquid shake cultures when grown on galacturonic acid or sodium pectate (1.45 ± 0.16 and 1.03 ± 0.24 nkat∙mL−1forO.ulmi, respectively, and 1.30 ± 0.08 and 1.28 ± 0.26 nkat∙mL−1forO.novo-ulmi, respectively). Rhamnose and pectin also stimulated secretion but to a lesser extent, whereas on glucose, enzyme activity was barely detectable (≤0.01 nkat∙mL−1).Ophiostoma novo-ulmiwas shown by Q-Sepharose chromatography to form two β-galactosidases, named β-galactosidases I and II. In cultures grown on galacturonic acid β-galactosidase I accounted for approximately 75% of the total activity in the culture filtrate. β-Galactosidase I was further purified to apparent electrophoretic homogeneity by means of Sephacryl gel filtration chromatography, chromatofocusing, and Superdex75 gel filtration. The molecular mass of the enzyme was 135 kDa by SDS–PAGE and 123 kDa by gel filtration. Its isoelectric point, determined by chromatofocusing, was 4.9. The optimal pH for enzyme activity was 5.8 and the optimal temperature was 50 °C. TheKmvalues forp-nitrophenyl β-D-galactopyranoside and lactose were 7.52 and 14.23 mM, respectively, and the maximum velocities for these substrates were 1733 and 355 nkat∙mg protein−1, respectively. TheKivalue forD(−)-galactonic acid γ-lactone was 2.29 mM.Key words: Dutch elm disease, β-galactosidase,Ophiostoma ulmi,Ophiostoma no

ISSN:0008-4166    

DOI:10.1139/m97-146

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

The bacterial populations associated with different plant and fungal habitats of intactPinus sylvestris–Suillus bovinusorPinus sylvestris–Paxillus involutusectomycorrhizospheres grown in natural forest soil were examined by scanning and transmission electron microscopy. Surfaces of nonmycorrhizalPinus sylvestrisroots hosted large numbers of morphologically distinct bacteria. Bacteria were detected on the mantle surfaces and at inter- and intra-cellular locations in the mantle and Hartig net ofSuillus bovinusmycorrhizas. The fungal strands were colonized by only a few bacteria unlike the outermost external fine hyphae on which extensive monolayers of bacteria were attached. The mycorrhizas ofPaxillus involutuswere mostly devoid of bacteria, but the intact external mycelium supported both bacterial colonies and solitary bacteria. Intracellular bacteria were not present inPaxillus involutushyphae. In both mycorrhizal systems, bacterial aggregation and attachment to hyphae were mediated with electron-dense or -translucent material. Our study shows that thePinus sylvestrismycorrhizospheres formed by two different ectomycorrhizal fungi are clearly dissimilar habitats for mycorrhizosphere-associated bacteria. Additionally, the spatially and physiologically defined mycorrhizosphere habitats were shown to host distinct populations of bacteria.Key words: ectomycorrhiza, intracellular bacteria,Paxillus involutus, soil bacteria,Suillus bovinus.

ISSN:0008-4166    

DOI:10.1139/m97-147

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

We investigated the effect of visible light onEscherichia coliin seawater microcosms.Escherichia colilost its ability to form colonies in marine environments when exposed to artificial continuous visible light. Survival of illuminated bacteria during the stationary phase was drastically reduced in the absence of the σsfactor (RpoS or KatF) that regulates numerous genes induced in this phase. In the stationary phase, double catalase mutantskatE katGand mutants defective in the protein Dps (both catalase and Dps are involved in resistance to hydrogen peroxide (H2O2)), were more sensitive to light. In the exponential phase, a mutation inoxyR, the regulatory gene of the adaptive response to H2O2, increased sensitivity to light, further suggesting that deleterious effects might be associated with H2O2production. However, in the stationary phase, thekatE katG dpsmutant was considerably more resistant to visible light than therpoSmutant, suggestingrpoS-dependent protection against deleterious effects other than those related to H2O2. The deleterious action of visible light was less important when the salinity decreased. In freshwater,rpoSandkatE katG dpsmutants did not show a drastic difference in sensitivity to light suggesting that osmolarity sensitizesE.colito those deleterious effects of visible light that are unrelated to H2O2.Key words:Escherichia coli, stationary phase, RpoS, visible light, seawater.

ISSN:0008-4166    

DOI:10.1139/m97-148

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

The facultative intracellular pathogenAeromonas salmonicidasurvives and replicates in macrophages, a virulence trait presumed to be associated with its ability to resist reactive radicals. The mechanisms used byA.salmonicidato resist reactive radicals in vitro were shown to have both structural and physiological determinants. The sensitivity ofA.salmonicidato exogenous H2O2, superoxide, and nitrogen radicals, as well as endogenous oxygen radicals, differed depending on growth conditions, cell surface structure, and preexposure to sublethal doses of radicals. Whereas sensitivities to exogenous oxygen radicals did not correlate with basal levels of catalase or Fe-superoxide dismutase, under similar culture conditions S-layer positive cells were more resistant to oxygen radicals than S-layer mutants. S-layer mutants recovered resistance when physically reconstituted with S-layer sheets. Hemin-coated S-layers, while protective against nitrogen radicals, sensitizedA.salmonicidato H2O2. Sublethal concentrations of H2O2or superoxide induced a highly protective response characterized by de novo synthesis of both catalase and Mn-superoxide dismutase. It is proposed that forA.salmonicidathe constitutive S-layer provides a first line of defense and the inducible catalase and Mn-superoxide dismutase provide a powerful second line of defense against macrophage-mediated killing via reactive oxygen species.Key words:Aeromonas salmonicida, oxygen radicals, nitrogen radicals, oxidative stress, S-layers.

ISSN:0008-4166    

DOI:10.1139/m97-149

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

The major (26 kDa) autolysin fromPseudomonas aeruginosawas purified to apparent homogeneity by a combination of preparative electrophoresis, ion-exchange, and dye–ligand chromatographies. This purification was facilitated by the development of a spot-assay that involved the spotting and subsequent incubation of autolysin samples on polyacrylamide gels containing peptidoglycan. The pI of the 26-kDa autolysin was determined to be between 3.5 and 4 and disulfide bonds within the enzyme were essential for activity. The autolysin catalyzed the release of reducing sugars from the peptidoglycans ofPseudomonas aeruginosaandEscherichia coliindicating it to be a β-glycosidase. It was ineffective at hydrolysing the peptidoglycan from Gram-positive bacteria and theO-acetylated peptidoglycans from eitherProteus mirabilisorStaphylococcus aureus. The N-terminal sequence of the purified autolysin was determined to be His-Glu-Pro-Pro-Gly. The 26-kDa autolysin together with a 29-kDa autolysin was determined to be secreted into the medium by a mechanism that involves the production and release of surface membrane vesicles during normal growth, but the enzymes were not found free and active in culture broth supernatants.Key words: autolysin, purification,Pseudomonas aeruginosa, membrane vesicles, muramidase.

ISSN:0008-4166    

DOI:10.1139/m97-150

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

RandomSau3AI DNA fragments from the temperateLactobacillusbacteriophage A2 were cloned into the promoter-probe plasmid pGKV210. Seven DNA fragments with promoter activity were selected, after transformation ofEscherichia coliandLactococcus lactis, subsp.lactis, through the chloramphenicol resistance they conferred to the corresponding clones. The seven promoters were functional inLactobacillus casei. Their strength was analysed by measuring the levels of chloramphenicol resistance and chloramphenicol acetyltransferase activity induced in each host. The nucleotide sequences of these fragments were determined and primer extension analysis was used to locate the initiation site of transcription from each promoter inE.coli. The promoters contained −10 and −35 regions similar to the consensus sequences ofE.coliandLactobacilluspromoters.Key words: bacteriophage,Lactobacillus, promoter.

ISSN:0008-4166    

DOI:10.1139/m97-151

出版商:NRC Research Press    

年代:1997

数据来源: NRC

摘要:

Eight bacteriophages specific toSerratia entomophila, a commercially available bacterial pathogen of the New Zealand grass grub (Costelytra zealandica), were characterized by host range determination, morphology and restriction endonuclease patterns of DNA. Phages were originally isolated from grass grub larvae and fermenter broth where phages had disrupted large-scale production ofS.entomophila. Seven of the phages (CW1–CW5, BC, and BT) had heads similar in size (approximately 60 × 60 nm) and long noncontractile tails (185 × 10 nm). Phage AgRP8 (P8) had a smaller head and a short tail structure. Restriction endonuclease analysis divided the phages into four groups: CW2, CW4, CW5, BC, and BT gave identical patterns, while CW1, CW3, and P8 each gave different patterns. Six distinct phage groups were distinguished by host range determination, after screening phages against 70 bacterial isolates: CW1, CW2/CW4, CW3, CW5, BC/BT, and P8. While confirming the indicated groupings by DNA analysis, it was possible to distinguish between some of the phages in the largest group: CW2/4 could be distinguished from CW5 and BC/BT. Screening of soil bacterial isolates ofS.entomophilaagainst nondiluted phages will aid in monitoring the establishment and persistence of strains applied for biological control of the grass grub.Key words:Serratia entomophila, bacteriophage, morphology, phage typing, host range.

ISSN:0008-4166    

DOI:10.1139/m97-152

出版商:NRC Research Press    

年代:1997

数据来源: NRC