摘要:

This is the first study on the characterization of lactococcal phages isolated in Canada. Thirty lactococcal phages were isolated from Quebec cheese plants reporting partial loss of starter activity. Phages were characterized by electron microscopy, DNA homology, protein profile, and host range. All phages belonged to theSiphoviridaefamily. Seventeen phages (57%) has prolate heads (60 × 40 nm) and 100 nm long, noncontractile tails (morphotype B2, species c2). They showed strong DNA homology with other prolate-headed phages isolated from other countries (Australia, United States). In addition to normal prolate phages, most lysates contained pairs of empty heads (no DNA) connected by a small bridge. Thirteen phages (43%) had small isometric heads (55 nm in diameter) and long, noncontractile tails (morphotype B1). Based on DNA homology, 11 of these phages were found related to phage species 936 despite differences in tail length (140 to 200 nm). The two other small isometric phages, UL36 and UL39, hybridized with phage P335 DNA, and therefore belong to this species. No DNA homology was observed between prolate and small isometric phages. Phages with prolate heads showed a broader host range than small isometric-headed phages. The DNA of phage UL36, which has a relatively narrow host range, has more restriction endonuclease sites than other lactococcal bacteriophages.Key words: lactococci, bacteriophage, taxonomy, cheese, whey.

ISSN:0008-4166    

DOI:10.1139/m92-143

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

The prototrophicPseudomonas syringaepv.tomatomutant DC3481, which is the result of a single-site Tn5insertion, cannot grow and cause disease on tomato plants and cannot use the major organic acids of tomato, i.e., citric, malic, succinic, and tartaric acids, as sole carbon sources. Although nonpathogenic, strain DC3481 can still induce a hypersensitive reaction in nonhost plants. We have identified a 30-kb fragment ofP.syringaepv.tomatowild-type DNA that can complement this mutant.EcoRI fragments from this region were subcloned and individually subjected to functional complementation analysis. The 3.8-kb fragment, which was the site of the Tn5insertion, restored pathogenicity and the ability to use all the major organic acids of tomato as carbon sources. It shares sequence homology with severalP.syringaepathovars but not other bacterial tomato pathogens. Our results indicate that sequences on the 3.8-kbEcoRI fragment are required for both the ability to grow on tomato leaves (and thus cause disease) and the utilization of carboxylic acids common to tomato. The 3.8-kb fragment may contain a sequence (or sequences) that regulates both traits.Key words:Pseudomonas syringaepv.tomato, phytopathogenicity, Tn5, tricarboxylic acid metabolism, bacterial speck, growthin planta.

ISSN:0008-4166    

DOI:10.1139/m92-144

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

Chitinase (EC 3.2.1.14) was isolated from the culture supernatant of a marine bacterium,Alteromonassp. strain O-7. The enzyme (Chi-A) was purified by anion-exchange chromatography (DEAE-Toyopearl 650 M) and gel filtration (Sephadex G-100). The purified enzyme showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The molecular size and pI of Chi-A were 70 kDa and 3.9, respectively. The optimum pH and temperature of Chi-A were 8.0 and 50 °C, respectively. Chi-A was stable in the range of pH 5–10 up to 40 °C. Among the main cations, such as Na+, K+, Mg2+, and Ca2+, contained in seawater, Mg2+stimulated Chi-A activity.N-Bromosuccinimide and 2-hydroxy-5-nitrobenzyl bromide inhibited Chi-A activity. The amino-terminal 27 amino acid residues of Chi-A were sequenced. This enzyme showed sequence homology with chitinases from terrestrial bacteria such asSerratia marcescensQMB1466 andBacillus circulonsWL-12.Key words: marine bacterium,Alteromonassp., chitinase.

ISSN:0008-4166    

DOI:10.1139/m92-145

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

Extracellular wood-degrading enzymes of the brown-rot fungusPostia placentawere localized using colloidal gold labeled monoclonal antibodies to the β-1,4-xylanase (32 to 36 kDa) fraction ofP.placenta.Postia placentawas grown from agar onto glass cover slips, immunolabeled with or without prior fixation, and examined by scanning electron microscopy. Enzymes were localized on the hyphal surface and on the clumped fibrillar elements mycofibrils of the hyphal sheath following fixation with glutaraldehyde. If fixation was omitted, labeling was diffuse and not localized on individual or clumped mycofibrils. We conclude that extracellular decay enzymes are weakly bound (noncovalently) to, but not identical with, the linear mycofibrillar elements of the hyphal sheath. The linear structural elements of the hyphal sheath may play an important role in transport and presentation of wood-degrading enzymes during the decay process.Key words: brown-rot fungi, enzymes, mycofibrils, hyphal sheath, immunolabeling, monoclonal antibodies, colloidal gold, scanning electron microscopy.

ISSN:0008-4166    

DOI:10.1139/m92-146

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

Evidence is provided for the existence of linear extracellular fibrillar elements in the brown-rot fungusPostia placenta. These elements appear as structural components of the hyphal sheath and more closely resemble mycofibrils than fungal fimbriae. Mycofibrils are associated with and appear to originate from the hyphal surface when hyphae are grown on wood or inert substrates, such as glass cover slips and polycarbonate filters. These extracellular structures have a nominal diameter of 10–50 nm and are up to 25 μm in length. We conclude that mycofibrils are linear structural extensions of the hyphal cell wall. The precise function of mycofibrils in the brown-rot decay process of wood remains to be elucidated.Key words:Postia placenta, mycofibrils, fungal fimbriae, hyphal sheath, electron microscopy.

ISSN:0008-4166    

DOI:10.1139/m92-147

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

A skimmed-milk clearing assay was used to identify, in a multicopyStreptomyces lividans66 genomic library, DNA fragments that lead to increased expression of protease activity inS.lividans66. Three independent loci were identified. The majority class (slpA, which represented 68 of 71 clones) produced large zones of clearing. Two other classes (designatedslpBandslpC) showed smaller zones thanslpA. Subcloning and deletion analysis of theslpAlocus delineated the relevant DNA to within a 2.5 kilobase pair fragment. DNA sequence analysis revealed a structural gene associated with the appearance of an extracellular protein in the culture medium. The derived amino acid sequence indicated the presence of a zinc-binding motif, which was previously noted to be characteristic of metalloprotease enzymes. However, the relatively small size of the protein (apparent molecular weight 20 000 – 24 000) suggests that it represents a novel class of neutral proteases distinct from the thermolysin-type enzymes. An adjacent divergent open reading frame was identified and shown to cause a significant increase in protease activity when present together with the protease structural gene on a multicopy plasmid inS.lividans66. The derived amino acid sequence of this open reading frame showed homology with previously characterized regulatory proteins of the LysR family of transcriptional regulator proteins.Key words:Streptomyces, extracellular proteases, putative transcriptional regulator.

ISSN:0008-4166    

DOI:10.1139/m92-148

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

To evaluate immobilized bacteria technology for the removal of low levels of glyphosate (N-phosphonomethylglycine) from aqueous industrial effluents, microorganisms with glyphosate-degrading activity obtained from a fill and draw enrichment reactor inoculated with activated sludge were first exposed to glyphosate production wastes containing 500–2000 mg glyphosate/L. The microorganisms were then immobilized by adsorption onto a diatomaceous earth biocarrier contained in upflow Plexiglas® columns. The columns were aerated, maintained at pH 7.0–8.0, incubated at 25 °C, supplemented with NH4NO3(50 mg/L), and exposed to glyphosate process wastes pumped upflow through the biocarrier. Glyphosate degradation to aminomethylphosphonic acid was initially >96% for 21 days of operation at flows yielding hydraulic residence times (HRTs) as short as 42 min. Higher flow rate studies showed >98% removal of 50 mg glyphosate/L from the waste stream could be achieved at a HRT of 23 min. Glyphosate removal of >99% at a 37-min HRT was achieved under similar conditions with a column inoculated with a pure culture ofPseudomonassp. strain LBr, a bacterium known to have high glyphosate-degrading activity. After acid shocking (pH 2.8 for 18 h) of a column of immobilized bacteria, glyphosate-degrading activity was regained within 4 days without reinoculation. Although microbial growth and glyphosate degradation were not maintained under low organic nutrient conditions in the laboratory, the low levels of degradable carbon (45–94 mg/L) in the industrial effluent were sufficient to support prolonged glyphosate-degrading activity. The results demonstrated that immobilized bacteria technology is effective in removing low levels of glyphosate in high-volume liquid waste streams.Key words: glyphosate, degradation, immobilized bacteria technology.

ISSN:0008-4166    

DOI:10.1139/m92-149

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

Five majoranfH-hybridizing mRNA species accumulated inAzotobacter vinelandiicells derepressed for nitrogenase-3 (an alternative nitrogenase, which appears to lack Mo and V). UsinganfH-,anfD-,anfG-,anfK-, andorf1orf2-specific probes and mutant strains ofA.vinelandiithese mRNA species have been identified as encodinganfHDGKorf1orf2(6.0 kb),anfHDGK(4.3 kb),anfHD(2.6 kb),vnfHorfFd(1.3 kb), andvnfHand (or)anfH(1.0 kb). A 0.6-kb mRNA species, which hybridized only to theorf1orf2-specific probe, and a 3.5-kb mRNA species, which hybridized toanfDoranfK, also accumulated under these conditions. Northern blot analysis and S1 nuclease mapping indicate that transcription of theanfstructural gene cluster initiates at a uniquenifconsensus promoter situated 127 base pairs upstream from theanfHcoding region. Observation ofanfH-hybridizing mRNA species that accumulate in strains derepressed for nitrogen fixation demonstrates that transcription of theanfHDGKorf1orf2cluster is normally repressed by Mo, V, and NH4+, whereas transcription of thevnfHorfFdcluster does not require the presence of V and is repressed only by Mo, but not NH4+. Analysis of the accumulation of mRNAs in a tungsten-tolerant strain revealed that Mo and V repression ofanftranscription must occur by different mechanisms.Key words:Azotobacter vinelandii, nitrogenase-3, transcripts, regulation, molybdenum, vanadium.

ISSN:0008-4166    

DOI:10.1139/m92-150

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

Staphylococcal toxic shock syndrome toxin-1 (TSST-1) as well as staphylococcal enterotoxin A (SEA) and B (SEB) have recently been shown to bind directly to the class II major histocompatibility antigen, HLA-DR. Whereas others have characterized TSST-1 and SEA binding to HLA-DR on transfected L cells or B lymphoma cell lines, we sought evidence for direct binding of TSST-1 and SEA to HLA-DR on purified human monocytes. A single class of high-affinity receptors was found for both TSST-1 (dissociation constant (Kd) 40 nM, 3.4 × 104receptors per cell) and SEA (Kd12 nM, 3.2 × 104receptors per cell) on normal human monocytes. Affinity cross-linking of125I-labeled toxins to monocytes revealed the presence of two membrane protein subunits with molecular masses consistent with the α and β chains of human HLA-DR (35 and 28 kDa, respectively). The anti-HLA-DR monoclonal antibody L243, but not L203 or 2.06, inhibited radiolabeled toxin binding to human monocytes and neutralized the mitogenic response of human T lymphocytes to both toxins. However, L243 was consistently more effective in blocking radiolabeled TSST-1 than SEA binding to human monocytes from the same donors, suggesting that TSST-1 and SEA may be binding to overlapping epitopes rather than to the same epitope on HLA-DR. Because TSST-1 and SEB bind to distinct epitopes on HLA-DR and because SEA cross competes with both TSST-1 and SEB on the HLA-DR receptor, we postulate that SEA occupies a binding site within HLA-DR that overlaps both TSST-1 and SEB. Future studies focused on receptor-mediated binding of these toxins to human monocytes and T lymphocytes from normal donors and toxic shock syndrome patients may reveal the underlying anomalies that predispose particular individuals to toxic shock syndrome.Key words: monocytes, staphylococcal toxic shock syndrome toxin-1, receptors, HLA-DR, staphylococcal enterotoxin A.

ISSN:0008-4166    

DOI:10.1139/m92-151

出版商:NRC Research Press    

年代:1992

数据来源: NRC

摘要:

The unique conditions associated with discontinuous (batch) dialysis culture of the diatomPhaeodactylum tricornutumelicit different nutritional responses from those observed in nondialysis culture. Simultaneous determinations of the nitrogen biomass concentration and of the nitrogen nutrients (NO3−+ NO2−) in the culture chamber, as well as in the seawater nutrient medium at the entrance and exit of the dialyzer, revealed that nitrogen-biomass production in dialysis cultures is achieved mainly (>90%) during postexponential growth, when the concentration of nutrients is limiting (smaller than Michaelis-Menten constant). Almost half of this biomass is produced at the diffusion limit of the apparatus, i.e., when the mass transfer of nutrient substrates, which determines the total uptake activity of the culture, reaches a maximum. In contrast, in nondialysis discontinuous cultures, in which the postexponential growth phase is short, most of the total cellular nitrogen is accumulated during active growth. Certain physiological indices relating to the active uptake and assimilation of inorganic nitrogen are consistent with the different nutritional responses elicited by these two types of culture techniques and explain the high biomass levels obtained in dialysis culture.Key words: dialysis culture, diatom, nitrogen metabolism.

ISSN:0008-4166    

DOI:10.1139/m92-152

出版商:NRC Research Press    

年代:1992

数据来源: NRC