ISSN:0008-4166    

DOI:10.1139/m93-153

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

Listeria monocytogenes4b and its forms without cell walls (L forms of a protoplastic type) were used to study in vivo interactions with host cells. Samples of peritoneal lavage fluid were obtained from rats intraperitoneally inoculated at intervals between 1 and 15 days after challenge, for scanning electron microscopic, bacteriological, biochemical, and cytometrical investigations. Scanning electron microscopic examination revealed continuous adhesion of L forms on the macrophage surface up to 15 days after inoculation. The persistence of the L forms within the peritoneal cavity was also shown bacteriologically at all sample times, while the parental bacterial forms were isolated from the peritoneal cavity up to 7 days after challenge. The total count of peritoneal exudative cells determined by automated flow peroxidase cytometry peaked on the 15th day in animals infected with parental forms, while in animals infected with L forms the peak was lower and the macrophage population was predominant. The glycolytic and acid phosphatase activity of peritoneal exudative cells was two times higher in rats infected with L forms as compared with rats infected with theL.monocytogenesparental forms on the 3rd day after challenge. An understanding of the nature of the interactions between L forms ofL.monocytogenesand peritoneal exudative cells found in vivo could be used to establish the influence of L forms on host cellular defense mechanisms.Key words:Listeria monocytogenes, L forms, peritoneal exudative cells, electron microscopy.

ISSN:0008-4166    

DOI:10.1139/m93-154

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

The competitiveness ofBradyrhizobium japonicumstrains G49 and SMGS1 was first studied in the greenhouse in sterilized sand, with or without added soil. Strain SMGS1 was more competitive than strain G49 with soybean (Glycine maxL.) cultivar Labrador but the two strains showed equivalent competitiveness with cultivar Kingsoy. When soil was added, nodule occupancy of strain G49 was only 22% with this cultivar. In field experiments, conducted over 2 years in soils already containing strain G49 (1.5 × 103to 4.0 × 104cells/g of soil), nodule occupancy of inoculated strain SMGS1 ranged from 20 to 90%. Nodule occupancy was 3–22% higher when inoculation was done by peat seed coating or with liquid inoculation in the row than with peat-coated clay microgranulars. Nodule occupancy was also dependent on the physiological state of the inoculated cells. When an inoculum stored at 28 °C for 1 year was used at the same viable cell rate, nodule occupancy of strain SMGS1 was 4–20% lower than with a recently made inoculum. Pot experiments with soil from field experiments carried out in the 1st year showed that the inoculated strain continued forming nodules without further inoculation, with a recovery rate equivalent to that of field experiment in the previous year.Key words:Bradyrhizobium japonicum, interstrain competition, inoculation technology, ELISA, field trials.

ISSN:0008-4166    

DOI:10.1139/m93-155

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

In this article we report on the polymerase chain reaction amplification of a partial 16S rRNA gene from the plant pathogenic bacteriumClavibacter michiganensissubsp.sepedonicus. A partial sequence (about 400 base pairs) of the gene was determined that covered two variable regions important for oligonucleotide probe development. A specific 24mer oligonucleotide probe targeted against the V6 region of 16S rRNA was designed. Specificity of the probe was determined using dot blot hybridization. Under stringent conditions (60 °C), the probe hybridized with all 16Cl.michiganensissubsp.sepedonicusstrains tested. Hybridization did not occur with 32 plant pathogenic and saprophytic bacteria used as controls under the same conditions. Under less stringent conditions (55 °C) the relatedClavibacter michiganensissubsp.insidiosus,Clavibacter michiganensissubsp.nebraskensis, andClavibacter michiganensissubsp.tesselariusalso showed hybridization. At even lower stringency (40 °C), allCl.michiganensissubspecies tested includingClavibacter michiganensissubsp.michiganensisshowed hybridization signal, suggesting that under these conditions the probe may be used as a species-specific probe forCl.michiganensis.Key words:Clavibacter michiganensissubsp.sepedonicus, PCR, 16S rRNA, oligonucleotide probe.

ISSN:0008-4166    

DOI:10.1139/m93-156

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

Properties of soil proteases and proteases fromBacillusspp. obtained from water-logged paddy fields treated with organic manure or chemical fertilizer or not treated with fertilizer were compared to elucidate the sources of soil proteases. The major extractable soil proteases were metal chelator sensitive neutral proteases that were active in hydrolyzing benzyloxycarbonyl-L-phenylalanyl-L-leucine and benzyloxycarbonyl-L-phenylalanyl-L-tyrosyl-L-leucine. In this respect they resembled extracellular proteases fromBacillus subtilis(six isolates),Bacillus cereus(four isolates), andBacillus mycoides(three isolates) isolated from the same fields. The major extractable soil protease from the manured field was a serine neutral protease that was active in hydrolyzing casein. It resembled an extracellular protease fromB.subtilis(eight isolates) isolated from the same field. Extractable soil proteases accounted for 18–96% of the total soil protease in the aforementioned soil. We concluded that a major source of soil protease in water-logged paddy fields is proteolyticBacillusspp.Key words: soil protease, metal chelator sensitive neutral protease, serine neutral protease, proteolyticBacillusspp.

ISSN:0008-4166    

DOI:10.1139/m93-157

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

The ability to catabolize crown-gall opines is found inAgrobacterium tumefaciensand various types of nonagrobacteria. Among the 75Rhizobium melilotistrains tested in this work, 6 utilized the opines octopine and octopinic acid as the sole carbon and nitrogen source. From a genomic library of one of these six strains,R.melilotiA3, a clone conferring the octopine catabolism (Occ) phenotype was identified and named pJMA. A different Occ clone, which had been obtained fromR.melilotiRm41, did not hybridize with clone pJMA from strain A3. However, some fragments of clone pJMA hybridized to a 20-kilobaseKpnI fragment containing the Ti plasmid genes of octopine catabolism (occgenes) fromA.tumefaciens15955. Shorter probes carrying the octopine permease genes or part of the octopine oxidase and ornithine cyclodeaminase genes fromA.tumefaciensalso hybridized with pJMA. TheR.melilotiDNA carried by pJMA was localized to a megaplasmid of the wild-type strain A3. Thus, it appears possible that genes represented on the Occ clone from strain A3 share a common origin with the corresponding genes from the Ti plasmid.Key words:Rhizobium meliloti,Agrobacterium tumefaciens, octopine catabolism.

ISSN:0008-4166    

DOI:10.1139/m93-158

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

A model was developed to study the fate of the fish pathogenAeromonas salmonicidain vivo, inside a specialized intraperitoneal chamber implanted in rainbow trout,Oncorhynchus mykiss. Although normally recalcitrant to lytic agents in vitro, owing to the presence of its regular surface array (S layer),A.salmonicidawas rapidly killed in the peritoneal cavity by a host-derived, soluble lytic activity present in peritoneal fluid. Peritoneal fluid was also found to kill other bacteria and lyse various types of erythrocytes, but was particularly lytic toA.salmonicida. Intraperitoneal survival of injected (free)A.salmonicidacells was several orders of magnitude higher than survival of implanted (restrained) cells. Injected free cells could evade the lytic activity of peritoneal fluid because they readily spread, initiating lethal infections. One evasion strategy was envisioned to be the penetration of peritoneal and (or) tissue macrophages. In spite of the killing mechanisms of these phagocytic cells,A.salmonicidawas still able to survive and even replicate inside head kidney macrophages, thereby supporting the notion ofA.salmonicidaas a facultatively intracellular pathogen. Intraperitoneal chambers in rainbow trout may constitute a valuable experimental tool for studying the in vivo fate ofA.salmonicida, and perhaps of other fish pathogens as well.Key words:Aeromonas salmonicida, intraperitoneal chambers, rainbow trout, complement-mediated cell lysis.

ISSN:0008-4166    

DOI:10.1139/m93-159

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

Direct specimen testing was performed on 186 peripheral blood specimens to identify the presence of antigen to cytomegalovirus (viz., the cytomegalovirus antigenemia (CMV-Ag) assay). Confirmatory testing was performed using the shell vial indirect immunofluorescence assay (SVA-IFA), the indirect immunoperoxidase assay (TC-IPA), and conventional tube culture isolation (TC-CPE). The primary reagent for the CMV-Ag assay consisted of anti-CMV monoclonal antibody directed against the internal matrix structural phosphoprotein (1C3; Clonatec-Biosoft, France). The 72-kDa early nuclear antigen (Dupont) was utilized in the SVA-IFA and the TC-IPA. All test systems received an equal number of polymorphonuclear leukocytes in the inoculum. CMV was detected and isolated from 30% (55/186) of the specimens evaluated by either one or a combination of the tests. Detection and (or) isolation of CMV from blood by the CMV-Ag assay, SV-IFA, TC-IPA, and TC-CPE occurred at a rate of 17 (31/186), 12 (22/186), 16 (29/186), and 26% (49/186). Three of 55 positive specimens were identified only by the CMV-Ag assay; each patient in question, however, had at least one previous CMV isolate. No significant differences in sensitivity occurred between the CMV-Ag assay, the SVA-IFA, or the TC-IPA. However, TC-CPE including the blind passage of all negative tube cultures yielded a significantly larger number of positive blood specimens than either of the rapid detection methodologies. The CMV-Ag assay encompasses the benefits of a nonculture system, is simple to perform and easy to read, permits a same-day diagnosis, and requires less reagents than the routinely used SVA-IFA or TC-IPA. The CMV-Ag assay and TC-CPE including a blind passage are recommended assays for the rapid and then long-term identification, respectively, of CMV in the blood of AIDS patients.Key words: cytomegalovirus, antigenemia, blood, AIDS.

ISSN:0008-4166    

DOI:10.1139/m93-160

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

A two-step membrane filter procedure was evaluated to determine the ability to differentiateEscherichia colifrom other coliform bacteria recovered from water. M-Endo LES agar incubated at 35 °C for 24 ± 2 h was used as the initial isolation medium. Membranes containing coliform colonies were transferred to nutrient agar plus 4-methylumbelliferyl β-D-glucuronide (MUG) and incubated for an additional 4 h at 35 °C.Escherichia colicolonies were distinguished by fluorescence when viewed under a long-wavelength ultraviolet light. A total of 119 MUG-positive colonies were isolated from 15 water sources, of which 115 (96.6%) were identified asE.coli. An examination of 182 pure culture environmentalE.coliisolates revealed that 167 isolates (91.8%) exhibited fluorescence on the nutrient agar plus MUG medium. Survivors ofE.colicultures exposed to chlorination were also capable of producing a positive MUG reaction.Key words: membrane filtration, 4-methylumbelliferyl β-D-glucuronide (MUG),Escherichia coli, water.

ISSN:0008-4166    

DOI:10.1139/m93-161

出版商:NRC Research Press    

年代:1993

数据来源: NRC

摘要:

The structure of two rhamnolipid biosurfactants produced byPseudomonas aeruginosaUG2 was studied. Analyses by gas chromatography - mass spectrometry and nuclear magnetic resonance spectroscopy showed these two rhamnolipids to be α-L-rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoate and 2-O-α-L-rhamnopyranosyl-α-L-rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoate. The ability of UG2 rhamnolipid biosurfactants to enhance removal of naphthalene, anthracene, phenanthrene, fluorene, 2,2′,5,5′-tetrachlorobiphenyl, and 3,3′,4,4′,5,5′-hexachlorobiphenyl into the aqueous phase was affected by soil type, hydrocarbon equilibration time, and biosurfactant adsorption to soil. Partially purified UG2 biosurfactants at a concentration of 5 g/L removed approximately 10% more hydrocarbon from a sandy loam soil than silt loam soil. High levels of UG2 rhamnolipids adsorbed to soil. In 18% (w/v) soil slurries 74, 49, 38, and 20% of 0.5, 1, 2, and 5 g UG2 rhamnolipids/L, respectively, were bound to the soil phase. Sodium dodecyl sulphate recovered lower levels and Witconol SN70 higher levels of phenanthrene and 2,2′,5,5′-tetrachlorobiphenyl than UG2 biosurfactants.Key words: biosurfactant, hydrocarbon,Pseudomonas aeruginosa, remediation.

ISSN:0008-4166    

DOI:10.1139/m93-162

出版商:NRC Research Press    

年代:1993

数据来源: NRC