摘要:

A naphthalene (Nap) and salicylate (Sal) degrading microorganism,Pseudomonas putidaRKJ1, is chemotactic towards these compounds. This strain carries a 83 kb plasmid. A 25 kbEcoRI fragment of the plasmid contains the genes responsible for Nap degradation through Sal. RKJ5, the plasmid-cured derivative of RKJ1, is neither capable of degradation nor is chemotactic towards Nap or Sal. The recombinant plasmid pRKJ3, which contained a 25 kbEcoRI fragment, was transferred back into the plasmid-free wild-type strain RKJ5, and the transconjugant showed both degradation and chemotaxis. The recombinant plasmid pRKJ3 was also transferred into motile, plasmid-freeP. putidaKT2442. The resulting transconjugant (RKJ15) showed chemotaxis towards both Nap and Sal. Two mutant strains carrying deletions in pRKJ3 (in KT2442) with phenotypes Nap-Sal+and Nap-Sal-, were also tested for chemotaxis. It was found that the Nap-Sal+mutant strain showed chemotaxis towards Sal only, whereas the Nap-Sal-mutant strain is non-chemotactic towards both the compounds. These results suggest that the metabolism of Nap and Sal may be required for the chemotactic activity.Key words:Pseudomonas putida, plasmid-encoded chemotaxis, naphthalene, salicylate.

ISSN:0008-4166    

DOI:10.1139/w99-118

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

Evidence accumulated from studies based on physiological, biochemical, and molecular characteristics has pointed to the heterogeneity of the ubiquitous anamorphic basidiomycetous yeast speciesCryptococcus albidus(Saito) Skinner, with its current varieties and synonyms. The taxonomic status of this species has not been reappraised because different studies, mostly involving limited numbers of strains, have not been integrated. To assess species diversity within the clade containingCryptococcus albidusand other phylogenetically relatedCryptococcusandFilobasidiumspecies, we determined ribosomal DNA (rDNA) sequences of 69 strains from the 5prime end of the 26S gene, D1/D2 region, and in some cases, the non-coding ITS2 region. Analysis of the sequence data together with available physiological, biochemical, and molecular characteristics, showed the segregation ofC. albidusinto at least 12 species, leading to the elevation of former varieties to the rank of species (C. aerius,C. diffluens), the reinstatement of synonyms (C. liquefaciens,C. terricola), and the proposal of new species (C. arrabidensis,C. chernovii,C. cylindricus,C. oeirensis,C. phenolicus,C. saitoi,C. uzbekistanensis,C. wieringae). The overall analyses of the results argue in favour of the use of rDNA sequence data to improve species delineation when integrated with other available physiological and molecular characteristics.Key words: basidiomycetous yeasts, taxonomy,Cryptococcus albidus, ribosomal DNA, molecular systematics.

ISSN:0008-4166    

DOI:10.1139/w99-108

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

The relative cell surface hydrophobicity (CSH) of 18 soil isolates ofPseudomonas fluorescens, determined by phase exclusion, hydrophobic interaction chromatography (HIC), electrostatic interaction chromatography (ESIC), and contact angle, revealed large degrees of variability. Variation in the adhesion efficiency toMacrophomina phaseolinaof the hyphae/sclerotia of these isolates was also examined. Two such isolates with maximum (32.8%; isolate 12-94) and minimum (12%; isolate 30-94) CSH were selected for further study. Early- to mid-log exponential cells of these isolates were more hydrophobic than those in stationary phase, and the CSH of these isolates was also influenced by fluctuations in temperatures and pH. Isolate 12-94 exhibited high CSH (32.3%) at 30°C, compared to lower values (28-24%) in the higher temperature range (35-40°C). Increasing concentrations of either Zn2+, Fe3+, K+, and Mg2+in the growth medium were associated with the increased CSH. Trypsin, pepsin, and proteinase K (75 to 150 &mgr;g·mL-1) reduced the CSH of isolate 12-94 cells. CSH was reduced, following exposure to DTT, SDS, Triton X-100, or Tween 80. Prolonged exposure of cells to starvation (60 days) also caused a significant decline in CSH. Several protein bands (18, 21, 23, 26 kDa) of the outer cell membrane were absent in 60-day starved cells compared to unstarved cells. In conclusion, our findings demonstrate that CSH ofP. fluorescensisolates may contribute to non-specific attachment/adhesion ontoM. phaseolinahyphae/sclerotia, and the efficiency of adhesion is regulated by growth and other environmental conditions.Key words: adhesion, hydrophobicity,Pseudomonas fluorescens,Macrophomina phaseoli

ISSN:0008-4166    

DOI:10.1139/w99-104

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

Sixty-four species of filamentous fungi from five flare pits in northern and western Canada were tested for their ability to degrade crude oil using gas chromatographic analysis of residual hydrocarbons following incubation. Nine isolates were tested further using radiorespirometry to determine the extent of mineralization of model radiolabelled aliphatic and aromatic hydrocarbons dissolved in crude oil. Hydrocarbon biodegradation capability was observed in species representing six orders of the Ascomycota. Gas chromatography indicated that species capable of hydrocarbon degradation attacked compounds within the aliphatic fraction of crude oil,n-C12-n-C26; degradation of compounds within the aromatic fraction was not observed. Radiorespirometry, usingn-[1-14C]hexadecane and [9-14C]phenanthrene, confirmed the gas chromatographic results and verified that aliphatic compounds were being mineralized, not simply transformed to intermediate metabolites. This study shows that filamentous fungi may play an integral role in the in situ biodegradation of aliphatic pollutants in flare pit soils.Key words: bioremediation, filamentous fungi, flare pits, hydrocarbon degradation, petroleum.

ISSN:0008-4166    

DOI:10.1139/w99-117

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

A new species ofPichiaand two new species ofCandidaare described and were determined to be genetically isolated from all other currently recognized ascomycetous yeasts from their sequence divergence in the species-variable D1/D2 domain of large subunit (26S) ribosomal DNA. The three species were primarily isolated from the frass of wood-boring insects living in pine and spruce trees. The new species and their type strains are the following:Pichia ramenticolaNRRL YB-1985 (CBS 8699), mating type alpha (NRRL YB-3835, CBS 8700, mating typea),Candida piceaeNRRL YB-2107 (CBS 8701), andCandida wyomingensisNRRL YB-2152 (CBS 8703).Pichia ramenticolaandC. piceaeassimilate methanol as a carbon source;P. ramenticolais the first known heterothallic ascomycetous yeast to utilize this compound.Key words:Pichia,Candida, new yeast species, ribosomal DNA, molecular systematics, methanol utilization.

ISSN:0008-4166    

DOI:10.1139/w99-115

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

TheappAgene that was previously shown to code for an acid phosphatase instead codes for a bifunctional enzyme exhibiting both acid phosphatase and phytase activities. The purified enzyme with a molecular mass of 44 708 Da was further separated by chromatofocusing into two isoforms of identical size with isoelectric points of 6.5 and 6.3. The isoforms had identical pH optima of 4.5 and were stable at pH values from 2 to 10. The temperature optimum for both phytase isoforms was 60°C. When heated at different pH values the enzyme showed the greatest thermal resistance at pH 3. The pH 6.5 isoform exhibitedKmandVmaxvalues of 0.79 mM and 3165 U·mg-1of protein for phytase activity and 5.5 mM and 712 U·mg-1of protein for acid phosphatase, respectively. The pH 6.3 isoform exhibited slightly lowerKmandVmaxvalues. The enzyme exhibited similar properties to the phytase purified by Greiner et al. (1993), except the specific activity of the enzyme was at least 3.5-fold less than that previously reported, and the N-terminal amino acid sequence was different. The Bradford assay, which was used by Greiner et al. (1993) for determination of enzyme concentration was, in our hands, underestimating protein concentration by a factor of 14. Phytase production using the T7 polymerase expression system was enhanced by selection of a mutant able to grow in a chemically defined medium with lactose as the carbon source and inducer. Using this strain in fed-batch fermentation, phytase production was increased to over 600 U·mL-1. The properties of the phytase including the low pH optimum, protease resistance, and high activity, demonstrates that the enzyme is a good candidate for industrial production as a feed enzyme

ISSN:0008-4166    

DOI:10.1139/w99-084

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

Kinetic models for microbial growth describe the specific growth rate (&mgr;) as a function of the concentration of the growth-limiting nutrient (s) and a set of parameters. A typical example is the model proposed by Monod, where &mgr; is related tosusing substrate affinity (Ks) and the maximum specific growth rate (&mgr;max). The preferred method to determine such parameters is to grow microorganisms in continuous culture and to measure the concentration of the growth-limiting substrate as a function of the dilution rate. However, owing to the lack of analytical methods to quantify sugars in the microgram per litre range, it has not been possible to investigate the growth kinetics ofEscherichia coliin chemostat culture. Using an HPLC method able to determine steady-state concentrations of reducing sugars, we previously have shown that the Monod model adequately describes glucose-limited growth ofE. coliML30. This has not been confirmed for any other sugar. Therefore, we carried out a similar study with galactose and found steady-state concentrations between 18 and 840 &mgr;g·L-1for dilution rates between 0.2 and 0.8·h-1, respectively. With these data the parameters of several models giving the specific growth rate as a function of the substrate concentration were estimated by nonlinear parameter estimation, and subsequently, the models were evaluated statistically. From all equations tested, the Monod model described the data best. The parameters for galactose utilisation were &mgr;max= 0.75·h-1andKs= 67 &mgr;g·L-1. The results indicated that accurateKsvalues can be estimated from a limited set of steady-state data when employing &mgr;maxmeasured during balanced growth in batch culture. This simplified procedure was applied for maltose, ribose, and fructose. For growth ofE. coliwith these sugars, &mgr;maxandKswere for maltose 0.87·h-1, 100 &mgr;g·L-1; for ribose 0.57·h-1, 132 &mgr;g·L-1, and for fructose 0.70·h-1, 125 &mgr;g·L-1.Key words: monod model, continuous culture, galactose, glucose, fructose, maltose, ribose.

ISSN:0008-4166    

DOI:10.1139/w99-113

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

Bradyrhizobium japonicummutant strain NAD163, containing a 30-kb deletion mutant encompassing thehsfAgene, was inoculated onto a broad range of legume species to test host-specificity. Most legume species formed ineffective nodules exceptVigna angularisvar. Chibopat andGlycine maxvar. Pureunkong. AhsfAinsertion mutant, BjjC211, gave similar results to strain NAD163, implying that many legume species require HsfA for host-specific nitrogen fixation. To determine whether other genes in the deleted region of NAD163 are also necessary, thehsfAgene was conjugally transferred into the NAD163 mutant. The transconjugant formed effective nodules on the host legume plants, which earlier had formed ineffective nodules with mutant NAD163. Thus, we conclude that thehsfAgene in the 30-kb region is the only factor responsible for host-specific nitrogen fixation in legume plants.Key words: host-specific nitrogen fixation, legume-Rhizobiumsymbiosis,hsfAgene, host-specificity.

ISSN:0008-4166    

DOI:10.1139/w99-111

出版商:NRC Research Press    

年代:1999

数据来源: NRC

摘要:

Minke whales consume large amounts of pelagic crustaceans. Digestion of the prey is initiated by indigenous bacteria in a rumen-like forestomach system. A major structural component of the crustacean exoskeleton is chitin, the &bgr;-1,4-linked polymer ofN-acetyl-D-glucosamine. The exoskeletons appear to dissolve completely in the non-glandular forestomach. Bacteria in the forestomach fluid of six krill-eating minke whales were enumerated and isolated using an anaerobic habitat-simulating culture medium. Median viable population densities ranged between 6.0 × 106and 9.9 × 109bacterial cells per mL forestomach fluid. Bacterial isolates (n= 44) cultured from the forestomach fluid of one minke whale mainly resembled strains ofEubacterium(25%),Streptococcus(18%),Clostridium(14%), andBacteroides(11%). As much as 12% of the bacterial isolates were chitinolytic, while &bgr;-N-acetylglucosaminidase activity was demonstrated in 54% of the isolates, and utilisation ofN-acetyl-D-glucosamine was observed in 73%. The chitinolytic isolates resembled strains ofBacteroides,Bacteroidaceae,Clostridium, andStreptococcus. Scanning and transmission electron microscopy of partly digested krill from the minke whale forestomach revealed bacteria close to and inside the chitinous exoskeleton. The bacterial chitinase may act on the chitinous crustacean exoskeletons, thereby allowing other bacteria access to the nutritious soft inner tissues of the prey, and thus initiating its degradation and fermentation.Key words: chitinase, &bgr;-N-acetylglucosaminidase, cetacea, symbiotic microbial digestio

ISSN:0008-4166    

DOI:10.1139/w99-112

出版商:NRC Research Press    

年代:1999

数据来源: NRC