Canadian Journal of Microbiology

Bacteria-free clones of the small centric diatom Cyclotella nana Hustedt were isolated, three from estuarine localities, one from Continental Shelf waters, and one from the Sargasso Sea. Detonula confervacea was isolated from Narragansett Bay. Morphology of all clones was studied with the light and electron microscopes. Morphological differences between clones of C. nana do not at present warrant separating any as distinct species.Clones of C. nana require only vitamin B₁₂; D. confervacea has no vitamin requirement.Growth of the estuarine clones of C. nana was unaffected by salinity down to 0.5‰ and increased with temperature to 25 °C. The Shelf clone grew more rapidly at salinities above 8‰ and at temperatures between 10° and 20 °C. The Sargasso Sea clone did not survive below 15 °C or 17.5‰, while D. confervacea did not survive at temperatures above 15° or at salinities below 8‰. The physiological differences between clones correspond roughly to the conditions obtaining in nature where each was collected.

The relationship of polyketide melanogenesis molecular biology to that of nonmelanin-producing pathways in a wide range of fungi and other organisms is discussed. Analytical methods and fundamental properties of melanins are discussed and fungal melanin properties are compared with those of animal and bacterial melanins. The enzymatic degradation of melanins by lignin peroxidases is described.Key words: fungal melanin, polyketide melanin, DHN melanin, melanin degradation, melanin properties, melanin analysis.

The enzymatic conversion of cellulose is catalyzed by a multiple enzyme system. The Trichoderma enzyme system has been studied extensively and has insufficient β-glucosidase (EC 3.2.1.21) activity for the practical saccharification of cellulose. The black aspergilli (A. niger and A. phoenicis) were superior producers of β-glucosidase and a method for production of this enzyme in liquid culture is presented. When Trichoderma cellulase preparations are supplemented with β-glucosidase from Aspergillus during practical saccharifications, glucose is the predominant product and the rate of saccharification is significantly increased. The stimulatory effect of β-glucosidase appears to be due to the removal of inhibitory levels of cellobiose.

Root colonization by introduced bacteria is an important step in the interaction of beneficial bacteria with the host plant. Investigators attempting to measure root colonization by bacteria must face several issues. A clear concept or definition of root colonization should be stated in each research summary, as several different definitions have been proposed. We consider true root colonists to be those bacteria that colonize roots in competitive conditions, i.e., natural field soils. Different methods of processing root samples are required if one is measuring external root colonization alone, internal colonization alone, or both. Given that most beneficial bacterial strains currently under investigation as root colonists are members of taxa naturally found in soils, a marking system is required to differentiate the introduced strain from members of the indigenous rhizosphere community. Spontaneous antibiotic resistance, immunological approaches, and foreign DNA sequences are among the marking systems that have been used and each has some possible advantages and disadvantages. More research is needed in the development and comparison of marking systems. The design of experiments to measure root colonization should take into account several statistical issues. One must decide what constitutes the sample unit for each replication of a given treatment, e.g., whole root systems or root segments. Consideration should also be given to how best to express the estimated population of root colonists (e.g., cfu/g fresh or dry weight root, cfu/cm root, or cfu/surface area root). Statistical analysis by standard analysis of variance tests should be used whenever possible to separate treatment means of colonization levels; however, one must determine that the underlying assumptions of these tests are correct for each experiment. Finally, in quantification of populations on roots, one will almost certainly encounter replications with no bacteria, i.e., zeros. There are several options for how to calculate treatment means when one or more replications is a zero, and the implications of these options are discussed. Key words: bioluminescence, genetic markers, plant growth-promoting rhizobacteria, rhizosphere bacteria, root, colonization.

Berberine sulfate was shown to possess antimicrobial activity against a wide variety of microorganisms including Gram-positive and Gram-negative bacteria, fungi, and protozoa. The antibacterial activity against Vibrio cholerae and Staphylococcus aureus was dependent on the inoculum size of the test organism and pH of the medium. A method of microbiological assay sensitive to 5–10 μg/ml of the drug was developed. The drug was shown to exert a more rapid antibacterial activity than chloramphenicol and tetracycline on V. cholerae, the K values being 2.4 ×10⁻² sec⁻¹, 7.8 × 10⁻³ sec⁻¹, and 5.2 × 10⁻³ sec⁻¹ respectively. Berberine sulfate was shown to be bacteriocidal to V. cholerae and bacteriostatic to S. aureus, at concentrations of 35 and 50 μg/ml. In both these organisms concentrations of 35 and 50 μg/ml of the drug inhibited ribonucleic acid (RNA) and protein synthesis almost immediately after the addition of the drug. There was little effect on deoxyribonucleic acid (DNA) synthesis at these concentrations.

The formation and activity of nitrogenase² in Azotobacter vinelandii OP was examined using a cell-free assay system. A lag period of about 30 min occurred between the exhaustion of the combined nitrogen source and growth on N₂. Cells grown on ammonium acetate or potassium nitrate had no detectable nitrogenase activity. Nitrogenase activity appeared in cells, grown under a flowing gas phase of 20% O₂ – 60% He, about 45 min after the exhaustion of ammonia. Nitrogenase formation was inhibited in a closed system with an atmosphere containing 40% O₂ but not by one containing 20% O₂. Hydrogen did not inhibit enzyme formation. The question of whether N₂ is required for the formation of the enzyme could not be answered as this gas could not be completely eliminated from the growth system. Chloramphenicol prevented the formation of the enzyme and inhibited nitrogen fixation in whole cells, but had no effect on cell-free enzyme activity. A brief rise in turbidity which occurred during nitrogenase formation appeared to be due to a color change in the cells from reddish brown to dark brown. Spectrophotometric examination of extracts from ammonia- and N₂-grown cells did not reveal any components responsible for this color difference, but this result may reflect only the presence of interfering substances in the crude extract.

A Flavobacterium sp., isolated from paddy water by enrichment culture technique, decomposed diazinon in a mineral medium as sole carbon source. The bacterium readily hydrolyzed diazinon to 2-isopropyl-6-methyl-4-hydroxy-pyrimidine which was then converted to carbon dioxide. The bacterium also converted parathion to p-nitrophenol. The enzyme involved in the hydrolysis was constitutive.

Microbial biomass and soil respiration rate decreased after application of 150 kg NH₄NO₃–N∙ha⁻¹ to different coniferous forest podzols. The decrease was already found 3 months after fertilization and was still evident after 3–5 years. Changes in pH, organic matter, or water content in the soils could not explain the decreases. In laboratory experiments, several unfertilized forest soils were treated with 2 mg of NH₄NO₃–N or of urea–nitrogen∙g wet soil⁻¹. The ammonium nitrate addition resulted in severe depressions of the respiration rates during and up to 175 days of incubation and the decrease was evident after about 1 week. The urea treatment initially increased the respiration rate of the soils, but this appeared to be a transitory effect.

Several media designed for use in a most probable number (MPN) determination of petroleum-degrading microorganisms were compared. The best results, i.e., largest numbers, were obtained using a buffered (32 mM PO₄^≡) liquid medium containing 1% hydrocarbon substrate. Of 104 presumptive oil degraders tested, 20 grew on oil agar medium but did not utilize oil or a mixture of pure paraffinic hydrocarbons (C₁₀ to C₁₆n-alkanes) in liquid (MPN) medium. Visible turbidity in the liquid medium was correlated with hydrocarbon utilization. Counts of petroleum degraders obtained using liquid medium (MPN) were in most cases higher than those obtained on an oil-amended silica gel medium. Both procedures yield an estimation of oil degraders, and the oil-amended agar permits growth of organisms which do not degrade crude oil. All strains of oil-degrading microorganisms examined in this study were lipolytic, but the converse was not always true.

We investigated the effects of two different plant species (corn and soybean) and three different soil types on microbial community structure in the rhizosphere. Our working hypothesis was that the rhizosphere effect would be strongest on fast-growing aerobic heterotrophs, while there would be little or no rhizosphere effect on oligotrophic and other slow-growing microorganisms. Culturable bacteria and fungi had larger population densities in the rhizosphere than in bulk soil. Communities were characterized by soil fatty acid analysis and by substrate utilization assays for bacteria and fungi. Fatty acid analysis revealed a very strong soil effect but little plant effect on the microbial community, indicating that the overall microbial community structure was not affected by the rhizosphere. There was a strong rhizosphere effect detected by the substrate utilization assay for fast-growing aerobic heterotrophic bacterial community structure, with soil controls and rhizosphere samples clearly distinguished from each other. There was a much weaker rhizosphere effect on fungal communities than on bacterial communities as measured by the substrate utilization assays. At this coarse level of community analysis, the rhizosphere microbial community was impacted most by soil effects, and the rhizosphere only affected a small portion of the total bacteria.Key words: rhizosphere, microbial community, fatty acid, substrate utilization.