Fungal and bacterial biomass in tundra soils along an arctic transect from Taimyr Peninsula, central Siberia

Microscopic analyses of tundra soils from northern central Siberia, Taimyr Peninsula (74.5°N, 98.5°E) were performed in order to investigate spatial variation of fungal and bacterial biomass. Biomass figures of fungi and bacteria (µg C g^-1 dry wt.) were measured from 11 permafrost soil pits. Fungal biovolume of up to 3.5 mm³ g^-1 dry wt. (median 0.19 mm³ g^-1 dry wt.) and a maximum hyphal length of 393 m g^-1 dry wt. (median 21 m g^-1 dry wt.) were determined. Fungal biomass was found up to 455 µg C g^-1 dry wt. (median 24 µg C g^-1 dry wt.). The amounts generally decreased with depth but increased within organic horizons. Little fungal biomass was found in the unvegetated soils or deep horizons above the permafrost table. Bacterial counts ranged from 0.16 to 7.38*10⁹ g^-1 dry wt. and bacterial biomass ranged from 0.68 to 20.38 µg C g^-1 dry wt. (median 6.19 µg C g^-1 dry wt.) because of small cell volume (median 0.04 µm³). Microbial biomass was generally dominated by fungi as shown by the ratio of fungal to bacterial biomass, which was between 0 and 174.1 (median 4.5). Plant cover and soil organic matter content were found to be the important keys in understanding microbial ecology in arctic tundra soils.     

Conversion of Biovolume Measurements of Soil Organisms, Grown Under Various Moisture Tensions, to Biomass and Their Nutrient Content

Direct microscopic measurements of biomass in soil require conversion factors for calculation of the mass of microorganisms from the measured volumes. These factors were determined for two bacteria, five fungi, and a yeast isolated from soil. Moisture stress conditions occurring in nature were simulated by growth in two media using shake cultures, on agar plates, and on membranes held at 34, 330, and 1,390 kPa of suction. The observed conversion factors, i.e., the ratio between dry weight and wet volume, generally increased with increasing moisture stress. The ratios for fungi ranged from 0.11 to 0.41 g/cm³ with an average of 0.33 g/cm³. Correction of earlier data assuming 80% water and a wet-weight specific gravity of 1.1 would require a conversion factor of 1.44. The dry-weight specific gravity of bacteria and yeasts ranged from 0.38 to 1.4 g/cm³ with an average of 0.8 g/cm³. These high values can only occur at 10% ash if no more than 50% of the cell is water, and a specific conversion factor to correct past data for bacterial biomass has not yet been suggested. The high conversion factors for bacteria and yeast could not be explained by shrinkage of cells due to heat fixing, but shrinkage during preparation could not be completely discounted. Moisture stress affected the C, N, and P content of the various organisms, with the ash contents increasing with increasing moisture stress. Although further work is necessary to obtain accurate conversion factors between biovolume and biomass, especially for bacteria, this study clearly indicates that existing data on the specific gravity and the water and nutrient content of microorganisms grown in shake cultures cannot be applied when quantifying the soil microbial biomass.     

Separation of B and C Type Virions by Centrifugation in Gentle Density Gradients

Murine type B particles were separated from type C (Rauscher leukemia virus) by means of gentle (low-increment rate) density gradients. The best separation was obtained when the density ranged from 1.13 to 1.20 g/cm³ when sucrose was used and from 1.12 to 1.28 g/cm³ with CsCl. The buoyant densities of the B and C particle bands in sucrose were 1.18 and 1.16 g/cm³, respectively. The CsCl gradient gave a better separation with the B particles banding at a density of 1.20 g/cm³ and with the C particle density little different from its value in sucrose.     

An analysis of the bovine genome by Cs2SO4-Ag density gradient centrifugation

Calf DNA preparations having molecular weights of 5 to 7 × 10⁶ have been fractionated by preparative Cs₂SO₄—Ag⁺ density gradient centrifugation into a number of components. These may be divided into three groups: (1) the main DNA component (1.697 g/cm³; all densities quoted are those determined in CsCl density gradients), the 1.704 and 1.709 g/cm³ components form about 50, 25 and 10% of the genome, respectively; they are characterized by having symmetrical CsCl bands and melting curves, both of which have standard deviations close to those of bacterial DNAs of comparable molecular weight, and by their G + C contents being equal to 39, 48 and 54%, respectively; after heat-denaturation and reannealing, their buoyant densities in CsCl are greater than native DNA by 12, 10 and 3 mg/cm³, respectively. (2) The 1.705, 1.710, 1.714 and 1.723 g/cm³ components represent 4, 1.5, 7 and 1.5% of the DNA, respectively, and exhibit the properties of “satellite” DNAs; their CsCl bands and melting curves have standard deviations lower than those of bacterial DNAs; after heat-denaturation and reannealing, their buoyant densities are identical to native DNA, except for the 1.705 g/cm³ component, which remains heavier by 5 mg/cm³; in alkaline CsCl, only the 1.714 g/cm³ component shows a strand separation. (3) A number of minor components, forming 1% of the DNA, have been recognized, but they have not been investigated in detail; two of them (1.719 and 1.699 g/cm³) might correspond to ribosomal cistrons and mitochondrial DNA, respectively.     

Isolation and preliminary characterization of cryptic satellite DNA in pea

Optimum conditions have been established for isolation of ‘cryptic’ satellite DNA from the genome of pea (Pisum sativum), using gradients of CS₂SO₄ containing silver ions. At an Ag⁺ :DNA-P ratio (R) of 0.1, and at alkaline pH, four fractions are obtained: mainband (buoyant density 1.437 g cm³; 67% of total DNA), satellite I (buoyant density 1.582 g/cm³; 7% of total DNA), satellite II (buoyant density 1.520 g/cm³, 11% of total) and satellite III (buoyant density variable between 1.45 and 1.51 g/cm³; 15% of total). The reiterated DNA content of these four fractions has been investigated by reassociation experiments conducted over a Cot range of 1 × 10^?5 to 2.0. All four fractions contain at least two kinetic components; each fraction, including the mainband, consists at least partly of highly reiterated DNA. Ribosomal RNA hybridizes only to the mainband.     

Chromatographic resolution of nucleic acids extracted from Penicillium chrysogenum

Summary High molecular weight DNA extracted from Penicillium chrysogenum has been fractionated using RPC-5 Analog, into three distinct types designated 1, 2 and 3. Types 1 and 2 have the same buoyant density of 1.710 g/cm³ and together appear to comprise the nuclear DNA. Type 1 is enriched for repeated sequences which are normally observed in restriction digests of P. chrysogenum total DNA. Conversely, type 2 appears to be composed entirely of non-repetitive sequences. Type 3 has been identified as mitochondrial DNA, having a buoyant density of 1.695 g/cm³ and an estimated molecular weight of 31.6×10⁶ Daltons.     

Isolation and characterization of microbodies and symphyomicrobodies with different buoyant densities from the fungus Blastocladiella emersonii.

Microbodies isolated from sporangia of the aquatic fungus $\text{Blastocladiella emersonii}$ have a mean buoyant density of 1.222 g/cm³ after centrifugation through a linear sucrose gradient, and contain catalase, isocitrate lyase and malate synthase. Microbodies fuse to produce one symphyomicrobody per zoospore at the time of sporogenesis. An increase in density accompanies this process. The symphyomicrobody has a mean buoyant density of 1.292 g/cm³ while the spore's single mitochondrion has a buoyant density of 1.219 g/cm³. Statistical data are also provided for both starting levels and purification of symphyomicrobody and mitochondrial enzyme markers.     

Role of lysine in the replication of reovirus: I. Synthesis of complete and empty virions

Lysine is essential for the replication of infectious reovirus. Omission of lysine from the extracellular medium not only permitted the continued synthesis of structural viral proteins and viral double-stranded ribonucleic acid (RNA), but also caused an enhanced formation of viral structures which were separable by isopycnic sedimentation of CsCl into a top band consisting of empty particles with a buoyant density of 1.29 g/cm³ and essentially free of viral RNA, and two lower bands which were difficult to resolve and had an average buoyant density of 1.37 g/cm³. The lower bands contained most of the viral nucleic acid. The above effects were reversed when lysine was restored early after infection. In contrast, a single band with a buoyant density of 1.38 g/cm³ was obtained from lysine-plus infected cells.     

A new sensor,the “filtration probe,” for quantitative characterization of the penicillin fermentation. I. Mycelial morphology and culture activity

In this article the filtration resistance of fungal (Penicillium) mycelia is shown to be quantitatively related to the morphology of the cells. On this basis, it was possible to determine the intracellular volume by simple filtration measurements. Since the dry weight of the cells can also be measured, a morphological variable-the “hyphae density” (dry weight hyphae/volume hyphae)-can be quantified. This variable typically is measured to have values in the range of 0.35–0.20 g/cm³; the lower values represent the final stages of the fermentation. It can be shown that this loss of cell material during fermentation is limited to cytoplasmic components, and that most (70%) of the components which disappear are proteins. Concurrent with and linearly related to this loss of proteins, the culture activity in terms of both cell maintenance energy (respiration activity) and penicillin synthesis also decreased. The hyphae density measured by filtration is therefore a direct measure of the metabolic potential of the organism.     

Isolation and partial characterization of a mitochondrial deoxyribonucleic acid-protein complex from sea urchin embryos

Summary Lysis of mitochondria from sea urchin embryos with Triton X-100 led to a complete conversion of DNA-containing mitochondrial residues into protein-DNA complex with a density higher than 1.22 g/cm³ in sucrose solutions. This complex banded isopycnically in metrizamide gradients at a density of 1..26 g/cm³. Exposure to mixtures of Triton X-100 with Tween 80 resulted in progressively less delipitated and disorganized mitochondria over Tween/Triton weight ratios from 1 to 2, with the retention of the starting buoyant density in sucrose of approximately 1.16 g/cm³ at Tween/Triton ratios above 2.5. The DNA-internal protein complex sedimented with the bulk of the surviving mitochondrial structure under all conditions studied. No free DNA could be detected under any conditions of membrane removal.     

Location of Satellite and Homogeneous DNA Sequences on Human Chromosomes

HUMAN DNA^1,2 contains at least two satellite fractions—satellite I (0.5% of the genome) which bands at a density of 1.687 g/cm³ in neutral CsCl and satellite II (2% of the genome) which bands at 1.693 g/cm³. The main band DNA has an average buoyant density between 1.670 and 1.720 g/cm³ and a light shoulder (constituting 15% of the genome) with a buoyant density of 1.696 g/cm³, referred to as homogeneous mainband. Satellite DNA has been observed in many higher organisms³, usually with an unknown function, notable exceptions being cistrons coding for ribosomal RNA⁴ and the DNA coding for histone messenger RNA⁵. To investigate possible functions of human repetitive DNA we have studied the annealing of complementary RNA fractions to chromosomes using in situ hybridization^6,7. We describe here preliminary observations using human satellite II and homogeneous mainband fractions.     

A laboratory energy budget for the ormer Haliotis tuberculata L.

Each component of the energy budget (ingestion, egestion,somatic growth, reproductive investment, respiration, excretion, and mucus production) was measured for Haliotis tuberculata L. held at 15°C in a 12-h light: 12-h dark regime. Ulva lactuca L. was used as the food source throughout, and the budget was assessed over the whole size range of the animal. Ingestion rates ranged from 1.94 to 997.2 cal·animal⁻¹·day⁻¹ in 0.01 to 50 g dry wt (3.71 × 10⁻³ to 17.3 g dry flesh wt) animals, respectively. The major component of the energy budget was somatic growth (37.5% of I) in a 0.01-g dry wt animal while it was respiration (31.1% of I) in a 50-g dry wt animal. Mucus production formed a large part of the budget (from 23.3% of I in a 0.01-g dry wt animal to 29.l% of I in a 50-g dry wt animal). Scope for growth, I − (E + R + U + M), was calculated as ranging from 24.5% of ingestion in a 50-g dry wt animal to 36.8% in a 0.01-g dry wt animal.Each component was measured independently and allometric relationships with animal dry weight calculated. Exponents ranged from 0.60 (somatic growth) to 1.06 (reproductive investment).The calorific value of food was 3419 cal·g dry wt⁻¹ and for faeces was 2817 cal·g dry wt⁻¹. Absorption as a percentage of ingestion in terms of dry weight ranged from 78% for 95-mm length animals (50 g dry wt) to 81% in 6-mm animals (0.01-g dry wt⁻¹). Gross and net growth efficiencies (K₁ and K₂) were calculated on an energy basis and both were logarithmically related to animal dry weight.     

Particle properties of parsnip yellow fleck virus

Particle preparations of parsnip yellow fleck virus (PYFV) isolates A-421 and P-121, representing the two major serotypes, were made by clarifying leal extracts with ether or butan-1-ol and concentrating the virus particles by precipitation with polyethylene glycol and differential centrifugation. The preparations contained c. 31 nm-diameter particles comprising two sedimenting components. Top component (T) consisted of stain-penetrable protein shells with A₂₆₀/A₂₈₀= 0.8–0.9, sedimentation coefficient (S₂₀) = 56 S (A-421) or 60 S (P-121), and buoyant density = 1.297 g/cm³. Bottom component (B) consisted of nucleoprotein particles, not penetrable by negative stain, with A₂₆₀/A₂₈₀= 1.9, sedimentation coefficient (S⁰_20.w) = 148 S (A-421) or 153 S (P-121), and buoyant density = 1.520 g/cm³ (A-421) or 1.490 g/cm³ (P-121). Yields of B component particles were up to c. 1 mg/100 g leaf tissue (both isolates); yields of T component particles were up to c. 0.6 mg (A-421) or 5.5 mg (P-121) per 100 g leaf tissue. PYFV particles were found to contain a single RNA species (mol. wt c. 3.4 × 10⁶, c. 9800 nucleotides), constituting 40% of the particle weight, and three polypeptide species, of mol. wt (× 10 ³) 30 , 26 and 24 (A-421) or 31 , 26 and 23 (P-121).     

Olive latent ringspot virus, a newly recognised virus infecting olive in Italy

A manually transmissible virus, for which the name olive latent ringspot virus (OLRV) is proposed, was isolated from a symptomless olive tree. The virus was mechanically transmitted to test plants. Purified preparations of OLRV contained three classes of isometric particle, c. 28 nm in diameter, with sedimentation coefficients of 525 (T), 975 (M) and 1325 (B) and containing 0, 30 and 43% nucleic acid respectively. At equilibrium in CsCl gradients, the buoyant densities of T and M components were 1–29 and 1–43 g/cm³ respectively, whereas B component separated into two sub-components with buoyant densities of 1–51 g/cm³ (BJ and 1–52 g/cm³ (B₂). Particle preparations contained two species of single-stranded RNA with mol. wt 1–40 times 10⁶ and 2–65 times 10⁶, both necessary for infectivity. The coat protein of OLRV, dissociated under strong denaturing conditions, separated into four components in polyacrylamide gel electrophoresis. Over 75% of the protein was found in a band with mol. wt 57 600, but all four components were recognised as oligomers of a monomeric form with mol. wt 14 300. OLRV was serologically unrelated to 26 different isometric plant viruses including 17 recognised nepoviruses. Its properties strongly indicate that it is a hitherto undescribed member of the nepovirus group.     

Fractionation of somatic and sperm chromatin during spermatogenesis in fish

A new approach is suggested for studying changes in the interactions of protein with DNA in the cells. Measurements of the buoyant density of chromatin were performed in somatic cells and in cells undergoing meiosis in fish. During the process of spermatogenesis some of the somatic histones on the DNA are replaced by a new class of proteins; consequently, the mature sperm contains a unique type of protein having a low mol. wt and a high proportion of arginine.The chromatin obtained from mature sperm is composed of a single component with a density of 1.48–1.49 g/cm³ as measured by CsCl equilibrium sedimentation. On the other hand, somatic cells contain chromatin with lower densities. Chromatin obtained from erythrocytes contains a single component with a density of 1.41–1.42 g/cm³ while liver chromatin shows two components; a main component with a density of 1.45–1.46 and a more heterogeneous component with a lighter density (1.32–1.35). There is a correlation between the buoyant density of the chromatin, the type of its basic proteins and the level of biosynthetic activity in the cells.     

Changing proportions of DNA components in Euglena gracilis

The ratios of satellite deoxyribonucleic acid components to chromosomal deoxyribonucleic acid in Euglena gracilis Z were measured by analytical density gradient ultracentrifugation. Chloroplast deoxyribonucleic acid with a buoyant density of 1.685 g/cm³ exhibited a constant ratio to chromosomal deoxyribonucleic acid during exponential growth and increased twofold as the culture reached the end of the exponential growth phase. The quantity of a satellite deoxyribonucleic acid with a buoyant density of 1.691 g/cm³ was not sufficient to measure the ratio to chromosomal deoxyribonucleic acid during exponential growth but increased to approximately equal the quantity of chloroplast deoxyribonucleic acid as the culture approached the end of the exponential growth phase. The quantity of a deoxyribonucleic acid component with a buoyant density of 1.700 g/cm³ was not sufficient to measure the ratio to chromosomal deoxyribonucleic acid during exponential growth but represented approximately one-third of the total deoxyribonucleic acid as the culture entered the stationary phase of growth.     

Determination of the Biomasses of Small Bacteria at Low Concentrations in a Mixture of Species with Forward Light Scatter Measurements by Flow Cytometry

The forward light scatter intensity of bacteria analyzed by flow cytometry varied with their dry mass, in accordance with theory. A standard curve was formulated with Rayleigh-Gans theory to accommodate cell shape and alignment. It was calibrated with an extinction-culture isolate of the small marine organism Cycloclasticus oligotrophus, for which dry weight was determined by CHN analysis and ¹⁴C-acetate incorporation. Increased light scatter intensity due to formaldehyde accumulation in preserved cells was included in the standard curve. When differences in the refractive indices of culture media and interspecies differences in the effects of preservation were taken into account, there was agreement between cell mass obtained by flow cytometry for various bacterial species and cell mass computed from Coulter Counter volume and buoyant density. This agreement validated the standard curve and supported the assumption that cells were aligned in the flow stream. Several subpopulations were resolved in a mixture of three species analyzed according to forward light scatter and DNA-bound DAPI (4′,6-diamidino-2-phenylindole) fluorescence intensity. The total biomass of the mixture was 340 μg/liter. The lowest value for mean dry mass, 0.027 ± 0.008 pg/cell, was for the subpopulation of C. oligotrophus containing cells with a single chromosome. Calculations from measurements of dry mass, Coulter Counter volume, and buoyant density revealed that the dry weight of the isolate was 14 to 18% of its wet weight, compared to 30% for Escherichia coli. The method is suitable for cells with 0.005 to about 1.2 pg of dry weight at concentrations of as low as 10³ cells/ml and offers a unique capability for determining biomass distributions in mixed bacterial populations.     

Bacterial dry matter content and biomass estimations

Approximately 20% dry-matter content appears to be an accepted standard value for bacterial cells. We have found that the dry-matter content of bacteria may be more than twice as high as generally assumed. The main reason for the low estimates seems to be that proper corrections for intercellular water have not been made when estimating the wet weight of the cells. Using three different bacterial strains, we determined a dry-matter content of cells ranging from 31 to 57%, suggesting not only that the accepted standard value is much too low but also that it is far from standard. To convert bacterial biovolume into biomass (carbon content), we suggest that 0.22 g of C cm-3 should be used as a conversion factor.     

Characterization of deoxyribonucleic Acid species from castor bean endosperm: inability to detect a unique deoxyribonucleic Acid species associated with glyoxysomes

Three DNA buoyant density species (nuclear, 1.692 g cm⁻³; mitochondria 1.705 g cm⁻³; and proplastid, 1.713 g cm⁻³) can be detected in extracts from castor bean endosperm. No other buoyant density species can be identified. DNA extracts from sucrose density gradient purified glyoxysomes exhibit varying amounts of each of the three identified DNAs but no other distinguishable DNA species. RNA synthesized in vitro by Escherichia coli RNA polymerase using purified castor bean nuclear DNA as a template, hybridizes equally well with its template and with the 1.692 g cm⁻³ species from glyoxysome fractions. These results are discussed in terms of their relevance to microbody biogenesis.     

Bacterial dry matter content and biomass estimations.

Approximately 20% dry-matter content appears to be an accepted standard value for bacterial cells. We have found that the dry-matter content of bacteria may be more than twice as high as generally assumed. The main reason for the low estimates seems to be that proper corrections for intercellular water have not been made when estimating the wet weight of the cells. Using three different bacterial strains, we determined a dry-matter content of cells ranging from 31 to 57%, suggesting not only that the accepted standard value is much too low but also that it is far from standard. To convert bacterial biovolume into biomass (carbon content), we suggest that 0.22 g of C cm-3 should be used as a conversion factor.