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Ecological control analysis: being(s) in control of mass flux and metabolite concentrations in anaerobic degradation processes.

Identification of the functional groups of microorganisms that are predominantly in control of fluxes through, and concentrations in, microbial networks would benefit microbial ecology and environmental biotechnology: the properties of those controlling microorganisms could be studied or monitored specifically or their activity could be modulated in attempts to manipulate the behaviour of such networks. Herein we present ecological control analysis (ECA) as a versatile mathematical framework that allows for the quantification of the control of each functional group in a microbial network on its process rates and concentrations of intermediates. In contrast to current views, we show that rates of flow of matter are not always limited by a single functional group; rather flux control can be distributed over several groups. Also, control over intermediate concentrations is always shared. Because of indirect interactions, through other functional groups, the concentration of an intermediate can also be controlled by functional groups not producing or consuming it. Ecological control analysis is illustrated by a case study on the anaerobic degradation of organic matter, using experimental data obtained from the literature. During anaerobic degradation, fermenting microorganisms interact with terminal electron-accepting microorganisms (e.g. halorespirers, methanogens). The analysis indicates that flux control mainly resides with fermenting microorganisms, but can shift to the terminal electron-accepting microorganisms under less favourable redox conditions. Paradoxically, halorespiring microorganisms do not control the rate of perchloroethylene and trichloroethylene degradation even though they catalyse those processes themselves.

Bacteria, Anaerobic↗

Induced plant uptake and transport of mercury in the presence of sulphur-containing ligands and humic acid.

The induced accumulation of mercury (Hg) by plants was investigated for the species Phaseolus vulgaris (Bush bean), Brassica juncea (Indian mustard), and Vicia villosa (Hairy vetch). All plants were grown in modified Hg-contaminated mine tailings and were treated with sulphur-containing ligands to induce Hg accumulation. The effects of varied substrate Hg concentration and humic acid (HA) level on the induced plant-Hg accumulation for B. juncea were examined. Thiosulphate salts (ammonium and sodium) mobilised Hg in the substrates and caused an increase in the Hg concentration of roots and shoots of all tested plant species. Root Hg accumulation was positively correlated to extractable Hg for (NH4)2S2O3-treated B. juncea plants grown in HA-amended substrates. However, shoot Hg translocation for this species was inhibited at 1.25 g HA kg(-1) of substrate. Mercury-thiosulphate complexes could be translocated and accumulated in the upper parts of the plants up to 25 times the Hg concentration in the substrate. We conclude that shoot Hg accumulation in the presence of thiosulphate salts is dependent upon plant species characteristics (e.g. root surface area) and humic acid content.

Biological Transport, Active↗

Real-time polymerase chain reaction quantification of canine DNA.

The accurate quantification of target DNA is an important step in the short tandem repeat analysis of forensic biological samples. By utilizing quantification data to control the amount of template DNA in the polymerase chain reaction (PCR), forensic scientists can optimize testing and minimize the consumption of limited samples. The ability to identify and quantify target DNA in mixed-species samples is crucial when it may be overwhelmed by nontarget DNA, as in cases of dog attack. We evaluated two quantitative real-time PCR assays for dynamic range, species specificity, and inhibition by humic acid. While both assays proved to be highly sensitive and discriminating, the Melanocortin-1 Receptor (MC1R) gene Taqman assay had the advantages of a shorter run time, greater efficiency, and safer reagents. In its application to forensic casework, the MC1R assay has been advantageous for quantifying dog DNA in a variety of mixed-species samples and facilitating the successful profiling of individual dogs.

Animals↗

Strong coupling between natural Planctomycetes and changes in the quality of dissolved organic matter in freshwater samples.

Dilution-regrowth experiments coupled to fluorescence in situ hybridization were conducted with samples from two humic reservoirs in order to examine how inorganic nutrients (N, P) affect free-living bacterioplankton phylogenetic groups and subsequently the quality of dissolved organic matter (DOM). The experiments were complemented by analyses of the empirical relationships between the targeted bacteria, nutrients, DOM and grazers. The ratio of absorbance of waters (A) at 250 and 365 nm (A(250 nm):A(365 nm)), which has been found to increase with the proportion of small molecules in the DOM pool in other humic waters, was used as an index of DOM quality. When nutrient stimulated bacteria, both the responses of bacterial groups (in the absence of grazers) and the ratio A(250 nm):A(365 nm) were generally different between treatments (+N, +P, +NP), suggesting that in nutrient-poor systems, differences in the type of inorganic nutrient supply will ultimately cause differences in DOM quality. The ratio A(250 nm),:A(365 nm) peaked in the +N treatments where members of the Planctomycetes (PLA) were the most stimulated group, and across treatments, PLA best explained (positive relationship) variations in this ratio. Consistent with this, the in situ data showed that the removal of the negative effects of flagellates on PLA yielded the highest R(2) in attempts to use bacterial groups to explain variations in A(250 nm):A(365 nm). These findings provide lines of evidence, not previously demonstrated in natural waters, that Planctomycetes may be an important factor changing the DOM quality, particularly in nutrient-poor systems when supplied with inorganic N.

Animals↗

The Desulfitobacterium genus.

Desulfitobacterium spp. are strictly anaerobic bacteria that were first isolated from environments contaminated by halogenated organic compounds. They are very versatile microorganisms that can use a wide variety of electron acceptors, such as nitrate, sulfite, metals, humic acids, and man-made or naturally occurring halogenated organic compounds. Most of the Desulfitobacterium strains can dehalogenate halogenated organic compounds by mechanisms of reductive dehalogenation, although the substrate spectrum of halogenated organic compounds varies substantially from one strain to another, even with strains belonging to the same species. A number of reductive dehalogenases and their corresponding gene loci have been isolated from these strains. Some of these loci are flanked by transposition sequences, suggesting that they can be transmitted by horizontal transfer via a catabolic transposon. Desulfitobacterium spp. can use H2 as electron donor below the threshold concentration that would allow sulfate reduction and methanogenesis. Furthermore, there is some evidence that syntrophic relationships occur between Desulfitobacterium spp. and sulfate-reducing bacteria, from which the Desulfitobacterium cells acquire their electrons by interspecies hydrogen transfer, and it is believed that this relationship also occurs in a methanogenic consortium. Because of their versatility, desulfitobacteria can be excellent candidates for the development of anaerobic bioremediation processes. The release of the complete genome of Desulfitobacterium hafniense strain Y51 and information from the partial genome sequence of D. hafniense strain DCB-2 will certainly help in predicting how desulfitobacteria interact with their environments and other microorganisms, and the mechanisms of actions related to reductive dehalogenation.

Biodegradation, Environmental↗

Formation of stable chlorinated hydrocarbons in weathering plant material.

Though several chlorinated organic compounds produced by humans are carcinogenic and toxic, some are also produced by the biotic and abiotic processes in the environment. In situ x-ray spectroscopy data indicate that natural organic matter in soils, sediments, and natural waters contain stable, less volatile organic compounds with chlorinated phenolic and aliphatic groups as the principal Cl forms. These compounds are formed at rapid rates from the transformation of inorganic Cl during humification of plant material and, thus, play a critical role in the cycling of Cl and of several major and trace elements in the environment and may influence human health.

Benzopyrans↗

Microheterogeneity of singlet oxygen distributions in irradiated humic acid solutions.

Singlet oxygen (1O2) is a highly reactive species formed through solar irradiation of organic matter in environmental waters. Implicated in a range of reactions, it has proven difficult to quantify its spatial distribution in natural waters. We assessed the microheterogeneous distribution of 1O2 in irradiated solutions containing chromophoric dissolved organic matter (CDOM) by using molecular probes of varying hydrophobicity. The apparent 1O2 concentrations ([1O2]app), measured by recently developed hydrophobic trap-and-trigger chemiluminescent probe molecules, were orders of magnitude higher than those measured by the conventional hydrophilic probe molecule furfuryl alcohol. The differential [1O2]app values measured by these probes reflect a steep concentration gradient between the CDOM macromolecules and the aqueous phase. A detailed kinetic model based on the data predicts probabilistic 1O2 distributions under different solvent conditions.

Humic Substances↗

Radiocarbon: analysis of inorganic carbon of fossil bone and enamel.

Carbon dioxide from calcium carbonate in fossil bone can be selectively separated from carbon dioxide in bone apatite by hydrolyzing the sample first in acetic acid and then in hydrochloric acid. Radiocarbon analyses of the inorganic carbon dioxide in three samples of known age clearly show calcium carbonate in fossil bone to be secondary and the carbonate of bone apatite to be indigenous and suitable for dating in some cases. Agreement between dates on collagen-bone apatite pairs increases the level of confidence.

Acetates↗

Liberation of sulfate from sulfate esters by soils.

When incubated with acid, alkaline, and neutral soils, a variety of synthetic sulfate esters representing the various classes of these compounds was hydrolyzed by enzymes, probably of microbial origin. The appearance of sulfate in the soil water occurred immediately after introduction into the soils with some esters, whereas with others it occurred only after lag periods. Heat treatment destroyed the hydrolytic acitivity in the soils. The ester sulfate groups present in humic acid extracted from the soil appeared to be resistant to hydrolysis by a variety of sulfohydrolases extracted from bacteria and other organisms.

Alcaligenes↗

Cultural and environmental factors affecting the longevity of Escherichia coli in Histosols.

The survival of Escherichia coli in organic soils (Histosols) was examined. The death rate of this organism in Pahokee muck was less than that observed in Pompano fine sand. The number of viable E. coli cells found in the muck was approximately threefold greater than that found in the sand following 8 days of incubation. The initial population of the coliform affected the death rate. The rate of loss of viability varied 100-fold when the population size decreased from 2.5 x 10(7) to 3.4 x 10(4). Other factors affecting the viability of E. coli in muck were aerobic versus anaerobic growth of the organism and moist versus flooded conditions in the soil. The greatest survival of the coliform was noted with anaerobically grown cells amended to flooded soil. That the observed decrease in E. coli viability in soil was the result of biotic factors was demonstrated with amendment of sterile soil with E. coli. When 1.1 x 10(5) bacteria per g of soil were added to sterile muck, a population of 3.0 x 10(7) organisms per g of soil developed over a 10-day period. The role of the protozoa in eradication of the coliform from the muck was indicated by a sixfold increase in the protozoan population in natural soil amended with E. coli. Higher organic matter content in a Histosol compared with a mineral soil resulted in an increased survival of the fecal coliforms. Biotic factors are instrumental in the decline in coliform populations, but the potential for growth of the coliform in the organic soil could extend the survival of the organism.

Aerobiosis↗

Abiotic factors affecting the toxicity of lead to fungi.

The toxicity of lead (Pb) to fungi in pure culture was influenced by several abiotic factors: pH, inorganic anions, clay minerals, and particulate (humic acid) and soluble organic matter. The toxicity of Pb was potentiated under acidic conditions (pH 5 and 6), and phosphate or carbonate anions reduced the toxicity, apparently as a result of the formation of sparingly soluble Pb salts. Clay minerals (montmorillonite greater than attapulgite greater than kaolinite) and particulate humic acid protected against the toxicity of Pb, presumably as the result of sorption, by cation exchange of the Pb to the exchange complexes, which reduced its availability for uptake by the fungi. Soluble organics, such as tryptone, yeast extract, cysteine, succinic acid, and increasing concentrations of neopeptone, also reduced the toxicity of Pb.

Aluminum Silicates↗

Effects of humic and fulvic acids on poliovirus concentration from water by microporous filtration.

Because naturally occurring organic matter is thought to interfere with virus adsorption to microporous filters, humic and fulvic acids isolated from a highly colored, soft surface water were used as model organics in studies on poliovirus adsorption to and recovery from electropositive Virosorb 1MDS and electronegative Filterite filters. Solutions of activated carbon-treated tap water containing 3, 10, and 30-mg/liter concentrations of humic or fulvic acid were seeded with known amounts of poliovirus and processed with Virosorb 1MDS filters at pH 7.5 or Filterite filters at pH 3.5 (with and without 5 mM MgCl2). Organic acids caused appreciable reductions in virus adsorption and recovery efficiencies with both types of filter. Fulvic acid caused greater reductions in poliovirus recovery with Virosorb 1MDS filters than with Filterite filters. Fulvic acid interference with poliovirus recovery by Filterite filters was overcome by the presence of 5 mM MgCl2. Although humic acid reduced poliovirus recoveries by both types of filter, its greatest effect was on virus elution and recovery from Filterite filters. Single-particle analyses demonstrated MgCl2 enhancement of poliovirus association with both organic acids at pH 3.5. The mechanisms by which each organic acid reduced virus adsorption and recovery appeared to be different for each type of filter.

Adsorption↗

Effects of humic materials on virus recovery from water.

Humic and fulvic acids were tested for their ability to interfere with virus recovery by microporous filters. Two electropositively charged types of filter (Seitz S and Zeta Plus 60S) were used to concentrate poliovirus in the presence of humic materials. Humic acid inhibited virus adsorption, but even at the highest humic acid concentrations tested (200 mg/liter), 30 to 40% of the virus was recovered by the filters. Fulvic acid, tested with Zeta Plus filters, did not affect virus recovery. For comparison, two electronegatively charged filter types were tested (Cox and Balston). These two types of filter were more sensitive to interference at lower concentrations of humic acid than the more positively charged filters. With Balston filters, at humic acid concentrations above 10 mg/liter, most of the virus was recovered in the filtrate. Fulvic acid, tested with Balston filters, did not interfere with virus recovery. With the electropositively charged filters, the humic materials adsorbed efficiently, even at high input concentrations. Interference with virus adsorption occurred at humic acid concentrations which were below the level of saturation of the filters. In addition, in high-volume experiments, humic acid led to premature blockage of the filters. The efficiency of virus recovery by a second concentration step, organic flocculation of the filter eluate, was tested. For all the filter types tested, this procedure was not affected by the presence of humic or fulvic acid in the input water.

Benzopyrans↗

Electrostatic mechanism of survival of virulent Aeromonas salmonicida strains in river water.

The ecological mechanism of survival of Aeromonas salmonicida, the bacterial pathogen of fish furunculosis, in river water was investigated by laboratory-based experiments with two virulent strains (which were autoagglutinating) and two virulent strains (which were nonagglutinating). A difference in net electrical charge of A. salmonicida cells was detected by electrophoresis; cells of the virulent strains were negative, whereas cells of the avirulent strains were positive. Despite the loss of viable cells within a week in distilled water and physiological saline (0.85% sodium chloride), the cells of the virulent strains survived for more than 15 weeks in the presence of diluted humic acid (10 micrograms/ml), tryptone (10 micrograms/ml), and cleaned river sand (100 g/100 ml of medium), but loss of viable cells occurred within 5 weeks in the absence of sand. The cells of the avirulent strains lost viability within 2 weeks with no relation to the presence of sand. Using ion-exchange columns, humic acid and the amino acids of tryptone were found to be anionic and cationic in water (pH 7.0), respectively. Sand particles had a high capacity to adsorb humic acid alone and amino acid-humic acid complexes. Thirty to fifty times the environmental concentration of amino acids (10 micrograms/ml) were accumulated on the surface of sand particles, thereby permitting only bacterial cells carrying net negative electrical charges (virulent cells) to survive for a long period on the surface of the sand particles. These electrostatic interrelationships among river sand, humic acid, and bacterial cells are closely implicated in the mechanism of long-term survival of virulent A. salmonicida in river sediments.

Adsorption↗

Humic acid interference with virus recovery by electropositive microporous filters.

The effects of humic acid on poliovirus type 1 recovery from water by Zeta Plus 60S filters were investigated. The humic acid interfered by preventing virus adsorption to the filters, and the interference increased as a function of the amount of humic acid filtered. Humic acid decreased virus adsorption when filtered before the virus, but did not elute virus which had adsorbed to the filters. The effects on virus recovery were not due to alterations in virus titer or neutralizability. The addition of AlCl3, which improved virus recovery by electronegative filters in the presence of humic acid, did not aid in overall virus recovery by the Zeta Plus filters in the presence or absence of humic acid. However, the salt and humic acid in combination improved virus adsorption and concurrently reduced virus elution efficiency. The addition of NaH2PO4 had no direct effect on virus recovery and did not alter the effect of humic acid. In an attempt to identify the components of humic acid responsible for the interference, humic materials were fractionated by size by using Sephadex gel chromatography and dialysis, and the fractions were tested for interfering activity. Interference was not associated with specific size fractions of the humic materials.

Adsorption↗

Detection of hepatitis A virus in seeded estuarine samples by hybridization with cDNA probes.

The development and trials of a nucleic acid hybridization test for the detection of hepatitis A virus (HAV) in estuarine samples within 48 h are described. Approximately 10(4) physical particles of HAV per dot could be detected. Test sensitivity was optimized by the consideration of hybridization stringency, 32P energy level, probe concentration, and nucleic acid binding to filters. Test specificity was shown by a lack of cross-hybridization with other enteroviruses and unrelated nucleic acids. Potential false-positive reactions between bacterial DNA in samples and residual vector DNA contamination of purified nucleotide sequences in probes were eliminated by DNase treatment of samples. Humic acid at concentrations of up to 100 mg/liter caused only insignificant decreases in test sensitivity. Interference with hybridization by organic components of virus-containing eluates was removed by proteinase K digestion followed by phenol extraction and ethanol precipitation. The test is suitable for detecting naturally occurring HAV in samples from polluted estuarine environments.

Animals↗