Nitrogen and Phosphorus for Growth of Oil-Degrading Microorganisms in Seawater

Seawater was supplemented with NH₄⁺ and P to determine concentrations of N and P adequate for supporting exponential growth of bacteria utilizing crude oil, and to determine maximum rates of N and P uptake. Oil-degrading microorganisms were obtained by enrichment culture of indigenous oil-utilizing microorganisms in seawater. NH₄⁺ at a concentration of 5.5 µM was limiting to growth of bacteria on crude oil. Exponential growth occurred at concentrations higher than 30 µM NH₄⁺. The P concentration of 0.13 µM was limiting to growth of bacteria on crude oil. Exponential growth occurred at 1.8 |J,M P. The maximum NH4+ consumption rate was 426 ± 30 |J,g NH₄⁺ L^-1 hr^-1, and the maximum uptake rate of P was 48±4 µg P L^-1 hr^-1. Uptake of N and P with time showed zero-order kinetics, likely due to substrate solubility limitations. The uptake ratio of N:P was approximately 7:1 on a weight basis. Natural concentrations of N and P in marine and estuarine systems after hydrocarbon spillage initially may not limit oil biodegradation but may become limiting if adequate flux does not occur to replenish N and P depleted by microbial consumption.     

Sheen Screen, a Miniaturized Most-Probable-Number Method for Enumeration of Oil-Degrading Microorganisms

Sheen Screen is a miniaturized method for enumerating oil-degrading microorganisms. The technique relies on the ability of oil-degrading microorganisms to emulsify oil when provided as a sole carbon source in 24-well tissue culture plates. Sediments that actively respire hydrocarbons have high numbers of Sheen Screen-positive microorganisms.     

Diversity of Oil-Degrading Microorganisms in the Gulf of Finland (Baltic Sea) in Spring and in Summer

Diversity of the oil-degrading microbial strains isolated from the water and sediments of the Gulf of Finland (Baltic Sea) in winter and in summer was studied. Substrate specificity of the isolates for aliphatic and aromatic hydrocarbons was studied. The isolates belonged to 32 genera of the types Proteobacteria (alpha-, beta-, and gammaproteobacteria), Actinobacteria,Firmicutes, and Bacteroidetes. Seasonal variations of the oil-degrading microbial communities was revealed. The presence of the known genes responsible for the degradation of oil aliphatic and aromatic hydrocarbons was determined. The alkB sequence of the alkane hydroxylase gene was found in ~16% of the studied strains. The sequence of the phnAc phenanthrene 3,4- dioxygenase was found in Sphingobacterium sp. and Arthrobacter sp. isolates retrieved in winter and summer. In five Pseudomonas sp. strains from winter samples, the classical operons of naphthalene degradation (nah) were localized in catabolic plasmids, of which three belonged to IncР-9, one, to IncР-7, and two to an unidentified incompatibility group. Burkholderia and Delftia strains contained the operons for naphthalene degradation via salicylate and gentisate (nag). The presence of nag genes has not been previously reported for Delftia spp. strains. The sequences of the nagG salicylate 5-hydroxylase gene were also found in Achromobacter, Sphingobacterium, and Stenotrophomonas strains.     

Molecular Characterization of an Oil-Degrading Cyanobacterial Consortium

Recent studies have shown that the cyanobacterium Microcoleus chthonoplastes forms a consortium with heterotrophic bacteria present within the cyanobacterial sheath. These studies also show that this consortium is able to grow in the presence of crude oil, degrading aliphatic heterocyclic organo-sulfur compounds as well as alkylated monocyclic and polycyclic aromatic hydrocarbons. In this work, we characterize this oil-degrading consortium through the analysis of the 16S rRNA gene sequences. We performed the study in cultures of Microcoleus grown in mineral medium and in cultures of the cyanobacterium grown in mineral medium supplemented with crude oil. The results indicate that most of the clones found in the polluted culture correspond to well-known oil-degrading and nitrogen-fixing microorganisms, and belong to different phylogenetic groups, such as the Alpha, Beta, and Gamma subclasses of Proteobacteria, and the Cytophaga/Flavobacteria/Bacteroides group. The control is dominated by one predominant organism (88% of the clones) closely affiliated to Pseudoxanthomonas mexicana (similarity of 99.8%). The presence of organisms closely related to well-known nitrogen fixers such as Rhizobium and Agrobacterium suggests that at least some of the cyanobacteria-associated heterotrophic bacteria are responsible for nitrogen fixation and degradation of hydrocarbon compounds inside the polysaccharidic sheath, whereas Microcoleus provides a habitat and a source of oxygen and organic matter.     

Investigation of Microorganisms in Bentonite Deposits

Microbiological characteristics of bentonite deposits were investigated as a natural analogue of microbial behavior in the buffer material for geological disposal of radioactive waste. Distributions of microorganisms in bentonite were examined at four sites in two different bentonite deposits in Japan. The sites included pond bottom, wetland, and wet mine gallery environments where bentonite layers have been left undisturbed for 2 to 30 years. Excavation was performed without using drilling water and the center parts of the cores were used for microbial examination. Plate counts with R2A medium of aerobic and anaerobic bacteria at the drilling mouth ranged from 10⁵ to 10⁷/g DW (dry weight) and from 10³ to 10⁶/g DW, respectively. The CFDA-AM (Carboxyfluorescein diacetate acetoxymethyl ester) cell counts ranged from 10⁶ to 10⁹/g DW. Bacterial numbers in the bentonite layers declined with distance from the drilling mouth; both aerobes and anaerobes were less than 10²/g DW and CFDA-AM cell counts less than 10⁶/g DW for core samples taken from approximately 1 m depth, except at the pond bottom. These results suggest that microbial activity in natural bentonite is lower than in typical soils and aquatic sediments and does not spread easily.     

Persistence of Halophylic Methanogens and Oil-Degrading Bacteria in an Offshore Oil-Producing Facility

Microbial communities in a high saline, Tetrakis-Hydroxymethyl Phosphonium Sulfate (THPS) and nitrate-treated Nigerian oil-producing facilities were investigated. Methanogens in produced water samples preferred methanol, while those in pig-run samples (oily wastes from pipelines) preferred H₂/CO₂, as substrates to produce methane and stimulate metal corrosion. The results coincide with the dominance of methylotrophic and hydrogenotrophic methanogens in the respective samples. The same microbial populations were also THPS and high salinity tolerant. The nitrate reducers and hydrocarbon degraders were also dominant in the reservoir. A more inclusive and effective mitigation strategy is therefore required to effectively tackle biocide resistant methanogens in biocide treated oilfield.     

Development of an Oil-Degrading Biopreparation by Activation of Aboriginal Hydrocarbon-Oxidizing Microflora

A method of activation of aboriginal hydrocarbon-oxidizing microorganisms for remediation of soil and water basins polluted with oil products was developed. The optimum composition of activating additives was found (g/l): mineral components, 10.0; oil, 5.0; and a synthetic detergent, 0.2. The resulting biopreparations increased the degree of purification by factors of 4–8 in soil and 18–24 in water when applied at a concentration of 10⁷ cells/g(ml).     

Study of the Oil-Degrading Activity of Caspian Shore Microflora

Soil samples from the shore of Baku Bay (Caspian Sea) were studied. The content of oil products in the samples amounted to 2.7–8.0 wt %. The total counts of microorganisms were 1.8 × 10⁶ cells per g soil, which is by two orders of magnitude lower compared to soils free from oil pollution. In addition, the diversity of microflora is considerably narrower. The samples were used to isolate pure cultures of microbial degraders capable of assimilating 24 to 32% of the oil introduced into liquid nutrient medium in six days. It was demonstrated that the fraction composition of residual oil changed and both light and heavy oil fractions were degraded.     

Stability of Recombinant Plasmids in Transgenic Microorganisms under Different Environmental Conditions

The copy number of R plasmids weakly depends on the selective pressure of the respective antibiotic but does depend on the physiology of the host species and the type of plasmids and cloned genes, whose expression leads to a further load on the biosynthetic apparatus of cells. The last factor is critical in the maintenance of recombinant plasmids in transgenic microorganisms.     

An Experimental Investigation of Bioconvection in Three Species of Microorganisms*

SYNOPSIS. A series of experiments was carried out on bioconvection using cultures of Polytomella agilis, Tetrahymena pyriformis and Chlamydomonas moewusii. During convection these microorganisms disperse into more and less dense regions causing visible patterns. These patterns, called the cell plan forms, were found to change with increasing Rayleigh numbers above Rc, the lowest concentration gradient at which bioconvection occurs. Hexagonal convection cells were observed near the critical Rayleigh number, Rc. As the Rayleigh number was increased the cell plan form changed from hexagons to 2-dimensional rolls and then to square convection cells. The size of the observed convection cells was less than expected for such a physical phenomenon. This appears to result from an increasing tendency of the microorganisms to circulate near the upper surface at higher Rayleigh numbers. The square convection cells appear to be unique to bioconvection. The inhibition of bioconvection is directly linked to an increased mortality rate. Observations on the nature of negative geotaxis were made which tend to support the statocyst hypothesis.     

Natural Shorelines Promote the Stability of Fish Communities in an Urbanized Coastal System

Habitat loss and fragmentation are leading causes of species extinctions in terrestrial, aquatic and marine systems. Along coastlines, natural habitats support high biodiversity and valuable ecosystem services but are often replaced with engineered structures for coastal protection or erosion control. We coupled high-resolution shoreline condition data with an eleven-year time series of fish community structure to examine how coastal protection structures impact community stability. Our analyses revealed that the most stable fish communities were nearest natural shorelines. Structurally complex engineered shorelines appeared to promote greater stability than simpler alternatives as communities nearest vertical walls, which are among the most prevalent structures, were most dissimilar from natural shorelines and had the lowest stability. We conclude that conserving and restoring natural habitats is essential for promoting ecological stability. However, in scenarios when natural habitats are not viable, engineered landscapes designed to mimic the complexity of natural habitats may provide similar ecological functions.     

Selection of Xenobiotic-Degrading Microorganisms in a Biphasic Aqueous-Organic System

Microbial selection on mixtures of chlorinated and nonchlorinated compounds that are poorly soluble in water and/or toxic to growing microbial cells was examined in both biphasic aqueous-organic and monophasic aqueous systems. A biphasic system in which silicone oil was used as the organic phase permitted the acceleration of acclimation, leading to rapid selection and to an increase in xenobiotic compound degradation. In contrast, acclimation, selection, and degradation were very slow in the monophasic aqueous system. The variation in microbial growth rate with the degree of dispersion (i.e., dispersion at different silicone oil concentrations and agitation rates), and cell adhesion to the silicone oil indicate that the performance of the biphasic aqueous-organic system is dependent on the interfacial area between the two phases and that microbial activity is important at this interface. Therefore, the biphasic water-silicone oil system could be used for microbial selection in the presence of xenobiotic compounds that are toxic and have low water solubility.     

The First Results of an Investigation into the Phylogenetic Diversity of Microorganisms in Southern Baikal Sediments in the Region of Subsurface Discharge of Methane Hydrates

Phylogenetic analysis of the bacterial communities in Lake Baikal bottom sediments in the region of subsurface methane hydrate discharge has been carried out using data on 16S rRNA sequences. The composition of these microbial communities is shown to be different in different horizons. Methanotrophic bacteria are found in the surface layer (0–5 cm), and uncultured bacteria constitute a great portion of this population. In deeper sediment layers (92–96 cm), a change in the microbial community occurs; specifically, a decreased homology with the known sequences is observed. The new sequences form separate clusters on a phylogenetic tree, indicating the possibly endemic nature of the bacteria revealed. Organisms related to the genus Pseudomonas constitute the main portion of the population. An archaea-related sequence was found in a horizon containing gas hydrate crystals (100–128 cm). Uncultured bacteria remain predominant.__________Translated from Mikrobiologiya, Vol. 74, No. 3, 2005, pp. 370–377.Original Russian Text Copyright © 2005 by Shubenkova, Zemskaya, Chernitsyna, Khlystov, Triboi.     

Nucleosome Interactions and Stability in an Ordered Nucleosome Array Model System

Although it is well established that the majority of eukaryotic DNA is sequestered as nucleosomes, the higher-order structure resulting from nucleosome interactions as well as the dynamics of nucleosome stability are not as well understood. To characterize the structural and functional contribution of individual nucleosomal sites, we have developed a chromatin model system containing up to four nucleosomes, where the array composition, saturation, and length can be varied via the ordered ligation of distinct mononucleosomes. Using this system we find that the ligated tetranucleosomal arrays undergo intra-array compaction. However, this compaction is less extensive than for longer arrays and is histone H4 tail-independent, suggesting that well ordered stretches of four or fewer nucleosomes do not fully compact to the 30-nm fiber. Like longer arrays, the tetranucleosomal arrays exhibit cooperative self-association to form species composed of many copies of the array. This propensity for self-association decreases when the fraction of nucleosomes lacking H4 tails is systematically increased. However, even tetranucleosomal arrays with only two octamers possessing H4 tails recapitulate most of the inter-array self-association. Varying array length shows that systems as short as dinucleosomes demonstrate significant self-association, confirming that relatively few determinants are required for inter-array interactions and suggesting that in vivo multiple interactions of short runs of nucleosomes might contribute to complex fiber-fiber interactions. Additionally, we find that the stability of nucleosomes toward octamer loss increases with array length and saturation, suggesting that in vivo stretches of ordered, saturated nucleosomes could serve to protect these regions from histone ejection.     

Microorganisms of the Upper Atmosphere: IV. Microorganisms of a Land Air Mass as It Traverses an Ocean

The viable micropopulation at three altitudes (152, 1,066, and 1,981 meters) of a land air mass as it traversed an ocean was determined. At the low altitude, a consistent pattern of decrease in numbers of land-originating microorganisms with increasing distance from shore was observed. At the higher altitudes, the observed pattern was one of irregularity. At the lower altitude the percentages of bacteria and fungi were approximately equal near the coast, but as distance from the coast increased, fungi predominated over the bacteria at all sampling locations. The greatest number of different genera of both bacteria and fungi were obtained at or near the coast. Fungi of the genera Alternaria, Hormodendron, Penicillium, and Aspergillus, and bacteria of the genera Micrococcus and Bacterium predominated at all altitudes and locations.     

Response of Microorganisms to an Accidental Gasoline Spillage in an Arctic Freshwater Ecosystem

The response of microorganisms to an accidental spillage of 55,000 gallons of leaded gasoline into an Arctic freshwater lake was studied. Shifts in microbial populations were detected after the spillage, reflecting the migration pattern of the gasoline, enrichment for hydrocarbon utilizers, and selection for leaded-gasoline-tolerant microorganisms. Ratios of gasoline-tolerant/utilizing heterotrophs to “total” heterotrophs were found to be a sensitive indicator of the degree of hydrocarbon contamination. Respiration rates were elevated in the highly contaminated area, but did not reflect differences between moderately and lightly contaminated areas. Hydrocarbon biodegradation potential experiments showed that indigenous microorganisms could extensively convert hydrocarbons to CO₂. In situ measurement of gasoline degradation showed that, if untreated, sediment samples retained significant amounts of gasoline hydrocarbons including “volatile components” at the time the lake froze for the winter. Nutrient addition and bacterial inoculation resulted in enhanced biodegradative losses, significantly reducing the amount of residual hydrocarbons. Enhanced biodegradation, however, resulted in the appearance of compounds not detected in the gasoline. Since the contaminated lake serves as a drinking water supply, treatment to enhance microbial removal of much of the remaining gasoline still may be advisable.