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Microbial diversity in production waters of a low-temperature biodegraded oil reservoir.

Microbial communities in two production waters of a low-temperature and low-salinity petroleum reservoir in Canada were examined using cultural and molecular approaches. The predominant cultivated microorganisms were homoacetogens but sulfate-reducers, acetoclastic methanogens and denitrifiers also gave significant counts. The dominant members of the culturable population were affiliated with the Firmicutes, the "Deltaproteobacteria", the "Epsilonproteobacteria", the Spirochaetes and the Euryarchaeota. 16S rRNA gene clone libraries were also constructed from the total DNA collected from production waters. The bacterial library was entirely composed by a single phylotype related to Arcobacter. The archaeal phylotypes were generally very closely related to members of the orders Methanosarcinales and Methanomicrobiales. Consistent with earlier observations, our data suggest that methanogenesis is a dominant terminal process in the reservoir. Moreover, the cross-evaluation of culture-dependent and -independent techniques also indicates that, contrary to most studies, both acetoclastic and lithotrophic methanogens may be involved in this process. This first investigation of the microbial diversity in a non water-flooded low-temperature and low-salinity petroleum reservoir expands substantially our knowledge of the extent of microbial diversity and highlights the complexity of microbial communities involved in the oil field food chain.

Alberta↗

Adaptation of Bacillus subtilis to growth at low temperature: a combined transcriptomic and proteomic appraisal.

The soil bacterium Bacillus subtilis frequently encounters a reduction in temperature in its natural habitats. Here, a combined transcriptomic and proteomic approach has been used to analyse the adaptational responses of B. subtilis to low temperature. Propagation of B. subtilis in minimal medium at 15 degrees C triggered the induction of 279 genes and the repression of 301 genes in comparison to cells grown at 37 degrees C. The analysis thus revealed profound adjustments in the overall gene expression profile in chill-adapted cells. Important transcriptional changes in low-temperature-grown cells comprise the induction of the SigB-controlled general stress regulon, the induction of parts of the early sporulation regulons (SigF, SigE and SigG) and the induction of a regulatory circuit (RapA/PhrA and Opp) that is involved in the fine-tuning of the phosphorylation status of the Spo0A response regulator. The analysis of chill-stress-repressed genes revealed reductions in major catabolic (glycolysis, oxidative phosphorylation, ATP synthesis) and anabolic routes (biosynthesis of purines, pyrimidines, haem and fatty acids) that likely reflect the slower growth rates at low temperature. Low-temperature repression of part of the SigW regulon and of many genes with predicted functions in chemotaxis and motility was also noted. The proteome analysis of chill-adapted cells indicates a major contribution of post-transcriptional regulation phenomena in adaptation to low temperature. Comparative analysis of the previously reported transcriptional responses of cold-shocked B. subtilis cells with this data revealed that cold shock and growth in the cold constitute physiologically distinct phases of the adaptation of B. subtilis to low temperature.

Adaptation, Physiological↗

Low temperature regulates Arabidopsis Lhcb gene expression in a light-independent manner.

Low temperature treatment of dark-grown seedlings of Arabidopsis thaliana results in a rapid increase in the amount of mRNAs encoding for the major polypeptides of the light-harvesting complex of photosystem II (Lhcb1 genes). This increase is transient and seems to be due mainly to the accumulation of Lhcb1*3 transcripts, indicating that low temperature differentially regulates the expression of the Arabidopsis Lhcb1 gene family in the dark. A 1.34 kb fragment of the Lhcb1*3 promoter is sufficient to confer low temperature regulation to a reporter gene in transgenic Arabidopsis etiolated seedlings, suggesting that the regulation is occurring at the transcriptional level. The cold-induced accumulation of Lhcb1*3 mRNA is not part of a general response to stressful conditions since no accumulation is detected in response to water stress, anaerobiosis or salt stress. The amount of Lhcb1*3 mRNA decrease in response to exogenous abscisic acid (ABA) suggesting that this phytohormone acts as a negative regulator. Moreover, the accumulation of Lhcb1*3 mRNAs in cold-treated ABA deficient etiolated seedlings is higher than that of wild-type and ABA insensitive etiolated seedlings, indicating that low temperature regulation of Lhcb1*3 is not mediated by ABA.

Abscisic Acid↗

Damage and derepression in Escherichia coli resulting from growth at low temperatures.

Ng, Henry (University of California, Davis), John L. Ingraham, and Allen G. Marr. Damage and derepression in Escherichia coli resulting from growth at low temperatures. J. Bacteriol. 84:331-339. 1962.-Exponentially growing cultures of Escherichia coli ML30 were subjected to abrupt changes in temperature. If the change in temperature was made within the range of temperature in which the temperature characteristic, mu, is constant, exponential growth resumed immediately at a rate characteristic of the new temperature. But, if the shifts were made to or from a temperature below the range of constant mu, the initial growth rate was intermediate to the rates normal for the initial and final temperatures. The results indicate that growth at low temperature alters or damages the cell in a way that reduces the growth rate. A period of growth at higher temperature is required to correct the damage. Release from glucose repression of the induction of beta-galactosidase occurs at a temperature coincident with low-temperature damage. Derepression may be the damage that results from growth at low temperature.

Cold Temperature↗

Physiological response of Enterococcus faecalis JH2-2 to cold shock: growth at low temperatures and freezing/thawing challenge.

Growth at low positive temperatures and induced phenotypic resistance to extreme cold temperature (freezing/thawing cycles) of Enterococcus faecalis were investigated. The effect of low temperatures on the specific growth rates was studied; use of Arrhenius profile and Ratkovsky 'square-root' model allowed determination of the 'temperature characteristic' (mu approximately equal to 13,800 cal mol-1), the critical temperature (Tcrit approximately equal to 17.9 degrees C) and the notional minimum growth temperature (T0 approximately equal to 3.6 degrees C). Preincubation of Ent. faecalis cells at low temperatures (8-16 degrees C) during periods corresponding to their generation time resulted in an increased ability of the bacterial cells to withstand short periods of freezing/thawing (-20 degrees C/+37 degrees C) challenge. Moreover, the increase of the incubation period at low positive temperature led to a higher degree of adaptation.

Cold Temperature↗

mRNA stability and localisation of the low-temperature-responsive barley gene family blt14.

Transcription and translation inhibitors have been used to investigate the role of mRNA stability in the low-temperature-regulated expression of the post-transcriptionally controlled low temperature responsive barley gene family, blt14. Genomic clones (blt14.1, blt14.2) representing additional members of the blt14 gene family have been isolated and sequenced. Gene specific probes have been used to analyse the spatial expression of each individual member of the blt14 gene family. Findings indicate that all of the genes are responsive to low temperature, but the organ distribution is different for each gene. The results indicate that blt14.0 mRNA is stabilised by a low-temperature-dependent protein factor. Taken together, the results suggest that organ-specific post-transcriptional mechanisms are important in the low-temperature regulation of blt14 gene expression.

Amino Acid Sequence↗

Why low temperature embedding for X-ray microanalytical investigations? A comparison of recently used preparation methods.

Freeze-drying followed by infiltration with resin and polymerization by UV light at low temperatures and under constant vacuum conditions is an alternative tissue preparation technique for microprobe analysis. Embedding is carried out with the nonpolar low-temperature embedding resin (Lowicryl HM20) which allows infiltration and polymerization at temperatures down to -50 degrees C. Sections of low temperature embedded material can be cut dry at -60 degrees C or at room temperature. Sectioning at low temperatures is an alternative for preparations that are difficult to cut at room temperature. The morphological preservation is adequate for the identification of structures such as mitochondria, lysosomes and different types of endoplasmic reticulum in liver cells. Some physical properties of Lowicryl resins, such as mass loss under the electron beam and high contrast, are positive characteristics for the analysis of semi-thick sections. No significant differences in the elemental composition could be detected between tissue which was freeze-dried or freeze-substituted prior to embedding. Freeze-drying is less time consuming. By avoiding contact with organic solvents the risks of ion loss and redistribution are diminished. In contrast to freeze-dried thin cryosections, low temperature embedded material can be sectioned for light microscopy and areas of interest chosen for further thin sectioning. This is of great importance in work with tissues with complicated morphology and heterogeneous cell populations. The initial preparative step--the cryofixation--determines to a high degree the morphological preservation of freeze-dried and embedded tissue.

Animals↗

Differential expression of two related, low-temperature-induced genes in Arabidopsis thaliana (L.) Heynh.

Plant cold acclimation is correlated to expression of low-temperature-induced (lti) genes. By using a previously characterized lti cDNA clone as a probe we isolated a genomic fragment that carried two closely located lti genes of Arabidopsis thaliana. The genes were structurally related with the coding regions interrupted by three similarly located short introns and were transcribed in the same direction. The nucleotide sequences of the two genes, lti78 and lti65, predict novel hydrophilic polypeptides with molecular weights of 77,856 and 64,510, respectively, lti78 corresponding to the cDNA probe. Of the 710 amino acids of LTI78 and 600 amino acids of LTI65, 346 amino acids were identical between the polypeptides, which suggests that the genes may have a common origin. Both lti78 and lti65 were induced by low temperature, exogenous abscisic acid (ABA) and drought, but the responsiveness of the genes to these stimuli was markedly different. Both the levels and the temporal pattern of expression differed between the genes. Expression of lti78 was mainly responsive to low temperature, that of lti65 to drought and ABA. In contrast to the induction of lti78, which follows separate signal pathways during low-temperature, ABA and drought treatment, the drought induction of lti65 is ABA-dependent and the low-temperature induction appears to be coupled to the ABA biosynthetic pathway. This differential expression of two related genes may indicate that they have somewhat different roles in the stress response.

Abscisic Acid↗

Static magnetization of V15 cluster at ultra-low temperatures: precise estimation of antisymmetric exchange.

In this article, the low-temperature static (adiabatic) magnetization data of the nanoscopic V15 cluster present in K6[VIV15As6O42(H2O)] x 8H2O is analyzed. The cluster anion, which attracted much attention in the past, contains a triangular VIV3 array causing frustration as a function of applied field and temperature. In the analysis, a three-spin (S = 1/2) model of V15 was employed that includes isotropic antiferromagnetic exchange interaction and antisymmetric (AS) exchange in the most general form compatible with the trigonal symmetry of the system. It was shown that, along with the absolute value of AS exchange, the orientation of the AS vector plays a significant physical role in spin-frustrated systems. In this context, the role of the different components of the AS in the low-temperature magnetic behavior of V15 was analyzed, and we were able to reach a perfect fit to the experimental data on the staircaselike dependence of magnetization versus field in the whole temperature range including extremely low temperature. Furthermore, it was possible for the first time to precisely estimate the two components of the AS vector coupling constant in a triangular unit, namely, the effective in-plane component, Dperpendicular, and the perpendicular part, Dn.

Journal Article↗

Approaches for deciphering the structural basis of low temperature enzyme activity.

An increasing number of enzymes active at low temperature are being studied to help determine the structural features important for cold-activity. This review examines the diversity of prokaryotic cold-active enzymes and the features proposed to account for low temperature activity. We then consider the difficulty of identifying the key structural features needed for cold-activity and the need to compare enzymes having different temperature optima from phylogenetically related organisms to determine features responsible for low temperature activity. In addition to studying naturally occurring enzymes, directed evolution experiments are discussed as methods for examining the proposed mechanisms influencing the thermal dependence of activity.

Bacteria↗

Low temperature high-performance liquid chromatography of cis-trans proline dipeptides.

Isomeric dipeptides which contain proline at the C-terminus and undergo on-column cis-trans isomerization were separated by reversed-phase chromatography at subambient column temperature. Chromatography at low temperatures offers a convenient method for measuring the relative concentrations of the isomers and, thus, the calculation of the equilibrium constants for such cis-trans isomerization at various conditions of solvent, pH, and temperature. Pure fractions of the cis and trans isomers were collected and used in kinetic studies of the isomerization. High-performance liquid chromatography at temperatures near the freezing point of the eluent is a potentially useful tool for the study of a wide range of biochemical molecules.

Chromatography, High Pressure Liquid↗

Low temperature prevents potentiation of norepinephrine release by phenylephrine.

The effect of 1-phenylephrine (1-PE), an alpha(1)-receptor agonist, was investigated on the release of tritiated norepinephrine ([3H]NE). Pairs of guinea pig vasa deferentia were loaded with [3H]NE, superfused continuously, and stimulated electrically. 1-PE (10, 100 microM) enhanced the basal release of tritium in concentration-dependent manner. The stimulation-evoked release of radioactivity was significantly increased by 100 microM 1-PE. Both basal and stimulation-evoked release by 1-PE were reduced by desipramine (10 microM), a monoamine uptake inhibitor. The effect of 1-PE on basal release was independent on extracellular Ca(2+) concentration ([Ca(2+)](o)) and alpha(1)-adrenoceptor blockade. However, the 1-PE-induced release was temperature dependent: at low temperature 1-PE failed to increase either basal or stimulation-evoked release of NE. Using three different temperatures (7, 12, 17 degrees C, respectively), it was found that basal release was blocked at all three temperature values but the stimulation-evoked release was inhibited only at the lower values. The effect of 1-PE on the NE release appears to involve a desipramine-, and temperature-sensitive process. These results suggest that a non-receptorial and direct carrier-mediated mechanism is involved in NE releasing effect of 1-PE.

Adrenergic Uptake Inhibitors↗

[Structural transitions of serum proteins in solutions during high- and low-temperature dehydration].

Infrared and ultraviolet spectroscopy, potentiometric titration, and refractometry, as well as analysis of lysine, tryptophan, and tyrosine availability, and enzymic hydrolysis were used to establish that high-temperature and low-temperature dehydration of solutions of serum protein is accompanied by a rearrangement of its molecular structure, resulting in changes in the number of polar radicals and hydrophobic bonds, intermolecular and intramolecular interactions, and the formation of a new stable conformation. Comparison of the two methods of dehydration showed that cryoconcentration resulted in a greater accessibility of the protein molecule to chemical agents and proteolytic enzymes, whereas heating had an opposite effect.

Amino Acids↗

Physiological adaptations involved in alkane assimilation at a low temperature by Rhodococcus sp. strain Q15.

We examined physiological adaptations which allow the psychrotroph Rhodococcus sp. strain Q15 to assimilate alkanes at a low temperature (alkanes are contaminants which are generally insoluble and/or solid at low temperatures). During growth at 5 degrees C on hexadecane or diesel fuel, strain Q15 produced a cell surface-associated biosurfactant(s) and, compared to glucose-acetate-grown cells, exhibited increased cell surface hydrophobicity. A transmission electron microscopy examination of strain Q15 grown at 5 degrees C revealed the presence of intracellular electron-transparent inclusions and flocs of cells connected by an extracellular polymeric substance (EPS) when cells were grown on a hydrocarbon and morphological differences between the EPS of glucose-acetate-grown and diesel fuel-grown cells. A lectin binding analysis performed by using confocal scanning laser microscopy (CSLM) showed that the EPS contained a complex mixture of glycoconjugates, depending on both the growth temperature and the carbon source. Two glycoconjugates [beta-D-Gal-(1-3)-D-GlcNAc and alpha-L-fucose] were detected only on the surfaces of cells grown on diesel fuel at 5 degrees C. Using scanning electron microscopy, we observed strain Q15 cells on the surfaces of octacosane crystals, and using CSLM, we observed strain Q15 cells covering the surfaces of diesel fuel microdroplets; these findings indicate that this organism assimilates both solid and liquid alkane substrates at a low temperature by adhering to the alkane phase. Membrane fatty acid analysis demonstrated that strain Q15 adapted to growth at a low temperature by decreasing the degree of saturation of membrane lipid fatty acids, but it did so to a lesser extent when it was grown on hydrocarbons at 5 degrees C; these findings suggest that strain Q15 modulates membrane fluidity in response to the counteracting influences of low temperature and hydrocarbon toxicity.

Adaptation, Physiological↗

Genetic analysis of mutations affecting growth of Saccharomyces cerevisiae at low temperature.

A large number of genes control growth of the yeast Saccharomyces cerevisiae at low temperatures (< 10 degrees ). Approximately 47 percent of the mutants selected for inability to grow at 4-5 degrees C show increased sensitivity to cycloheximide. In 3 of 4 cases tested, supersensitivity to cycloheximide and inability to grow at the low temperature segregate together and thus appear to be effects of the same mutation. Since many cold-sensitive mutants of bacteria have been found to have altered ribosomes and since cycloheximide resistance in yeast can be caused by ribosomal changes, this suggests that the mutants having low-temperature-sensitive growth may be defective in ribosome-assembly processes at the low temperatures. Two of the lts loci, lts1 and lts3 have been located on chromosome VII and another two, lts4 and lts10 on chromosome IV. A mutation, cyh10, conferring cycloheximide resistance, but not cold sensitivity, has been located close to the centromere on chromosome II.

Chromosome Mapping↗

Low temperature microspectrofluorometry: design of a 'cold chamber'.

Low temperature microspectrofluorometry allows an improvement of spectral resolution and an increase of fluorescence intensity. Suppression of fluorescence fading, or at least a marked reduction, is also obtained. A cooling chamber for microspectrofluorometric measurement is described which allows the cooling, under a microscope, of a biological sample down to liquid nitrogen temperature. Objectives with numerical apertures better than 1.0 and a magnification power up to 100X can be used. Low temperature measurements on a histological sample are presented and discussed.

Bacillus cereus↗

Molecular analysis and spatial expression pattern of a low-temperature-specific barley gene, blt101.

A cDNA clone of the previously unreported low-temperature-induced gene blt101 was isolated after a differential screen of a cDNA library prepared from low-temperature (6 degrees C day/2 degrees C night) grown barley shoot meristems. Southern blot analysis of barley ditelosomic addition lines was used to assign this single-copy gene to the long arm of chromosome 4. Analysis of steady-state levels of blt101 mRNA showed the induction of this transcript in shoot meristems upon transfer of barley (cv. Igri) plants from control (20 degrees C/15 degrees C) to low (6 degrees C/2 degrees C) temperature treatment. Further, the high level of this transcript is maintained at low temperatures but is reduced on transfer from low to control temperatures. The gene is not induced by drought or by foliar application of ABA. Analysis of segregating doubled haploid lines shows that there is no specific association of this gene with either spring/winter growth habit or frost hardiness. Examination of the spatial expression pattern revealed ubiquitous expression of blt101 in low-temperature (6 degrees C/2 degrees C) grown barley shoot meristems, mature leaves and roots.

Abscisic Acid↗

Increase of myocardial diastolic tonus at low temperatures: a Ca-phenomenon?

Myocardial diastolic tonus increases at low temperature. The chief aim of the present series of experiments was to decide whether this phenomenon should be interpreted as being caused by thermoelastic alteration or by an impairment of the Ca-pump. All experiments were performed on isolated right ventricular rabbit papillary muscles. In order to impose ramp-like and sinusoidally modulated changes of the temperature on the perfusate, a Peltier-battery was used. In Krebs-Henseleit solution with normal Ca-content (2.52 mmoles/l) changes of diastolic tonus of the driven muscle within a narrow range at low temperature amount approximately to isometric twitch tension (10(-2) up to 3.10(-2) N mm-2). Non-stimulated preparations show tension alterations of 2.10(-4) N mm-2 at best. With increasing stimulation interval the hysteresis-like loop representing changes of diastolic tonus shifts to lower temperatures. Under threefold Ca2+-concentration of the perfusate the diastolic tension varies over the total range of periodically modulated temperature in both stimulated and non-stimulated muscles. From these results it may be concluded that increasing myocardial diastolic tonus under low temperature is caused by an insufficiency of the Ca-sequestration system.

Animals↗