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Reversible transition between active and dormant microbial states in soil.

The rate of respiration obtained in the substrate-induced respiration (SIR) method can be divided into the respiration rate of growing (r) and non-growing (K) microorganisms. The fraction of r is generally small (5-20%) in soils with no recent addition of substrates, but can be 100% in soils with high substrate availability. This suggests that substrate availability determines the proportion of biomass between these groups, and implies that transitions between them can take place reversibly. These hypotheses were tested by adding three different amounts of glucose which induced first-order, zero-order, and growth-associated respiration kinetics to three soils at four pre-incubation times (4, 12, 27, and 46 days) before the SIR measurement. An abiotic flush of CO(2) in the SIR measurement was detected and corrected for before data analysis. Accumulated CO(2)-C over 4 days after glucose addition, corrected for the respiration in unamended controls, corresponded to 41-50% mineralization of the glucose-C, and the relative amount mineralized by each soil was independent of the glucose amount added. The high glucose concentration gave an increased SIR, which reverted to the initial value within 27-46 days. In a specific sample, the maximum respiration rate induced during the pre-incubation, and the amount of organisms transformed from the K to the r state, as quantified in respiration rate units in the SIR measurement, were identical to each other, and these parameters were also highly correlated to the initial glucose concentration. The K-->r transition was very fast, probably concurrent with the instantaneous increase in the respiration rate obtained by the glucose amendment. Thereafter, a slow first-order back-transition from the r to the K state ensued, with half-lives of 12, 23, and 70 days for the three soils. The results suggest the existence of community-level controls by which growth within or of the whole biomass is inhibited until it has been completely transformed into the r state. The data also suggest that the microbial specific activity is not related to the availability of exogenous substrate in a continuous fashion, rather it responds as a sharp transition between dormant and fully active. Furthermore, the inherent physiological state of the microbial biomass is strongly related to its history. It is proposed that the normal dynamics of the soil microbial biomass is an oscillation between active and dormant physiological states, while significant growth occurs only at substantial substrate amendment.

Journal Article↗

Identification of the sources of energy for nitrogen fixation and physiological characterization of nitrogen-fixing members of a marine microbial mat community.

Experimental manipulations of a microbial mat community were performed to determine sources of energy and reductant used for nitrogen fixation and to physiologically characterize the responsible diazotrophs. The dominant photolithotrophic members of this community were nonheterocystous cyanobacteria, but other potential nitrogen-fixing microorganisms were also present. Pronounced diel variability in rates of acetylene reduction was observed, with nighttime rates a factor of three to four higher than daytime rates. Acetylene reduction measured at night was dependent upon the occurrence of oxygenic photosynthesis the preceding day; mats incubated in the dark during the daytime reduced acetylene at rates comparable to those of light-incubated mats but were not able to reduce acetylene at the normally high rates the following night. The addition of various exogenous carbon compounds to these dark-incubated mats did not elicit nighttime acetylene reduction. Nighttime acetylene reduction apparently proceeds under anoxic conditions in these mats; the highest rates of acetylene reduction occur late at night. Additions of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (an inhibitor of oxygenic photosynthesis) to mats resulted in a pronounced stimulation of acetylene reduction during the day, but acetylene reduction the next night proceeded at greatly reduced rates (relative to untreated mats). This daytime stimulation, under the 3-(3,4-dichlorophenyl)-1,1-dimethylurea-induced anoxic conditions in the experimentally treated mats, was light dependent. These results suggest that nitrogen fixation in these mats may be attributed to the activities of nonheterocystous cyanobacteria utilizing storage products of oxygenic photosynthesis under anoxic conditions at night.

Journal Article↗

Molecular diversity in gene-encoded, cationic antimicrobial polypeptides.

Gene-encoded, ribosomally synthesised antimicrobial peptides (AMPs) are an ancient and pervasive component of the innate defence mechanisms used by multicellular organisms to control the natural flora and combat pathogens. Bacteria also produce such AMPs to maintain ecological niches free of rival strains. Several hundred different peptides have been characterised to date, and they show a marked degree of variability in both sequence and structure, having evolved to act against distinct microbial targets in different physiological contexts. Many of these peptides appear to function via a selective, but not receptor-mediated, permeabilisation of microbial membranes, while others interact with specific membrane associated or intracellular targets. This review presents a broad survey of different amp structural classes, emphasising both their molecular diversity and underlying similarities. The mode of action of these peptides and potential for biomedical and other application is also briefly discussed.

Amino Acid Sequence↗

Variations in digestive physiology of rats after short duration flights aboard the US space shuttle.

The purpose of this work was to assess the influence of microgravity on several endogenous and microbial parameters of digestive physiology. On the occasion of two Spacelab Life Sciences missions, SLS-1 (a 9-day space flight) and SLS-2 (a 14-day space flight), Sprague-Dawley rats flown aboard the US space shuttle were compared to age-matched ground-based controls. In both flights, exposure to microgravity modified cecal fermentation: concentration and profile of short-chain fatty acids were altered, whereas urea and ammonia remained unchanged. Only in SLS-1 was there an induction of intestinal glutathione-S-transferase. Additional analyses in SLS-2 showed a decrease of hepatic CYP450 and of colonic goblet cells containing neutral mucin. After a postflight recovery period equal to the mission length, only modifications of the hepatic and intestinal xenobiotic metabolizing enzymes still persisted. These findings should help to predict the alterations of digestive physiology and detoxification potential likely to occur in astronauts. Their possible influence on health is discussed.

Animals↗

Role of cryptic genes in microbial evolution.

Cryptic genes are phenotypically silent DNA sequences, not normally expressed during the life cycle of an individual. They may, however, be activated in a few individuals of a large population by mutation, recombination, insertion elements, or other genetic mechanisms. A consideration of the microbial literature concerning biochemical evolution, physiology, and taxonomy provides the basis for a hypothesis of microbial adaptation and evolution by mutational activation of cryptic genes. Evidence is presented, and a mathematical model is derived, indicating that powerful and biologically important mechanisms exist to prevent the loss of cryptic genes. We propose that cryptic genes persist as a vital element of the genetic repertoire, ready for recall by mutational activation in future generations. Cryptic genes provide a versatile endogenous genetic reservoir that enhances the adaptive potential of a species by a mechanism that is independent of genetic exchange.

Biological Evolution↗

Microbial genome sequencing.

Complete genome sequences of 30 microbial species have been determined during the past five years, and work in progress indicates that the complete sequences of more than 100 further microbial species will be available in the next two to four years. These results have revealed a tremendous amount of information on the physiology and evolution of microbial species, and should provide novel approaches to the diagnosis and treatment of infectious disease.

Anti-Infective Agents↗

Analysis of Microbial Communities in a Landfill Leachate Polluted Aquifer using a New Method for Anaerobic Physiological Profiling and 16S rDNA Based Fingerprinting.

Databases containing information regarding presence and activity of microbial communities will be very useful for determination of the potential for intrinsic bioremediation in landfill leachate polluted aquifers. Simple analyses such as community-level physiological profiling (CLPP) and denaturing gradient gel electrophoresis (DGGE) of 16S rDNA fragments yield large sets of data for inclusion into such databases. In this study we describe the development of a method for anaerobic CLPP, using commercially available Biolog plates. Incubation at the in situ temperature of the aquifer (10 degrees C) for 28 days was optimal for obtaining a specific, reproducible physiological profile. Anaerobic incubation was essential for profiling anaerobic communities. The anaerobic cultivation-dependent CLPP method and cultivation-independent DGGE were applied to groundwater and sediment samples from the aquifer near the Coupépolder landfill in The Netherlands. A combination of computer-assisted CLPP and DGGE analysis of both groundwater and sediment samples yielded the best separating power for characterizing microbial communities in the aquifer. Communities in groundwater were significantly different from those in the corresponding sediment. Microbial communities present in subsamples from sediment cores usually were similar for the various sampling locations. Variation was observed for the heterogeneous sediment beneath the landfill. Both anaerobic CLPP and DGGE analysis clearly separated microbial communities from the polluted aquifer underneath the landfill from those in the less or not polluted aquifer downstream and upstream of the landfill.

Journal Article↗

PCR bias in ecological analysis: a case study for quantitative Taq nuclease assays in analyses of microbial communities.

Succession of ecotypes, physiologically diverse strains with negligible rRNA sequence divergence, may explain the dominance of small, red-pigmented (phycoerythrin-rich) cyanobacteria in the autotrophic picoplankton of deep lakes (C. Postius and A. Ernst, Arch. Microbiol. 172:69-75, 1999). In order to test this hypothesis, it is necessary to determine the abundance of specific ecotypes or genotypes in a mixed background of phylogenetically similar organisms. In this study, we examined the performance of Taq nuclease assays (TNAs), PCR-based assays in which the amount of an amplicon is monitored by hydrolysis of a labeled oligonucleotide (TaqMan probe) when hybridized to the amplicon. High accuracy and a 7-order detection range made the real-time TNA superior to the corresponding end point technique. However, in samples containing mixtures of homologous target sequences, quantification can be biased due to limited specificity of PCR primers and probe oligonucleotides and due to accumulation of amplicons that are not detected by the TaqMan probe. A decrease in reaction efficiency, which can be recognized by direct monitoring of amplification, provides experimental evidence for the presence of such a problem and emphasizes the need for real-time technology in quantitative PCR. Use of specific primers and probes and control of amplification efficiency allow correct quantification of target DNA in the presence of an up to 10(4)-fold excess of phylogenetically similar DNA and of an up to 10(7)-fold excess of dissimilar DNA.

Bacteria↗

[Use of gamma-irradiation of soil for analyzing the state of microbial coeneses].

Increasing doses of gamma-irradiation eliminate some physiological and ecologo-trophical groups from microbial cenoses in soil. Soil fungi are least resistant to the action of gamma-irradiation (do not grow on agarized soil (AS) at a dose of 0.5 Mrad). Actinomycetes are more resistant (do not grow on AS at a dose of 0.75 Mrad). Bacteria are most resistant to irradiation. Asporogenous bacteria do not grow on agarized soil at a dose of 1.25 Mrad. Sporogenous bacteria survive at higher doses (1.75 Mrad). The method of gamma-irradiation permits to accomplish differential analysis of ecological groups of soil microflora and experimental synthesis of microbial associations. The effect of gamma-irradiation on soil enzymes is weak compared with autoclaving. gamma-Irradiated soils may serve as a substrate for model experiments during analysis of microbial associations.

Bacteria↗

Characterization of the microbial community of lotic organic aggregates ('river snow') in the Elbe River of Germany by cultivation and molecular methods.

Aerobic and anaerobic cultivation techniques, 16S rDNA-based phylogeny, and fluorescent in situ hybridization were used to describe the phylogenetic diversity and physiological versatility of lotic microbial aggregates ('river snow') obtained from the river Elbe. In the course of the year the 'river snow' community changed. It was characterized by a great bacterial diversity in spring, the predominant occurrence of algae in summer and reduction of the total bacterial cell count in autumn and winter. In all 'river snow' samples, more than 70% of the bacteria counted with the general DNA stain DAPI also hybridized with the Bacteria-specific probe EUB338. In situ analysis of the bacterial 'river snow' community with a comprehensive suite of specific rRNA-targeted probes revealed population dynamics to be governed by seasonal factors. During all seasons, beta-Proteobacteria constituted the numerically most important bacterial group forming up to 54% of the total cell counts. In contrast to this, the relative abundance of other major bacterial lineages ranged from 2% for the order Planctomycetales to 36% for Cytophaga-Flavobacteria. Cultivation of 'river snow' under aerobic and anaerobic conditions with a variety of different media resulted in the isolation of 40 new bacterial strains. Phenotypic and phylogenetic analyses revealed these new strains to be mostly unknown organisms affiliated to different bacterial phyla. Application of newly developed specific oligonucleotide probes proved the cultivated bacteria, including clostridia and the numerically abundant beta-Proteobacteria, as relevant in situ members of the 'river snow' community.

Journal Article↗

Microbial metabolomics: past, present and future methodologies.

Microbial metabolomics has received much attention in recent years mainly because it supports and complements a wide range of microbial research areas from new drug discovery efforts to metabolic engineering. Broadly, the term metabolomics refers to the comprehensive (qualitative and quantitative) analysis of the complete set of all low molecular weight metabolites present in and around growing cells at a given time during their growth or production cycle. This review focuses on the past, current and future development of various experimental protocols in the rapid developing area of metabolomics in the ongoing quest to reliably quantify microbial metabolites formed under defined physiological conditions. These developments range from rapid sample collection, instant quenching of microbial metabolic activity, extraction of the relevant intracellular metabolites as well as quantification of these metabolites using enzyme based and or modern high tech hyphenated analytical protocols, mainly chromatographic techniques coupled to mass spectrometry (LC-MS(n), GC-MS(n), CE-MS(n)), where n indicates the number of tandem mass spectrometry, and nuclear magnetic resonance spectroscopy (NMR).

Bacteria↗

Sheep-urine-induced changes in soil microbial community structure.

Soil microbial communities play an important role in nutrient cycling and nutrient availability, especially in unimproved soils. In grazed pastures, sheep urine causes local changes in nutrient concentration which may be a source of heterogeneity in microbial community structure. In the present study, we investigated the effects of synthetic urine on soil microbial community structure, using physiological (community level physiological profiling, CLPP), biochemical (phospholipid fatty acid analysis, PLFA) and molecular (denaturing gradient gel electrophoresis, DGGE) fingerprinting methods. PLFA data suggested that synthetic urine treatment had no significant effect on total microbial (total PLFA), total bacterial or fungal biomass; however, significant changes in microbial community structure were observed with both PLFA and DGGE data. PLFA data suggested that synthetic urine induced a shift towards communities with higher concentrations of branched fatty acids. DGGE banding patterns derived from control and treated soils differed, due to a higher proportion of DNA sequences migrating only to the upper regions of the gel in synthetic urine-treated samples. The shifts in community structure measured by PLFA and DGGE were significantly correlated with one another, suggesting that both datasets reflected the same changes in microbial communities. Synthetic urine treatment preferentially stimulated the use of rhizosphere-C in sole-carbon-source utilisation profiles. The changes caused by synthetic urine addition accounted for only 10-15% of the total variability in community structure, suggesting that overall microbial community structure was reasonably stable and that changes were confined to a small proportion of the communities.

Animals↗

Physiological and community responses of established grassland bacterial populations to water stress.

The effects of water stress upon the diversity and culturable activity of bacterial communities in the rhizosphere of an established upland grassland soil have been investigated. Intact monoliths were subjected to different watering regimens over a 2-month period to study community adaptation to moisture limitation and subsequent response to stress alleviation following rewetting. Genetic diversity was analyzed with 16S-based denaturing gradient gel electrophoresis (DGGE) of total soil-extracted DNA (rRNA genes) and RNA (rRNA transcripts) in an attempt to discriminate between total and active communities. Physiological response was monitored by plate counts, total counts, and BIOLOG-GN2 substrate utilization analyses. Controlled soil drying decreased the total number of CFU on all the media tested and also decreased the substrate utilization response. Following rewetting of dried soil, culture-based analyses indicated physiological recovery of the microbial population by the end of the experiment. In contrast, DGGE analyses of community 16S rRNA genes, rRNA transcripts and cultured communities did not reveal any changes relating to the moisture regimens, despite the observed physiological effects. We conclude that the imposed moisture regimen modulated the physiological status of the bacterial community and that bacterial communities in this soil are resistant to water stress. Further, we highlight the need for a reexamination of rRNA transcript-based molecular profiling techniques as a means of describing the active component of soil bacterial communities.

Bacteria↗

Effects of parasitization by Varroa destructor on survivorship and physiological traits of Apis mellifera in correlation with viral incidence and microbial challenge.

Varroa mites (Varroa destructor) are serious ectoparasites of honey bees (Apis mellifera). This research addresses the impact of varroa mites on survivorship, viral incidence, and physiological traits of newly-emerged worker bees. RT-PCR confirmed our previous finding that varroa parasitization was linked to high levels of deformed wing virus (DWV). In non-treatment bees, varroa parasitization combined with increased viral levels altered survivorship curves from long-survival to shorter-survival types. After challenge with live Escherichia coli, the survivorship of mite-parasitized bees was significantly lower than mite-free bees. Deformed-wing, mite-parasitized bees died on average within 1 day, even without E. coli challenge. This was correlated with the absence of an important enzyme activity in insect immunity, phenol oxidase, lacking even in those bees challenged with immuno-elicitors. The lack of inducible phenol oxidase activity indicated that the bee immune system is not fully competent upon adult emergence. Varroa parasitism also significantly reduced body weight of the parasitized bees, but body weight was not significantly correlated with the survivorship of mite-parasitized bees. Our research indicates that the combination of mite parasitization, the interaction of DWV and microbes, and a developmental immune incompetency attribute to decreased worker survivorship and have a negative impact on colony fitness.

Animals↗

Rapid methane oxidation in a landfill cover soil.

Methane oxidation rates observed in a topsoil covering a retired landfill are the highest reported (45 g m day) for any environment. This microbial community had the capacity to rapidly oxidize CH(4) at concentrations ranging from <1 ppm (microliters per liter) (first-order rate constant [k] = -0.54 h) to >10 ppm (k = -2.37 h). The physiological characteristics of a methanotroph isolated from the soil (characteristics determined in aqueous medium) and the natural population, however, were similar to those of other natural populations and cultures: the Q(10) and optimum temperature were 1.9 and 31 degrees C, respectively, the apparent half-saturation constant was 2.5 to 9.3 muM, and 19 to 69% of oxidized CH(4) was assimilated into biomass. The CH(4) oxidation rate of this soil under waterlogged (41% [wt/vol] H(2)O) conditions, 6.1 mg liter day, was near rates reported for lake sediment and much lower than the rate of 116 mg liter day in the same soil under moist (11% H(2)O) conditions. Since there are no large physiological differences between this microbial community and other CH(4) oxidizers, we attribute the high CH(4) oxidation rate in moist soil to enhanced CH(4) transport to the microorganisms; gas-phase molecular diffusion is 10-fold faster than aqueous diffusion. These high CH(4) oxidation rates in moist soil have implications that are important in global climate change. Soil CH(4) oxidation could become a negative feedback to atmospheric CH(4) increases (and warming) in areas that are presently waterlogged but are projected to undergo a reduction in summer soil moisture.

Journal Article↗

Big questions, small worlds: microbial model systems in ecology.

Although many biologists have embraced microbial model systems as tools to address genetic and physiological questions, the explicit use of microbial communities as model systems in ecology has traditionally been more restricted. Here, we highlight recent studies that use laboratory-based microbial model systems to address ecological questions. Such studies have significantly advanced our understanding of processes that have proven difficult to study in field systems, including the genetic and biochemical underpinnings of traits involved in ecological interactions, and the ecological differences driving evolutionary change. It is the simplicity of microbial model systems that makes them such powerful tools for the study of ecology. Such simplicity enables the high degrees of experimental control and replication that are necessary to address many questions that are inaccessible through field observation or experimentation.

Journal Article↗

[Quantitative description of microbial growth in a batch culture depending on the physiologic state of inocula].

The dynamics of biomass production and the respiration rate of five microorganisms grown as batch cultures were studied in detail. Cell suspensions with a known physiological state, i.e. chemostat cultures grown at a particular D value, as well as quasi steady-state populations cultivated with slow feeding and long energy-source starvation were used as inocula. The microorganisms were arbitrary subdivided into two groups. The biomass of Pseudomonas fluorescens, Bacillus subtilis and Debaryomyces formicarius accumulated smoothly with a monotonic rise of the specific growth rate to the upper level of microns. A distinct inverse correlation was established between the duration of the lag phase and the specific growth rate of the inoculum. Arthrobacter globiformis and Lipomyces tetrasporus were characterised by biphase growth referred to as false diauxia: glucose was accumulated without its oxidation during the first phase, and actual growth (just as in the first group of microorganism) occurred during the second phase. Most of the results are satisfactorily described by a simplified modification of the synthetic chemostat model.

Arthrobacter↗