Phylogenetic assessment of culture collection strains of Thiobacillus thioparus, and definitive 16S rRNA gene sequences for T. thioparus, T. denitrificans, and Halothiobacillus neapolitanus.

The 16S rRNA gene sequences of 12 strains of Thiobacillus thioparus held by different culture collections have been compared. A definitive sequence for the reference type strain (Starkey; ATCC 8158(T)) was obtained. The sequences for four examples of the Starkey type strain were essentially identical, confirming their sustained identity after passage through different laboratories. One strain (NCIMB 8454) was reassigned as a strain of Halothiobacillus neapolitanus, and a second (NCIMB 8349) was a species of Thermithiobacillus. These two strains have been renamed in their catalog by the National Collection of Industrial and Marine Bacteria. The 16S rRNA gene sequence of the type strain of Halothiobacillus neapolitanus (NCIMB 8539(T)) was determined and used to confirm the identity of other culture collection strains of this species. The reference sequences for the type strains of Thiobacillus thioparus and Halothiobacillus neapolitanus have been added to the online List of Prokaryotic Names with Standing in Nomenclature. Comparison of the 16S rRNA gene sequences available for strains of Thiobacillus denitrificans indicated that the sequence for the type strain (NCIMB 9548(T)) should always be used as the reference sequence for new and existing isolates.

Complete genome sequence of Isosphaera pallida type strain (IS1B).

Isosphaera pallida (ex Woronichin 1927) Giovannoni et al. 1995 is the type species of the genus Isosphaera. The species is of interest because it was the first heterotrophic bacterium known to be phototactic, and it occupies an isolated phylogenetic position within the Planctomycetaceae. Here we describe the features of this organism, together with the complete genome sequence and annotation. This is the first complete genome sequence of a member of the genus Isosphaera and the third of a member of the family Planctomycetaceae. The 5,472,964 bp long chromosome and the 56,340 bp long plasmid with a total of 3,763 protein-coding and 60 RNA genes are part of the Genomic Encyclopedia of Bacteria and Archaea project.

Complete genome sequence of 'Thermobaculum terrenum' type strain (YNP1).

'Thermobaculum terrenum' Botero et al. 2004 is the sole species within the proposed genus 'Thermobaculum'. Strain YNP1(T) is the only cultivated member of an acid tolerant, extremely thermophilic species belonging to a phylogenetically isolated environmental clone group within the phylum Chloroflexi. At present, the name 'Thermobaculum terrenum' is not yet validly published as it contravenes Rule 30 (3a) of the Bacteriological Code. The bacterium was isolated from a slightly acidic extreme thermal soil in Yellowstone National Park, Wyoming (USA). Depending on its final taxonomic allocation, this is likely to be the third completed genome sequence of a member of the class Thermomicrobia and the seventh type strain genome from the phylum Chloroflexi. The 3,101,581 bp long genome with its 2,872 protein-coding and 58 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

A new process for organizing assessments of social, economic, and environmental outcomes: case study of wildland fire management in the USA.

Ecological risk assessments typically are organized using the processes of planning (a discussion among managers, stakeholders, and analysts to clarify ecosystem management goals and assessment scope) and problem formulation (evaluation of existing information to generate hypotheses about adverse ecological effects, select assessment endpoints, and develop an analysis plan). These processes require modification to be applicable for integrated assessments that evaluate ecosystem management alternatives in terms of their ecological, economic, and social consequences.We present 8 questions that define the steps of a new process we term integrated problem formulation (IPF), and we illustrate the use of IPF through a retrospective case study comparing 2 recent phases of development of the Fire Program Analysis (FPA) system, a planning and budgeting system for the management of wildland fire throughout publicly managed lands in the United States. IPF extends traditional planning and problem formulation by including the explicit comparison of management alternatives, the valuation of ecological, economic and social endpoints, and the combination or integration of those endpoints. The phase 1, limited prototype FPA system used a set of assessment endpoints of common form (i.e., probabilities of given flame heights over acres of selected land-resource types), which were specified and assigned relative weights at the local level in relation to a uniform national standard. This approach was chosen to permit system-wide optimization of fire management budget allocations according to a cost-effectiveness criterion. Before full development, however, the agencies abandoned this approach in favor of a phase 2 system that examined locally specified (rather than system-optimized) allocation alternatives and was more permissive as to endpoint form. We demonstrate how the IPF process illuminates the nature, rationale, and consequences of these differences, and argue that its early use for the FPA system may have enabled a smoother development path.

Fire drives transcontinental variation in tree birch defense against browsing by snowshoe hares.

Fire has been the dominant disturbance in boreal America since the Pleistocene, resulting in a spatial mosaic in which the most fire occurs in the continental northwest. Spatial variation in snowshoe hare (Lepus americanus) density reflects the fire mosaic. Because fire initiates secondary forest succession, a fire mosaic creates variation in the abundance of early successional plants that snowshoe hares eat in winter, leading to geographic variation in hare density. We hypothesize that fire is the template for a geographic mosaic of natural selection: where fire is greatest and hares are most abundant, hare browsing has most strongly selected juvenile-phase woody plants for defense. We tested the hypothesis at multiple spatial scales using Alaska birch (Betula neoalaskana) and white birch (Betula papyrifera). We also examined five alternative hypotheses for geographic variation in antibrowsing defense. The fire-hare-defense hypothesis was supported at transcontinental, regional, and local scales; alternative hypotheses were rejected. Our results link transcontinental variation in species interactions to an abiotic environmental driver, fire. Intakes of defense toxins by Alaskan hares exceed those by Wisconsin hares, suggesting that the proposed selection mosaic may coincide with a geographic mosaic of coevolution.

Use of the DiversiLab repetitive sequence-based PCR system for genotyping and identification of Archaea.

Repetitive elements are short stretches of DNA that are randomly distributed throughout the chromosomes of prokaryotes. The use of PCR primers to amplify intervening sequences of DNA between specific repetitive elements in Bacteria has become a standard method for rapidly genotyping bacterial strains and providing good resolution between multiple strains within a single species. Rapid, standardized methods for high resolution genotyping of Archaea are not widely available. We evaluated the DiversiLab system from Bacterial Barcodes that utilizes a kit-based repetitive sequence-based (rep-PCR) method that has been optimized for genotyping DNA was extracted from the source organisms using either a standard chemical DNA extraction kit or Whatman FTA paper. Rep-PCR was performed using an archaeal primer set and, the products were run on an Agilent, Lab-on-a-Chip DNA analyzer. Results were analyzed and compared using DiversiLab web-based software from Bacterial Barcodes. Seventy-nine strains representing 27 genera of Crenarchaeota and Euryarchaeota were analyzed. All the organisms could be successfully genotyped and the results were reproducible. We could not detect differences in rep-PCR profiles between DNA extracted using the chemical extraction kit and FTA paper. Thus far, 14 genera and 32 species of methanogens have been analyzed, and all yielded unique genotypes. For halophiles, 11 genera and 28 different species were analyzed, and all yielded unique genotypes. A comparison of 7 different strains of Halobacterium salinarium demonstrated that 6 of the 7 strains had a unique genotype. A comparison of 4 strains of Methanosarcina mazei indicated that each strain produced a unique genotype. There was little systematic inference that could be made from dendrograms comparing different strains, species, and genera of Archaea based on UPGMA cluster analysis. Based on these results, rep-PCR was a useful tool for the genotyping and strain identification of Archaea.

Growth characteristics of microorganisms on commercially available animal-free alternatives to tryptic soy medium.

The growth characteristics of a range of bacteria and fungi that are routinely used in quality control practices were compared for two representative vegetable-based tryptic soy formulations. All of the representative microorganisms grew well on the vegetable-based media and the media provided suitable recoveries of the organisms following simulated storage. Subtle phenotypic changes were observed between cultures grown on different media, but these did not significantly change the strain identification.

Glycine betaine as a cryoprotectant for prokaryotes.

Osmoprotectants are low molecular weight, hydrophilic, nontoxic molecules that assist a cell under osmotic stress to stabilize its concentration of internal solutes. These properties are similar to compounds used as cryoprotectants for the preservation of prokaryotic cells during freezing. This study tested the ability of a common compatible solute, glycine betaine (GB), to act as a cryoprotectant. In a series of freeze-drying studies using a variety of prokaryotes, GB performed as well, or better than, two commonly used cryoprotectants, sucrose/bovine serum albumin (S/BSA) and trehalose/dextran (T/D). GB did especially well maintaining cell viability after long-term storage (simulated equivalent of 20 years) for microorganisms like Neisseria gonorrhoeae and Streptococcus pneumoniae. GB was tested for its ability to preserve members of the genus Acidothiobacillus, a difficult genus to preserve. For two strains of Acidithiobacillus ferrooxidans that were preserved using liquid drying, GB performed as well as S/BSA. Results were more mixed for two strains of Acidithiobacillus thiooxidans; one strain could be preserved with S/BSA but not GB, the other strain gave low recoveries with both cryoprotectants. GB also proved to be a useful cryoprotectant for liquid nitrogen preservation yielding equivalent results to the cryopreservative, glycerol for halophilic archaea, and neutrophilic Fe-oxidizing bacteria. These results indicate that GB is a simple and useful cryoprotectant that works for a wide range of prokaryotic organisms under different cryopreservation regimens.

Desulfonatronum thiodismutans sp. nov., a novel alkaliphilic, sulfate-reducing bacterium capable of lithoautotrophic growth.

A novel alkaliphilic, sulfate-reducing bacterium, strain MLF1(T), was isolated from sediments of soda Mono Lake, California. Gram-negative vibrio-shaped cells were observed, which were 0.6-0.7x1.2-2.7 micro m in size, motile by a single polar flagellum and occurred singly, in pairs or as short spirilla. Growth was observed at 15-48 degrees C (optimum, 37 degrees C), >1-7 % NaCl, w/v (optimum, 3 %) and pH 8.0-10.0 (optimum, 9.5). The novel isolate is strictly alkaliphilic, requires a high concentration of carbonate in the growth medium and is obligately anaerobic and catalase-negative. As electron donors, strain MLF1(T) uses hydrogen, formate and ethanol. Sulfate, sulfite and thiosulfate (but not sulfur or nitrate) can be used as electron acceptors. The novel isolate is a lithoheterotroph and a facultative lithoautotroph that is able to grow on hydrogen without an organic source of carbon. Strain MLF1(T) is resistant to kanamycin and gentamicin, but sensitive to chloramphenicol and tetracycline. The DNA G+C content is 63.0 mol% (HPLC). DNA-DNA hybridization with the most closely related species, Desulfonatronum lacustre Z-7951(T), exhibited 51 % homology. Also, the genome size (1.6x10(9) Da) and T(m) value of the genomic DNA (71+/-2 degrees C) for strain MLF1(T) were significantly different from the genome size (2.1x10(9) Da) and T(m) value (63+/-2 degrees C) for Desulfonatronum lacustre Z-7951(T). On the basis of physiological and molecular properties, the isolate was considered to be a novel species of the genus Desulfonatronum, for which the name Desulfonatronum thiodismutans sp. nov. is proposed (the type strain is MLF1(T)=ATCC BAA-395(T)=DSM 14708(T)).

Aerobic growth of Anoxybacillus pushchinoensis K1(T): emended descriptions of A. pushchinoensis and the genus Anoxybacillus.

In this work, corrections are made to the descriptions of the species Anoxybacillus pushchinoensis corrig. and the genus ANOXYBACILLUS: Experiments to determine the relationship of A. pushchinoensis K1(T) to oxygen showed that it was capable of aerobic growth, but preferred to grow anaerobically. During aerobic growth, the redox indicator resazurin was reduced as a result of hydrogen gas production. The facultatively anaerobic nature of K1(T) was ascertained by cultivation in aerobic liquid medium, where growth began at the bottom of the tube. The anaerobic nature of K1(T) was also indicated by a negative catalase reaction. This work is submitted to correct the description of the species A. pushchinoensis from obligate anaerobe to aerotolerant anaerobe and to emend the description of the genus Anoxybacillus from obligate anaerobes or facultative anaerobes to aerotolerant anaerobes or facultative anaerobes.

Characterization of members of the Legionellaceae family by automated ribotyping.

In order to implement a new and reliable method for characterizing different species of Legionella, a genetic fingerprinting study with an automated ribotyping system (RiboPrinter) was completed with members of this genus which were deposited at the American Type Culture Collection. The RiboPrinter examined the different patterns of EcoRI digestion fragments from the rRNA operons of 110 strains, representing 48 of the 49 described Legionella species as well as 70 serogroups of those species. Distinctive and consistent patterns were obtained for the type strains of the 48 species investigated. Legionella pneumophila subsp. fraseri and L. pneumophila subsp. pascullei each generated a specific pattern, whereas L. pneumophila subsp. pneumophila produced six different fingerprint patterns. No correlation seemed to exist between the ribotypes obtained and the 15 serotypes of L. pneumophila. For the other species, those with two known serogroups presented two distinctive patterns with the RiboPrinter with the exception of L. hackeliae and L. quinlivanii, which yielded only one pattern. We also encountered ribotypes for strains which were not identified to the species level. The ribotypes generated for these strains with the RiboPrinter did not match those generated for known type strains, suggesting the putative description of new serogroups or species. Although the automated system did not have sufficient discriminatory ability to serve as an epidemiological tool in a clinical setting, it appeared to be a powerful tool for general genomic analysis of the Legionella isolates (e.g., determination of new species) and assessment of the interrelationship among Legionella strains through the RiboPrinter database connection.