Salinivibrio costicola subsp. vallismortis subsp. nov., a halotolerant facultative anaerobe from Death Valley, and emended description of Salinivibrio costicola.

Strain DVT, a halotolerant, Gram-negative, facultatively anaerobic bacterium, was isolated from a hypersaline pond located in Death Valley, California. The cells were non-spore-forming, motile, curved rods (1.0-1.8 x 0.5-0.6 microns) and occurred singly, in pairs or rarely in chains. Strain DVT was oxidase-, catalase-, Voges-Proskauer-, amylase-, gelatinase- and lipase-positive and indole-negative. Nitrate, sulfate and fumarate were not used as electron acceptors. Carbohydrates served as energy sources both aerobically and anaerobically. Strain DVT grew optimally at 37 degrees C (temperature range 20-50 degrees C) with 2.5% NaCl (NaCl range 0-12.5%) and pH 7.3 (pH range of 5.5-8.5) in a glucose/yeast extract medium with a doubling time of 20 min (aerobically) or 41 min (anaerobically). The end products of glucose fermentation were ethanol, isobutyrate, propionate, lactate, formate and CO2. Strain DVT was resistant to penicillin, D-cycloserine, streptomycin and tetracycline (200 micrograms ml-1). The G + C content was 50 mol%. 16S rRNA gene sequence analysis indicated that it was closely related to Salinivibrio costicola (97.7%) and this was confirmed by DNA-DNA hybridization (93% relatedness). However, phenotypic characteristics such as halotolerance, gas production, growth at 50 degrees C, antibiotic resistance, sugar-utilization spectrum and phylogenetic signatures are sufficiently different from Salinivibrio costicola to warrant designating strain DVT as a new subspecies of Salinivibrio costicola, Salinivibrio costicola subsp. vallismortis subsp. nov. (= DSM 8285T).

Isolation and characterization of desulfovibrio senezii sp. nov., A halotolerant sulfate reducer from a solar saltern and phylogenetic confirmation of desulfovibrio fructosovorans as a new species

A new halotolerant Desulfovibrio, strain CVLT (T = type strain), was isolated from a solar saltern in California. The curved, gram-negative, nonsporeforming cells (0.3 x 1.0-1.3 &mgr;m) occurred singly, in pairs, or in chains, were motile by a single polar flagellum and tolerated up to 12.5% NaCl. Strain CVLT had a generation time of 60 min when grown in lactate-yeast extract medium under optimal conditions (37 degreesC, pH 7.6, 2.5% NaCl). It used lactate, pyruvate, cysteine, or H2/CO2 + acetate as electron donors, and sulfate, sulfite, thiosulfate, or fumarate as electron acceptors. Elemental sulfur, nitrate, or oxygen were not used. Sulfite and thiosulfate were disproportionated to sulfate and sulfide. The G+C content of the DNA was 62 mol%. Phylogenetic analysis revealed that Desulfovibrio fructosovorans was the nearest relative. Strain CVLT is clearly different from other Desulfovibrio species, and is designated Desulfovibrio senezii sp. nov. (DSM 8436).

Caldicellulosiruptor owensensis sp. nov., an anaerobic, extremely thermophilic, xylanolytic bacterium.

An anaerobic, extremely thermophilic, xylanolytic, non-spore-forming bacterium was isolated from a sediment sample taken from Owens Lake, California, and designated strain OLT (T = type strain). Strain OLT had a Gramnegative reaction and occurred as short rods which sometimes formed long chains containing a few coccoid cells. It grew at 50-80 degrees C, with an optimum at 75 degrees C. The pH range for growth was 5.5-9.0 with an optimum at about pH 7.5. When grown on glucose at optimal conditions, its doubling time was 7.3 h. In addition to glucose, the isolate utilized sucrose, xylose, fructose, ribose, xylan, starch, pectin and cellulose. Yeast extract stimulated growth on carbohydrates but was not obligately required. The end products from glucose fermentation were lactate, acetate, ethanol, H2 and CO2. The G + C content of strain OLT was 36.6 mol%. The 16S rDNA sequence analysis indicated that strain OLT was a member of the subdivision containing Gram-positive bacteria with DNA G + C content of less than 55 mol% and clustered with members of the genus Caldicellulosiruptor. Because strain OLT is phylogenetically and phenotypically different from other members of this genus, it is proposed to designate this isolate Caldicellulosiruptor owensensis sp. nov. Strain OLT is the type strain (= ATCC 700167T).

LEMMA: a language for easy medical models analysis.

The quality of health care systems and processes is becoming a prominent problem and more and more efforts are devoted to define methodologies and tools to measure and assure quality of care. New methods are required to optimize health care processes to guarantee high quality standards within (limited) available resources. Resource optimizations able to preserve the quality of treatments require good models of medical processes. This paper presents LEMMA, a new notation to model medical processes. LEMMA provides physicians with intuitive graphical elements to design their models. At the same time a high level timed Petri net corresponding to the designed model is built automatically. In this way, LEMMA models are ascribed formal semantics and can be executed and analyzed automatically. The dual language approach followed in this paper allows physicians to gain all the benefits of formal methods without being proficient in them. Medical users manage simple graphical elements, while Petri nets ensure formality and validation capabilities. In this way LEMMA mixes formal and informal notations, overcoming the problems of both the approaches. The definition of the notation has been supported by the development of an environment to design LEMMA models. The environment, besides letting us experiment with the notation, has been employed to define and analyze real case studies.

Haloanaerobium alcaliphilum sp. nov., an anaerobic moderate halophile from the sediments of Great Salt Lake, Utah.

A strictly anaerobic, moderately halophilic, gram-negative, rod-shaped bacterium was isolated from Great Salt Lake, Utah, sediments and designated GSLST (T = type strain). Strain GSLST grew optimally at pH 6.7 to 7.0 but had a very broad pH range for growth (pH 5.8 to 10.0). The optimum temperature for growth was 37 degrees C, and no growth occurred at 15 or 55 degrees C. The optimum salt concentration for growth was 10%. Strain GSLST required yeast extract and Trypticase peptone to ferment carbohydrates, pyruvate, and glycine betaine. Strain GSLST was resistant to penicillin, D-cycloserine, tetracycline, and streptomycin. The G + C content of this isolate was 31 mol%. The fermentation products from glucose utilization were acetate, butyrate, lactate, CO2, and H2, and in addition strain GSLST fermented glycine betaine to acetate and trimethylamine. All of these traits distinguish this organism from all previously described halophilic anaerobes. The 16S rRNA gene sequence of strain GSLST was found to be similar to, but also significantly different from, the 16S rRNA sequences of Haloanaerobium salsugo and Haloanaerobium praevalens. Therefore, strain GSLST (= DSM 8275T) is described as a new species, Haloanaerobium alcaliphilum.

Waste treatment integration in space.

The circumstances and criteria for space-based waste treatment bioregenerative life-support systems differ in many ways from those needed in terrestrial applications. In fact, the term "waste" may not even be appropriate in the context of nearly closed, cycling, ecosystems such as those under consideration. Because of these constraints there is a need for innovative approaches to the problem of "materials recycling". Hybrid physico-chemico-biological systems offer advantages over both strictly physico-chemico or biological approaches that would be beneficial to material recycling. To effectively emulate terrestrial cycling, the use of various microbial consortia ("assemblies of interdependent microbes") should be seriously considered for the biological components of such systems. This paper will examine the use of consortia in the context of a hybrid-system for materials recycling in space.

Increased beta-adrenergic receptors in the light-density membrane fraction in lungs from senescent rats.

There is a dynamic balance between cell surface and intracellular beta-adrenergic receptors. To investigate whether this balance is altered with aging, we determined the number of beta-adrenergic receptors in the heavy- (48,000 x g) and light-density (200,000 x g) membrane fractions from lungs and hearts of 4-, 13-, and 25-month-old rats. There were no differences in the density of receptors with age in the heavy membrane fraction. However, the density of receptors in the light-density fraction from lungs increased with age (109 +/- 7, 230 +/- 43, and 225 +/- 36, F(2,12) = 4.4, p less than .05). Receptors in the heavy membrane fraction expressed both high and low affinity binding for isoproterenol, whereas receptors in light-density fraction expressed only low affinity binding. In contrast to the lung, there were no changes in the number of receptors with age in the light-density membrane fraction from hearts. These data indicate there are increased numbers of beta-adrenergic receptors in the light membrane fraction in lungs from aged rats, which are separated from the other components of the adenylate cyclase complex.

Glucocorticoids modulate beta-adrenoceptor subtypes and adenylate cyclase in brown fat.

Thermogenesis in brown adipose tissue (BAT) serves as a regulator of body temperature and weight maintenance. Thermogenesis can be stimulated by catecholamine activation of adenylate cyclase through the beta-adrenergic receptor. Glucocorticoids potentiate the action of catecholamines in some tissues by increasing the expression of beta-adrenergic receptors. Paradoxically, glucocorticoids suppress and adrenalectomy enhances BAT thermogenesis. To further study the reasons for this discrepancy, we assessed the effects of methylprednisolone administration, adrenalectomy, and adrenalectomy with corticosterone replacement on adenylate cyclase activity in BAT and on beta-adrenergic receptor density in lungs and BAT of rats. In lungs, the density of the beta 2-adrenergic receptor subtype increases after methylprednisolone administration and decreases after adrenalectomy. There was no change in BAT receptor density, but isoproterenol-, NaF-, and forskolin-stimulated adenylate cyclase activity was reduced by 20-35% after methylprednisolone treatment. There was a two- to threefold increase in adenylate cyclase activity after adrenalectomy, which was reversed by corticosterone administration. These data suggest that one mechanism by which glucocorticoids regulate BAT thermogenesis is by modulating the beta-adrenergic pathway at the level of adenylate cyclase activation.

Modulation of receptors and adenylate cyclase activity during sucrose feeding, food deprivation, and cold exposure.

Thermogenesis in brown adipose tissue (BAT) serves as a regulator of body temperature and weight maintenance. Thermogenesis can be stimulated by catecholamine activation of adenylate cyclase through the beta-adrenergic receptor. To investigate the effects of sucrose feeding, food deprivation, and cold exposure on the beta-adrenergic pathway, adenylate cyclase activity and beta-adrenergic receptors were assessed in rat BAT after 2 wk of sucrose feeding, 2 days of food deprivation, or 2 days of cold exposure. beta-Adrenergic receptors were identified in BAT using [125I]iodocyanopindolol. Binding sites had the characteristics of mixed beta 1- and beta 2-type adrenergic receptors at a ratio of 60/40. After sucrose feeding or cold exposure, there was the expected increase in BAT mitochondrial mass as measured by total cytochrome-c oxidase activity but a decrease in beta-adrenergic receptor density due to a loss of the beta 1-adrenergic subtype. This BAT beta-adrenergic receptor downregulation was tissue specific, since myocardial beta-adrenergic receptors were unchanged with either sucrose feeding or cold exposure. In contrast, food deprivation did not alter BAT beta-adrenergic receptor density. Forskolin-stimulated adenylate cyclase activity increased in BAT after sucrose feeding or cold exposure but not after food deprivation. The ratio of isoproterenol-stimulated to forskolin-stimulated adenylate cyclase activity decreased in the sucrose-fed and cold-exposed rats but not in the food-deprived rats. These data suggest that in BAT, sucrose feeding or cold exposure result in downregulation of beta-adrenergic receptors and that isoproterenol-stimulated adenylate cyclase activity was limited by receptor availability.

Genetic transformation in the methanogen Methanococcus voltae PS.

Mutations causing requirements for histidine, purine, and vitamin B12 were obtained in strain PS of Methanococcus voltae (archaebacteria) upon irradiation with UV or gamma rays. The first two mutations were shown to revert at low frequencies and were used to demonstrate the occurrence of transformation with homologous, wild-type DNA. The transformation rates obtained for these presumably chromosomal markers were in the range of 2 to 100 transformants per microgram of DNA. Mutants resistant to 2-bromoethanesulfonate and to 5-methyl-DL-tryptophan were also isolated.

A grading model for stress urinary incontinence.

A classification of stress incontinence is described. The stress test makes it possible to diagnose sphinctero-urethral incompetence and to identify the cases that are objectively negligible. Thus, surgical treatment can be limited to those who require it.

Desensitization and resensitization of beta-adrenergic receptors in a smooth muscle cell line.

Exposure to the beta-adrenergic agonist, metaproterenol, elicits extensive receptor loss and desensitization of adenylate cyclase activity in the hamster DDT1, MF-2 cell line. The reappearance of beta-adrenergic receptors and restoration of adenylate cyclase activity were investigated. Receptor reappearance was investigated under conditions in which lost receptors were not detectable either on the cell surface or within the cell. Exposure to metaproterenol resulted in a 3-5-fold decrease in beta-adrenergic receptor affinity for agonist, an 85% reduction in beta-adrenergic receptor number per cell, and a 65% reduction in isoproterenol-stimulated adenylate cyclase activity without any change in NaF-stimulated enzyme activity. The rate of reappearance of the lost receptors was proportional to the concentration of metaproterenol to which the cells were initially exposed. Metaproterenol, at a concentration of 250 microM, induced long-term receptor loss which required 16 days in fresh media devoid of metaproterenol before the full complement of receptors reappeared. This prolonged receptor loss may be due to residual metaproterenol; however, the resensitization of isoproterenol-stimulated adenylate cyclase activity was restored 2 days after removal of metaproterenol. The lag period for the reappearance of receptors was shortened by incubation with either the beta-adrenergic antagonist, nadalol, or the glucocorticoid, methylprednisolone. Both pharmaceuticals reversed receptor down-regulation and up-regulated receptor number in control cells, although the extent and time course of restoration were different. These data suggest that the process of resensitization in DDT1 cells involves rapid restoration of adenylate cyclase activity and a slower reappearance of receptors over a time period of six population doublings.

Methanogenic cleavage of acetate by lysates of Methanosarcina barkeri.

Cell lysates of acetate-grown Methanosarcina barkeri 227 were found to cleave acetate to CH4 and CO2. The aceticlastic reaction was identified by using radioactive methyl-labeled acetate. Cell lysates decarboxylated acetate in a nitrogen atmosphere, conserving the methyl group in methane. The rate of methanogenesis from acetate in the cell lysates was comparable to that observed with whole cells. Aceticlastic activity was found in the particulate fraction seperate from methylcoenzyme M methylreductase activity, which occurs in the soluble fraction. Pronase treatment eliminated methylcoenzyme M methylreductase activity in lysates and stimulated aceticlastic activity, indicating the aceticlastic activity was not derived from unbroken cells, which are unaffected by proteolytic treatment.

Levels of coenzyme F420, coenzyme M, hydrogenase, and methylcoenzyme M methylreductase in acetate-grown Methanosarcina.

Methanosarcina barkeri strain 227 maintained on an acetate medium for 2 years was found to possess hydrogenase, methylcoenzyme M methylreductase, coenzyme F420, and coenzyme M. The levels of these constituents in acetate-grown cells were similar to those found in cells of the same strain grown on methanol or hydrogen and carbon dioxide.

Methanogenesis from acetate: enrichment studies.

An acetate enrichment culture was initiated by inoculating anaerobic sludge from a mesophilic methane digestor into a mineral salts medium with calcium acetate as the sole carbon and energy source. This enrichment was maintained indefinitely by weekly transfer into medium of the same composition. A study of this enrichment disclosed an unexpected age-dependent inhibition of methanogenesis by H2 and formate which apparently differed from the inhibition by chloroform and benzyl viologen. This age-dependent inhibition indicated that microbial interactions of the mixed enrichment population may play a regulatory role in methane formation. Futhermore, stimulation of methanogenesis in the acetate enrichment by addition of yeast extract showed a nutrient limitation which indicated that syntrophic interactions leading to formation of growth factors may also occur. A model is presented to illustrate the possible interrelationships between methanogenic and nonmethanogenic bacteria in their growth and formation of methane and carbon dioxide from acetate.

Studies on an acetate-fermenting strain of Methanosarcina.

An acetate-fermenting strain of Methanosarcina was isolated from an acetate enrichment culture inoculated with anaerobic sludge from a waste treatment digestor. In pure culture, this organism fermented acetate in the absence of added hydrogen at rates comparable in magnitude to those found in digestor systems. This rate was significantly higher than previously obtained for pure cultures of this genus. Mineral components of yeast extract were highly stimulatory for cultures growing on methanol. Comparable stimulation was not observed for cultures growing on acetate. Labeling studies indicated that acetate was converted to methane and CO2 as predicted by previous studies on mixed cultures. Total oxidation or reduction of acetate was not the mechanism of conversion of acetate to methane by the pure culture. The ability of this strain to form colonies or to produce methane from acetate was apparently influenced by the choice of substrate and conditions used for growing the inoculum.