NeuroThera® Efficacy and Safety Trial-3 (NEST-3): a double-blind, randomized, sham-controlled, parallel group, multicenter, pivotal study to assess the safety and efficacy of transcranial laser therapy with the NeuroThera® Laser System for the treatment of acute ischemic stroke within 24 h of stroke onset.

RATIONALE: Transcranial laser therapy is undergoing clinical trials in patients with acute ischemic stroke. The NeuroThera® Efficacy and Safety Trial-1 was strongly positive for 90-day functional benefit with transcranial laser therapy, and post hoc analyses of the subsequent NeuroThera® Efficacy and Safety Trial-2 trial suggested a meaningful beneficial effect in patients with moderate to moderately severe ischemic stroke within 24 h of onset. These served as the basis for the NeuroThera® Efficacy and Safety Trial-3 randomized controlled trial. AIM: The purpose of this pivotal study was to demonstrate safety and efficacy of transcranial laser therapy with the NeuroThera® Laser System in the treatment of subjects diagnosed with acute ischemic stroke. DESIGN: NeuroThera® Efficacy and Safety Trial-3 is a double-blind, randomized, sham-controlled, parallel group, multicenter, pivotal study that will enroll 1000 subjects at up to 50 sites. All subjects will receive standard medical management based on the American Stroke Association and European Stroke Organization Guidelines. In addition to standard medical management, both groups will undergo the transcranial laser therapy procedure between 4·5 and 24 h of stroke onset. The study population will be randomized into two arms: the sham control group will receive a sham transcranial laser therapy procedure and the transcranial laser therapy group will receive an active transcranial laser therapy procedure. The randomization ratio will be 1:1 and will be stratified to ensure a balanced subject distribution between study arms. STUDY OUTCOMES: The primary efficacy end point is disability at 90 days (or the last rating), as assessed on the modified Rankin Scale, dichotomized as a success (a score of 0-2) or a failure (a score of 3 to 6).

A comparative evaluation of ultrasound molecular imaging, perfusion imaging, and volume measurements in evaluating response to therapy in patient-derived xenografts.

Most pre-clinical therapy studies use the change in tumor volume as a measure for disease response. However, tumor size measurements alone may not reflect early changes in tumor physiology that occur as a response to treatment. Ultrasonic molecular imaging (USMI) and Dynamic Contrast Enhanced-Perfusion Imaging (DCE-PI) with ultrasound are two attractive alternatives to tumor volume measurements. Since these techniques can provide information prior to the appearance of gross phenotypic changes, it has been proposed that USMI and DCE-PI could be used to characterize response to treatment earlier than traditional methods. This study evaluated the ability of tumor volume measurements, DCE-PI, and USMI to characterize response to therapy in two different types of patient-derived xenografts (known responders and known non-responders). For both responders and non-responders, 7 animals received a dose of 30 mg/kg of MLN8237, an investigational aurora-A kinase inhibitor, for 14 days or a vehicle control. Volumetric USMI (target integrin:α av ß3) and DCE-PI were performed on day 0, day 2, day 7, and day 14 in the same animals. For USMI, day 2 was the earliest point at which there was a statistical difference between the untreated and treated populations in the responder cohort (Untreated: 1.20 ±â€…0.53 vs. Treated: 0.49 ±â€…0.40; p < 0.05). In contrast, statistically significant differences between the untreated and treated populations as detected using DCE-PI were not observed until day 14 (Untreated: 0.94 ±â€…0.23 vs. Treated: 1.31 ±â€…0.22; p < 0.05). Volume measurements alone suggested no statistical differences between treated and untreated populations at any readpoint. Monitoring volumetric changes is the "gold standard" for evaluating treatment in pre-clinical studies, however, our data suggests that volumetric USMI and DCE-PI may be used to earlier classify and robustly characterize tumor response.

WE-C-218-01: Ultrasound Contrast Agents.

Medical ultrasound has long been used in clinical applications both as a primary modality and as a supplement to other diagnostic procedures. The basis for ultrasound imaging is the transmission of high frequency (megaHertz) sound waves that propagate through tissue. These sound waves backscatter from the interfaces between tissue components with different acoustic properties and are detected by the imaging system, allowing the creation of images based on tissue characteristics and spatial location. Thus, traditional ultrasound has focused primarily on the imaging of anatomical structures and analysis of blood flow in large vessels. Unfortunately, blood is a weak scatterer, which can make vascular diagnostic applications (example: echocardiography) challenging especially with larger patients. Contrast agents help to improve on this shortcoming by enhancing the visualization of blood flow, thus improving the quality of diagnostics. The use of contrast agents for ultrasound was first reported in 1968 when Gramiak and Shah discovered that there was an increased backscatter of ultrasound caused by injected microbubbles. This is because the mismatch in acoustic impedance (a function of an object's density and compressibility) between the microbubble gas core and blood (or tissue) is several orders of magnitude, which results in substantially higher scattering from a bubble than an equivalent volume of tissue or blood. Additionally, microbubbles oscillate in response to an ultrasound field, and respond non-linearly to acoustic pulses even at low energies, unlike tissue. The non-linear property of microbubbles in an ultrasound field allows for the use of various pulsing and signal processing strategies to detect the backscattered signal from contrast agents and segment it from tissue, thus providing a high contrast-to- noise ratio. Due to these unique acoustic properties, a clinical ultrasound system can detect even single microbubble contrast agents, providing exquisite sensitivity and the ability to perform advanced diagnostic procedures. Over the last several decades, ultrasound contrast agents have been improved for enhanced stability and increased persistence times. Although preclinical studies as well as clinical use in Europe and Asia strongly suggest that the use of contrast ultrasound can substantially improve diagnostic capabilities in both cardiology and radiology applications, contrast use in the US is still very limited. Obstacles to the widespread use of microbubbles include safety concerns, the need for optimization of approaches for contrast use, and general understanding of their potential by physicians. This course covers the basic principles of contrast agents used in ultrasound imaging including their stability, shell properties and their behavior within an acoustic field. In addition, we will cover many new techniques that are being evaluated in preclinical studies including: p er fus ion-based techniques, molecular imaging, gene therapy, drug delivery, and acoustic angiography. Finally, basic safety concerns and biological effects will be reviewed. LEARNING OBJECTIVES: 1. Understand the basic principles of ultrasound contrast agents a. What are microbubble contrast agents? b. Properties of microbubbles c. Safety concerns and biological effects 2. Understand basic contrast imaging techniques a. Harmonic and suharmonic imaging techniques b. Pulse inversion techniques 3. Understand the use of contrast agents in various vascular applications a. Traditional methods (Cardiovascular, Abdominal) b. Advanced perfusion imaging techniques 4. Understand the role of contrast agents in preclinical applications a. Ultrasound molecular imaging b. Gene therapy c. Drug delivery d. Acoustic angiography.

Factors affecting the survival of Bradyrhizobium applied in liquid cultures to soya bean [Glycine max (L.) Merr.] seeds.

AIMS: To determine the impact of medium composition, bacterial strain, trehalose accumulation, and relative humidity during seed storage on the survival of Bradyrhizobium japonicum on soya bean [Glycine max (L.) Merr.] seeds. METHODS AND RESULTS: Bacteria in liquid cultures were applied to seeds, and the number of survivors was quantified after 2, 24, 48, or 96 h. Addition of yeast extract to a defined medium increased on-seed survival 50- to 80-fold. Addition of 40 mmol l(-1) of NaCl to the medium doubled or tripled the accumulation of trehalose in cells and increased survival several fold, and the addition of both salt and trehalose had an additive effect. There was a threefold difference among strains in survival, and survival of the various strains was significantly correlated with differences in the accumulation of trehalose. The correlation between trehalose accumulation by bacteria and survival was also highly significant in other experiments. Studies in controlled humidity environments showed 100-fold or more differences in survival. CONCLUSIONS: The consistently significant correlation of trehalose content of cells with survival on seed suggests that trehalose is an important component of the survival mechanisms. When some of the factors (salt and trehalose in the medium plus humidity control) were studied in combination, several 100-fold increases in survival of bacteria on seeds were recorded. SIGNIFICANCE AND IMPACT OF THE STUDY: It is possible by manipulation of several parameters--strain selection, salt and trehalose content of the medium, control of relative humidity--to achieve substantial improvements in survival of Bradyrhizobium on soya bean seeds.

Ga-As (808 nm) laser irradiation enhances ATP production in human neuronal cells in culture.

OBJECTIVE: The aim of the present study was to investigate whether Ga-As laser irradiation can enhance adenosine triphosphate (ATP) production in normal human neural progenitor (NHNP) cells in culture. METHODS: NHNP were grown in tissue culture and were treated by Ga-As laser (808 nm, 50 mW/cm(2), 0.05 J/cm(2)), and ATP was determined at 10 min after laser application. RESULTS: The quantity of ATP in laser-treated cells was 7513 +/- 970 units, which was significantly higher (p < 0.05) than the non-treated cells, which comprised 3808 +/- 539 ATP units. CONCLUSION: Laser application to NHNP cells significantly increases ATP production in these cells. These findings may explain the beneficial effects of low-level laser therapy (LLLT) in stroked rats. Tissue culture of NHNP cells might offer a good model to study the mechanisms associated with promotion of ATP production in the nervous system by LLLT.

Three enzymes for trehalose synthesis in Bradyrhizobium cultured bacteria and in bacteroids from soybean nodules.

alpha,alpha-Trehalose is a disaccharide accumulated by many microorganisms, including rhizobia, and a common role for trehalose is protection of membrane and protein structure during periods of stress, such as desiccation. Cultured Bradyrhizobium japonicum and B. elkanii were found to have three enzymes for trehalose synthesis: trehalose synthase (TS), maltooligosyltrehalose synthase (MOTS), and trehalose-6-phosphate synthetase. The activity level of the latter enzyme was much higher than those of the other two in cultured bacteria, but the reverse was true in bacteroids from nodules. Although TS was the dominant enzyme in bacteroids, the source of maltose, the substrate for TS, is not clear; i.e., the maltose concentration in nodules was very low and no maltose was formed by bacteroid protein preparations from maltooligosaccharides. Because bacteroid protein preparations contained high trehalase activity, it was imperative to inhibit this enzyme in studies of TS and MOTS in bacteroids. Validamycin A, a commonly used trehalase inhibitor, was found to also inhibit TS and MOTS, and other trehalase inhibitors, such as trehazolin, must be used in studies of these enzymes in nodules. The results of a survey of five other species of rhizobia indicated that most species sampled had only one major mechanism for trehalose synthesis. The presence of three totally independent mechanisms for the synthesis of trehalose by Bradyrhizobium species suggests that this disaccharide is important in the function of this organism both in the free-living state and in symbiosis.

Effect of trehalose on survival of Bradyrhizobium japonicum during desiccation.

AIMS: A major reason for the ineffectiveness of legume inoculants in the field is the rapid death of rhizobia because of desiccation. The major purpose of this study was to identify conditions under which alpha,alpha-trehalose would improve survival of Bradyrhizobium japonicum during desiccation. METHODS AND RESULTS: Trehalose was added to cultures just prior to desiccation or was supplied to bacteria during the 6-day growth period. A wide variety of trehalose concentrations was tested. Trehalose added to cultures at the time of desiccation improved survival slightly, but trehalose loading during growth was much more effective in protection against desiccation. Growth of bacteria with 3 mmol l-1 trehalose increased trehalose concentration in cells by about threefold and increased survival of cells placed on soya bean [Glycine max (L.) Merr.] seeds by two- to four-fold after 2 or 24 h. Average of overall results indicate that growth of bacteria with trehalose in the medium resulted in a 294% increase in survival after 24 h of desiccation. The concentration of trehalose in cells was very highly correlated with survival of bacteria. When trehalose-loaded cells were suspended in buffer or water, 60-85% of cellular trehalose was lost in about 1 h and, in spite of these losses, survival during desiccation was not reduced. CONCLUSIONS: Accumulation of trehalose in the cytoplasm is critical to the survival of B. japonicum during desiccation. Increasing the periplasmic concentration of trehalose is also beneficial but is not so critical as the concentration of trehalose in the cytoplasm. Because B. japonicum cannot utilize trehalose as a carbon source, cells can be loaded with trehalose by providing the disaccharide during the growth period. SIGNIFICANCE AND IMPACT OF THE STUDY: Although it may not be practical to use trehalose as a carbon source in inoculant production, it may be possible to engineer greater trehalose accumulation in rhizobia. Trehalose concentration in cells should be a useful predictor of survival during desiccation.

Biosynthesis of trehalose from maltooligosaccharides in Rhizobia.

Previously, the enzymes for trehalose synthesis that are present in Escherichia coli were demonstrated in Bradyrhizobium japonicum and B. elkanii. An alternative mechanism recently reported for the synthesis of trehalose from maltooligosaccharides was considered based on the fact that high concentrations of sugars in liquid culture stimulated the accumulation of trehalose. An assay for the synthesis of trehalose from maltooligosaccharides using crude, gel-filtered protein preparations was developed. Analysis of a variety of the Rhizobiaceae indicates that the "maltooligosaccharide mechanism" is present in B. japonicum, B. elkanii, Rhizobium sp. NGR234, Sinorhizobium meliloti, R. tropici A, R. leguminosarum bv viciae, R. I. bv trifolii, and Azorhizobium caulinodans. Synthesis of trehalose from maltooligosaccharide could not be detected in R. tropici B or R. etli.

Simultaneous extraction and derivatization of carbohydrates from green plant tissues for analysis by gas-liquid chromatography.

Simultaneous extraction and derivatization of carbohydrates was performed by mixing dry ground plant tissue with derivatization reagents in pyridine; trimethylsilyl derivatives were analyzed by gas-liquid chromatography. This "direct analysis" was compared to analysis of samples prepared by exhaustive ethanol extraction of the same ground plant tissues. Comparisons included leaf blades from apple, grape, corn, and tomato and leaf blade, petiole, stem, and pod tissues from soybean plants. Direct analysis gave superior quantification of sucrose, glucose, and fructose because of sucrose hydrolysis during ethanol extraction. Sucrose hydrolysis was highly variable among plant species and use of hot ethanol at the first extraction step reduced sucrose hydrolysis but did not always abolish it. Sucrose hydrolysis was probably due to the activity of hydrolytic enzymes in 75% ethanol at room temperature. Direct analysis was inferior for the quantification of cyclitols in the fibrous tissues of soybean but provided acceptable results for cyclitol analysis in leaf blade tissue. When the time for extraction/reaction was extended from 40 to 60 min, some improvement in recovery of cyclitols was observed, but recovery remained 10 to 20% below that obtained with exhaustive ethanol extraction. Mannitol was vacuum infiltrated into the five types of leaf tissue and recovery averaged 100% by the direct method relative to ETOH extraction for apple, grape, corn, and soybean leaves but was only 76% for tomato leaves. Direct analysis provides very large time savings and is clearly the method of choice when the analysis of large numbers of samples of plant tissues for carbohydrate composition is required.

The structure of a novel polysaccharide produced by Bradyrhizobium species within soybean nodules.

Certain strains of Bradyrhizobium japonicum and B. elkanii produce a polysaccharide within the root nodules of their legume host, soybean. These nodule polysaccharides (NPSs) were isolated and characterized. The NPS produced by B. elkanii strains proved to be identical in glycosyl composition and linkages to the extracellular polysaccharide (EPS) of this species indicating that the NPS and EPS for B. elkanii have identical structures (W.F. Dudman, Carbohydr. Res., 66 (1978) 9-23), [formula: see text] However, the structure of the NPS from B. japonicum proved to be quite different from that of its EPS. Methylation analysis of this NPS showed that it consists of 3-linked Gal, 3-linked Rha, 2,4-linked Rha, 4-linked Rha, and terminal 2-O-methyl GlcA in a 1:1:1:1:1 ratio. Stereochemical configurations of the glycosyl residues were determined by the preparation and analysis of trimethylsilyl (Me3Si) (-)-2-butyl glycosides. NMR spectroscopy (both 1H and 13C) showed that the Gal residue is alpha-linked, while all the other glycosyl residues are beta-linked. Oligosaccharides produced by periodate oxidation-Smith degradation were purified, as were oligosaccharides produced by partial acid hydrolysis. Characterization of the Smith degradation products by methylation analysis. NMR spectroscopy, electrospray-mass spectrometry, and characterization of the partial acid hydrolysate oligosaccharides showed that the repeating oligosaccharide unit of the NPS has the structure, [formula: see text]

Fate of Nodule-Specific Polysaccharide Produced by Bradyrhizobium japonicum Bacteroids.

A polysaccharide produced by Bradyrhizobium japonicum bacteroids in nodules (NPS) on soybean (Glycine max [L.] Merr.) roots is different in composition and structure from the extracellular polysaccharide produced in culture by this organism. Isogenic strains either capable or incapable of NPS synthesis supported similar rates of plant growth and nitrogenase activity, indicating that polysaccharide deposition was not detrimental. The possibility that NPS may have some protective or nutritional role for bacteroids was considered. Analysis of disintegrating nodules over periods of 1 to 3 months indicated greater recovery of viable bacteria from NPS+ nodules prior to the breakdown of NPS. During and after the breakdown of NPS, the decline in viable bacteria was similar for NPS+ and NPS- strains. Bacteroid destruction in senescing nodules may be accelerated by exposure to proteolytic enzymes in host cytoplasm; however, highly purified NPS had no significant effect on the in vitro activity of partially purified proteases, so protection of bacteroids via this mechanism is unlikely. B. japonicum USDA 438 did not utilize NPS as a carbon source for growth in liquid culture. In vitro assays of NPS depolymerase activity in cultured bacteria and bacteroids were negative using a variety of strains, all of which contained extracellular polysaccharide depolymerase. It seems highly unlikely that B. japonicum can utilize the polysaccharide it synthesizes in nodules, and NPS breakdown in senescing nodules is probably caused by saprophytic fungi.

Factors Influencing the Synthesis of Polysaccharide by Bradyrhizobium japonicum Bacteroids in Field-Grown Soybean Nodules.

Certain strains of Bradyrhizobium japonicum produce large quantities of polysaccharide in soybean (Glycine max (L.) Merr.) nodules, and nodule polysaccharide (NPS) is different from that produced in culture. A previous survey of field-grown plants showed highly variable levels of NPS among field sites. To obtain clues about the possible function of NPS, we conducted two additional surveys of field-grown plants. The amount of polysaccharide in bulk samples of nodules was not associated with soil type, texture, slope, drainage, or any of the measured soil chemical properties except pH and [Ca]. Correlations with pH and [Ca] were positive and highly significant for two independent surveys involving a total of 77 sites in two years. In a preliminary comparison of high and low levels of Ca supplied to soybean plants grown in silica sand in a greenhouse, a high level of Ca (200 mg of Ca liter) increased the NPS level and increased the Ca content of the polysaccharide fraction. B. japonicum isolates from 450 nodules collected at 10 field sites in 1993 were used to form nodules on soybean plants grown in sand culture in a greenhouse in order to examine bacterial phenotype under controlled conditions. Results showed that the NPS level in the bulk nodule sample from any given site was a function of the proportion of nodule occupants that were capable of NPS synthesis. Thus, a higher soil pH and/or [Ca] may positively influence the survival of B. japonicum capable of synthesis of the nodule-specific polysaccharide.

Labeling of Carbon Pools in Bradyrhizobium japonicum and Rhizobium leguminosarum bv viciae Bacteroids following Incubation of Intact Nodules with CO(2).

The aim of the work reported here was to ascertain that the patterns of labeling seen in isolated bacteroids also occurred in bacteroids in intact nodules and to observe early metabolic events following exposure of intact nodules to (14)CO(2). Intact nodules of soybean (Glycine max L. Merr. cv Ripley) inoculated with Bradyrhizobium japonicum USDA 110 and pea (Pisum sativum L. cv Progress 9) inoculated with Rhizobium leguminosarum bv viciae isolate 128C53 were detached and immediately fed (14)CO(2) for 1 to 6 min. Bacteroids were purified from these nodules in 5 to 7 min after the feeding period. In the cytosol from both soybean and pea nodules, malate had the highest radioactivity, followed by citrate and aspartate. In peas, asparagine labeling equaled that of aspartate. In B. japonicum bacteroids, malate was the most rapidly labeled compound, and the rate of glutamate labeling was 67% of the rate of malate labeling. Aspartate and alanine were the next most rapidly labeled compounds. R. leguminosarum bacteroids had very low amounts of (14)C and, after a 1-min feeding, malate contained 90% of the radioactivity in the organic acid fraction. Only a trace of activity was found in aspartate, whereas the rate of glutamate and alanine labeling approached that of malate after 6 min of feeding. Under the conditions studied, malate was the major form of labeled carbon supplied to both types of bacteroids. These results with intact nodules confirm our earlier results with isolated bacteroids, which showed that a significant proportion of provided labeled substrate, such as malate, is diverted to glutamate. This supports the conclusion that microaerobic conditions in nodules influence carbon metabolism in bacteroids.

Induction of Glutamine Synthetase Activity in Nonnodulated Roots of Glycine max, Phaseolus vulgaris, and Pisum sativum.

Nitrate or ammonium fertilization significantly increased glutamine synthetase (GS) activity in nonnodulated roots of French bean (Phaseolus vulgaris), soybean (Glycine max), and pea (Pisum sativum). Western analysis revealed substantial GS antibody-positive protein in root extracts that had minimal GS activity, indicating that an inactive form of GS may be present in nonfertilized plants.

Formation of Novel Polysaccharides by Bradyrhizobium japonicum Bacteroids in Soybean Nodules.

Certain strains of Bradyrhizobium japonicum form a previously unknown polysaccharide in the root nodules of soybean plants (Glycine max (L.) Merr.). The polysaccharide accumulates inside of the symbiosome membrane-the plant-derived membrane enclosing the bacteroids. In older nodules (60 days after planting), the polysaccharide occupies most of the symbiosome volume and symbiosomes become enlarged so that there is little host cytoplasm in infected cells. The two different groups of B. japonicum which produce different types of polysaccharide in culture produce polysaccharides of similar composition in nodules. Polysaccharide formed by group I strains (e.g., USDA 5 and USDA 123) is composed of rhamnose, galactose, and 2-O-methylglucuronic acid, while polysaccharide formed by group II strains (e.g., USDA 31 and USDA 39) is composed of rhamnose and 4-O-methylglucuronic acid. That the polysaccharide is a bacterial product is indicated by its composition plus the fact that polysaccharide formation is independent of host genotype but is dependent on the bacterial genotype. Polysaccharide formation in nodules is common among strains in serogroups 123, 127, 129, and 31, with 27 of 39 strains (69%) testing positive. Polysaccharide formation in nodules is uncommon among other B. japonicum serogroups, with only 1 strain in 18 (6%) testing positive.

Effects of Salt Stress on Amino Acid, Organic Acid, and Carbohydrate Composition of Roots, Bacteroids, and Cytosol of Alfalfa (Medicago sativa L.).

Ethanol-soluble organic acid, carbohydrate, and amino acid constituents of alfalfa (Medicago sativa) roots and nodules (cytosol and bacteroids) have been identified by gas-liquid chromatography and high performance liquid chromatography. Among organic acids, citrate was the predominant compound in roots and cytosol, with malonate present in the highest concentration in bacteroids. These two organic acids together with malate and succinate accounted for more than 85% of the organic acid pool in nodules and for 97% in roots. The major carbohydrates in roots, nodule cytosol, and bacteroids were (descending order of concentration): sucrose, pinitol, glucose, and ononitol. Maltose and trehalose appeared to be present in very low concentrations. Asparagine, glutamate, alanine, gamma-aminobutyrate, and proline were the major amino acids in cytosol and bacteroids. In addition to these solutes, serine and glutamine were well represented in roots. When alfalfa plants were subjected to 0.15 m sodium chloride stress for 2 weeks, total organic acid concentration in nodules and roots were depressed by more than 40%, whereas lactate concentration increased by 11, 27, and 94% in cytosol, roots, and bacteroids, respectively. In bacteroids, lactate became the most abundant organic acid and might contribute partly to the osmotic adjustment. On the other hand, salt stress induced a large increase in the amino acid and carbohydrate pools. Within the amino acids, proline showed the largest increase, 11.3-, 12.8-, and 8.0-fold in roots, cytosol, and bacteroids, respectively. Its accumulation reflected an osmoregulatory mechanism not only in roots but also in nodule tissue. In parallel, asparagine concentration was greatly enhanced; this amide remained the major nitrogen solute and, in bacteroids, played a significant role in osmoregulation. On the contrary, the salt treatment had a very limited effect on the concentration of other amino acids. Among carbohydrates, pinitol concentration was increased significantly, especially in cytosol and bacteroids (5.4- and 3.4-fold, respectively), in which this cyclitol accounted for more than 35% of the total carbohydrate pool; pinitol might contribute to the tolerance to salt stress. However, trehalose concentration remained low in both nodules and roots; its role in osmoregulation appeared unlikely in alfalfa.

Increased Accumulation of Trehalose in Rhizobia Cultured under 1% Oxygen.

The growth of rhizobia under 1% O(2) induced the accumulation of alpha,alpha-trehalose, and the effect of low O(2) was independent of medium composition and Rhizobium species. Trehalose concentration in cells declined rapidly when microaerobic cultures were supplied with 21% O(2). Trehalose formation in nodules may be induced by the microaerobic environment.

Periplasmic metabolism of glutamate and aspartate by intact Bradyrhizobium japonicum bacteroids.

In studies on the uptake and metabolism of [14C]glutamate by Bradyrhizobium japonicum bacteroids we found that, in the presence of unlabeled malate, succinate or alpha-ketoglutarate, substantial label was recovered in alpha-ketoglutarate in the reaction mixtures. As much as 30% of the total 14C supplied could be found in alpha-ketoglutarate in the reaction mixtures after 30 min and this occurred in the absence of detectable labeling of alpha-ketoglutarate in the cells. The labeling of alpha-ketoglutarate was almost completely inhibited by aminooxyacetate (aminotransferase inhibitor). Direct assay of aspartate aminotransferase in intact bacteroids was possible in the presence of very dilute Triton X-100 (less than or equal to 0.02%, w/v). The response of the aminotransferase to detergent was similar to the response of phosphodiesterase, a periplasmic marker, and different from malate dehydrogenase and beta-hydroxybutyrate dehydrogenase, cytoplasmic markers. Comparison of maximum enzyme activity assayable with intact bacteroids and maximum activity in sonicated bacteroids indicated that about half of the total cellular aminotransferase activity was accessible to the external medium. The combined labeling and enzyme assay results indicated that B. japonicum bacteroids have a capability for transamination in the periplasmic space. Although this may not be important in the transfer of reducing equivalents from host cytoplasm to bacteroids in nodules, the transamination capability may facilitate the acquisition of metabolites by free-living bacteria.

As young as you feel: age identification among the elderly.

1. Understanding and integrating an individual's perceptions about age and health status is fundamental to making accurate assessments and implementing interventions that are personally acceptable and clinically appropriate. 2. The concept of chronological age has limited validity for the explanation of behavior because it wrongly assumes homogeneity in individual lifestyles among age cohorts. 3. Older adults tend to perceive themselves as mentally and physically younger than their chronological age. 4. Nurses who interact with individuals in a manner based solely on their chronological age may exhibit disrespect and underestimate the potential for the client's involvement in self-care activities.