The mindful shield: The effects of mindfulness training on resilience and leadership in military leaders.

OBJECTIVES: The purpose of this study was to address military leader perceptions of their resilience, transformational leadership behaviors, and leadership effectiveness before and after experiencing Mindfulness-Based Attention Training (MBAT). METHODS: Participants were formal and informal leaders in the Kansas Air National Guard. The study used a mixed-methods sequential exploratory design. Phase I involved analyzing pretest and posttest results obtained from a Jha Lab study for three self-report assessments in an intervention group (n = 36) vs a control group (n = 37). The qualitative data in phase II was obtained from individual interviews of participants (n = 12) following the Jha Lab study. RESULTS: The phase I quantitative results confirmed the null hypotheses-no significant differences found-for all research questions. Phase II resulted in eight thematic codes, six of which were central to the experiences described by participants (Halting, Sensing, Being, Shielding, Considering, and Engaging) and two that were not (Obstructing, and Escaping). CONCLUSIONS: The key finding was that the descriptions of mindful thoughts and behaviors were consistent across participants indicating that MBAT accurately presents mindfulness during the course and the training had positive effects on participant mindfulness, primarily in the areas of being present to self, shielding the self through reperceiving, and then consciously altering behavior based on the new perspective. Results should direct future resiliency course development, leadership course curricula, and aid understanding of how leaders mentally conceptualize stress, incorporate resilient behaviors and then apply that knowledge to their own leadership behaviors.

Enzymes from high-temperature microorganisms.

Enzymes derived from microorganisms growing at extreme temperatures are of biotechnological use as highly thermostable biocatalysts and should provide insight into the intrinsic basis of protein stability. So far, only DNA polymerases from these organisms have been put to commercial use, although the application of other classes of highly thermostable enzymes is being considered. Problems in the cultivation of high-temperature microorganisms and in the production of their enzymes still hampers progress in this field.

Glycosyl hydrolases from hyperthermophilic microorganisms.

Glycosyl hydrolases from hyperthermophiles are, thus far, the most widely studied enzyme class from these organisms. Not only are there many biotechnological opportunities for these enzymes, but the rapidly increasing amount of information about their genetic, biochemical and biophysical characteristics (recently genomic sequencing data for both P. furiosus and P. horikoshi have been published on the Internet) make them ideal candidates for the study of biocatalysis and protein thermostability at extremely high temperatures.

Finding and using hyperthermophilic enzymes.

Recent developments have enhanced the prospects for the discovery of hyperthermophilic enzymes. This is important because the intrinsic basis underlying the extraordinary thermostability of hyperthermophilic enzymes has yet to be revealed, and so engineering this characteristic into less thermophilic enzymes is not possible at this time. Successful efforts to clone and express the genes encoding hyperthermophilic enzymes in mesophilic hosts have improved the availability of high-temperature biocatalysts. The remaining task is the identification of opportunities to make strategic use of the thermoactivity and thermostability of hyperthermophilic enzymes.

Homomultimeric protease in the hyperthermophilic bacterium Thermotoga maritima has structural and amino acid sequence homology to bacteriocins in mesophilic bacteria.

A novel homomultimeric protease (> 669 kDa), based on 31 kDa subunits, was purified from cell extracts of the hyperthermophilic bacterium Thermotoga maritima. This protease exhibits activity toward chymotrypsin and trypsin substrates, optimally at 90 degrees C and pH 7.1, and has a half-life of 36 min at 95 degrees C. Transmission electron microscopy established that the protease consists of a large globular assembly which appears circular from the front view. The function of this protease in T. maritima remains unclear, although putative homologs include a 29 kDa antigen from Mycobacterium tuberculosis and a 31 kDa monomer of a high molecular weight bacteriocin produced by Brevibacterium linens [Valdes-Stauber, N. and Scherer, S. (1996) Appl. Environ. Microbiol. 62, 1283-1286]. The relationship of these mesophilic proteins to the T. maritima protease suggests that their antibacterial activity may involve elements of proteolysis, and raises the prospect for antimicrobial ecological strategies in hyperthermophilic niches.

Cultivation Techniques for Hyperthermophilic Archaebacteria: Continuous Culture of Pyrococcus furiosus at Temperatures near 100 degrees C.

A system which allows continuous cultivation of hyperthermophilic archaebacteria at temperatures approaching 100 degrees C has been developed. Continuous cultivation of the hyperthermophile Pyrococcus furiosus was carried out with this system; the resulting dilution rate and gas production profiles are discussed.

The hyperthermophilic archaebacterium, Pyrococcus furiosus. Development of culturing protocols, perspectives on scaleup, and potential applications.

From this brief discussion, it is clear that there are many obstacles to overcome before hyperthermophilic archaebacteria will be an important aspect of biotechnology. Nevertheless, the prospects are intriguing. The nature of the environments that harbor these organisms and the consequent requirements for their controlled culture suggest that chemical and biochemical engineers can play an important role in elucidating their scientific and technological aspects.

Regulation of ribosomal RNA transcription by growth rate of the hyperthermophilic Archaeon, Pyrococcus furiosus.

We have studied the single rRNA gene cluster from the Archaeon, Pyrococcus furiosus. This isolate grows optimally at 100 degrees C and is thus a hyperthermophile. In P. furiosus, transcription of 16S rRNA is subject to regulation over a 7.5-fold range in response to a 20-fold increase in growth rate. The single cluster encoding the 16S and 23S rRNA genes of P. furiosus was cloned and the 1.9 kb region upstream of the 16S rRNA gene was sequenced.

Rabies as a transneuronal tracer of circuits in the central nervous system.

The ability of selected neurotropic viruses to move transneuronally in the central nervous system makes them particularly well suited for use as tracers in experimental neuroanatomy. Recently, techniques have been developed for using rabies virus as a transneuronal tracer. Several features of rabies infection make the virus particularly useful for this purpose. We examined transneuronal transport of rabies in the central nervous system of primates after intracortical and intramuscular injections. Rabies was transported in a time-dependent manner to infect synaptically-connected chains of neurons. Transport occurred exclusively in the retrograde direction. At the survival times we used, rabies infection was restricted to neurons and did not cause cell lysis. There are several methodological and safety issues that must be considered when designing studies that use rabies as a transneuronal tracer. When appropriate protocols and laboratory practices have been established, transneuronal transport of rabies can be a safe and efficient tool for revealing the organization of multi-synaptic circuits in the central nervous system.

Behavioral and neurobiological alterations induced by the immunotoxin 192-IgG-saporin: cholinergic and non-cholinergic effects following i.c.v. injection.

192-IgG-Saporin is an anti-neuronal immunotoxin that combines the 192 monoclonal antibody to the p75 neurotrophin receptor found on terminals and cell bodies of neurons in the cholinergic basal forebrain with the ribosome-inactivating protein saporin. Bilateral intraventricular injection of the 192-saporin produced a variety of dose-related behavioral, neurochemical, and histological alterations in adult male rats. While both the 2 micrograms and 4 micrograms dose produced comparable cholinergic hypofunction only the high dose produced behavioral changes. Behavioral deficits induced by the 4 micrograms dose of 192-saporin induced alterations in rotorod performance and reactivity on the hot-plate which recovered over 8 weeks. In addition, the 4 micrograms dose produced a persistent impairment in the acquisition and performance of standard Morris water maze task as well as a cued version of the task. The neurobiological alterations induced by 192-saporin involved both cholinergic and non-cholinergic systems. Both doses of 192-saporin produced a 60-80% decrease in high affinity choline transport in the hippocampus and cortex without altering this parameter in the striatum. In addition, there was a significant dose-related decrease of norepinephrine in the hippocampus in the high dose group. 192-saporin did not alter the content of dopamine, serotonin, or their metabolites in any region examined. 192-saporin also produced a loss of Purkinje cells in the cerebellum. This cell type also expresses the p75 receptor and appears to be a target for intraventricular 192-saporin. This complex interplay of factors makes the i.c.v. model of 192-saporin very problematic for studying the functional properties of the cholinergic basal forebrain. However, recent data suggest that injection of 192-saporin directly into components of the cholinergic basal forebrain can be used to further elaborate the function of this brain system and to model disorders of cholinergic hypofunction such as Alzheimer's disease.

Strategies to limit brain injury and promote recovery of function.

Neurotoxicology integrates the best aspects of pharmacology, neuroscience, and toxicology and has established unique approaches, questions, and model systems. It has contributed to (i) isolating the multitude of environmental factors that damage the nervous system, (ii) characterizing the nature of the insult in both developing and mature organisms, and (iii) identifying the populations at risk. These efforts have symbolized the scope of modern neurotoxicology but there is now a growing appreciation that neurotoxicology, neurology, and biological psychiatry share common interests that are based upon a set of unified conceptual questions. The unifying goals for all of these disciplines are to discern the biology of neural insult, its functional consequences, and the extent and underlying mechanisms of recovery of function. While research is beginning to unravel the biology of neurological disorders there is a widening gap between our understanding of the substrates of these diseases and our ability to prevent or treat them. A concerted effort must be mounted to develop new and appropriate treatments for neurodegenerative disorders. The vulnerability of the hippocampus to a broad spectrum of insults (cerebrovascular insufficiency, excitotoxins, age-related neurodegenerative disorders, drugs of abuse) and its involvement in cognitive function makes it a logical focal point for the study of the behavioral and neurobiological correlates of recovery of function. This article discusses models of neurodegeneration and different therapeutic strategies that might limit injury or promote recovery of function.

Preferential destruction of cerebellar Purkinje cells by OX7-saporin.

Saporin, a plant toxin derived from Saponaria officinalis, disrupts protein synthesis by inactivating the 60S portion of the ribosomal complex. OX7 is a mouse monoclonal antibody directed against the Thy-1.1 receptor that is differentially expressed on subpopulations of central nervous system neurons. Disulfide conjugation of OX7 to saporin permits delivery of saporin to target neurons. OX7-saporin was used to study the behavioral and morphological consequences of selective destruction of cerebellar Purkinje cells which abundantly express the Thy-1.1 antigen. Male Sprague-Dawley rats received bilateral intraventricular injections of 1- or 2 microg OX7-saporin or 8 microl artificial CSF. Rats were tested for behavioral changes 1 week before and 1, 2, and 8 weeks post-treatment. OX7-saporin treatment resulted in dose- and time-dependent changes in motor performance, activity, and negative geotaxis, but did not affect foot splay. Following behavioral testing, cerebellar sections were prepared for microscopic examination and the pattern of Purkinje cell loss was determined in anatomically matched sections. OX7-saporin induced dose-dependent death of Purkinje cells, particularly in the anterior and superior portions of cerebellar folia 1-6 and folium 9. Other brain regions appeared largely unaffected. Data suggest that intraventricular injection of rats with OX7-saporin is an effective model with which to examine the consequences of Purkinje cell destruction.

Isolation and characterization of Thermococcus barossii, sp. nov., a hyperthermophilic archaeon isolated from a hydrothermal vent flange formation.

A new hyperthermophilic microorganism, Thermococcus barossii, was isolated from rock fragments of a hydrothermal vent flange formation, located along the East Pacific Rise of the Juan de Fuca Ridge. This organism is obligately anaerobic and grows over a temperature range of at least 60-92 degrees C in artificial seawater-based media, containing elemental sulfur, tryptone and yeast extract. The addition of a maltooligosaccharide mixture and tungsten to this medium improved growth to some extent. At the Topt for growth (82.5 degrees C), cell densities as high as 4 x 10(8) cells/ml could be obtained in 18-liter batch fermentations, with a doubling time of approximately 40 minutes, if culture access to elemental sulfur was sufficient. In continuous culture at the same temperature, comparable cell densities could be obtained but only at slower growth rates. Morphologically, T. barossii is coccoid-shaped, forming irregularly-shaped spheres; under optimal conditions, these coccoids become more regular and smaller, a characteristic of other hyperthermophilic archaea. Negatively-stained preparations showed no pili or flagella associated with the cell surface. 16S rRNA sequencing reveals that T. barossii is most similar to Thermococcus celer (99.7%). Yet, further comparisons with T. celer showed that T. barossii is a new Thermococcus species: different growth temperature optimum (82.5 degrees C vs. 88 degrees C), obligate requirement for sulfur, higher G + C content (60% vs. 56.7%) and 47.7% DNA-DNA hybridization. The nucleotide and translated amino acid sequence for the gene encoding a DNA polymerase from T. barossii was compared to sequences of related genes from other Thermacoccales. The polymerase phylogenies were congruent with those obtained from the 16S rRNA phylogenetic analyses. Based on the high degree of similarity among members of the genus Termococcus for the criteria used thus far, aspects of enzymology may be an important mechanism of differenting one species from another.

Purification and functional characterization of a chaperone from Methanococcus jannaschii.

A chaperone from Methanococcus jannaschii has been purified to homogeneity with a single chromatographic step. The chaperone was identified and characterized using activity assays for characteristic chaperone abilities. The M. jannaschii chaperone binds unfolded proteins, protects proteins against heat-induced aggregation, and has a strongly temperature dependent ATPase activity. The chaperone has also been shown to inhibit the spontaneous refolding of a mesophilic protein at low temperatures. The purified chaperone complex has a M(r) of about 1,000,000 and consists of a single type of subunit with an approximate M(r) of 60,000. Analysis of partial sequence data reveals that this chaperone is the predicted protein product of the previously identified chaperonin gene in M. jannaschii (BULT et al., 1996). To our knowledge, this is the first functional characterization of a chaperone from a methanogen.

Continuous culture as a tool for investigating the growth physiology of heterotrophic hyperthermophiles and extreme thermoacidophiles.

Although there is great scientific and technological interest in examining the physiology and bioenergetics of microorganisms from extreme environments, difficulties encountered in their cultivation and lack of genetic systems hampers the investigation of these issues. As such, we have adapted methods for continuous cultivation of mesophilic organisms to extremes of temperature and pH to study extremophiles. Since the risk for contamination of extremophilic continuous cultures is relatively small, long-term, steady state experiments investigating physiological response to culture perturbations are possible. Experiments along these lines have provided insights into the significance of specific enzymes in the metabolism of particular substrates, in addition to providing a better understanding of stress response and unusual physiological characteristics of hyperthermophilic and extremely thermoacidophilic microorganisms. Several examples are provided here, including the thermal stress response of Metallosphaera sedula (T(opt) 74 °C) growing at pH 2.0, and the response of the heterotrophic hyperthermophiles Pyrococcus furiosus (T(opt) 98 °C), Thermococcus litoralis (T(opt) 88 °C) and T. maritima (T(opt) 80 °C) to changes in growth medium. Also discussed will be how the same experimental systems have been used to study exopolysaccharide production and biofilm formation by hyperthermophilic heterotrophs and facilitated the estimation of bioenergetic parameters for these organisms under a variety of growth conditions. Continuous culture, used in conjunction with genome sequence information, two-dimensional gel electrophoresis and differential gene expression, can provide important insights into the metabolism of high temperature extremophiles.

Controlling deamidation rates in a model peptide: effects of temperature, peptide concentration, and additives.

The rate of deamidation of the Asn residue in Val-Tyr-Pro-Asn-Gly-Ala (VYPNGA), a model peptide, was determined at pH 9 (400 mM Tris buffer) as a function of temperature and peptide concentration. Over the temperature range 5-65 degrees C, deamidation followed Arrhenius behavior, with an apparent activation energy of 13.3 kcal/mol. Furthermore, increasing the peptide concentration slows the rate of deamidation. Self-stabilization with respect to deamidation has not been reported previously. The rate of deamidation was also determined in the presence of sucrose and poloxamer 407 (Pluronic F127). In both cases, the rate of deamidation was retarded by up to 40% at 35 degrees C. In aqueous solutions containing poloxamer 407, the degree of stabilization is independent of formation of a reversible thermosetting gel. With sucrose, maximum reduction in the deamidation rate was attained with as little as 5% (w/v). Addition of sucrose results in a greater conformational preference for a type II beta-turn structure, which presumably is less prone to intramolecular cyclization and subsequent deamidation.

Preparation and in vitro characterization of gentamycin-impregnated biodegradable beads suitable for treatment of osteomyelitis.

A new method for preparing poly(L-lactide) (PLA) biodegradable beads impregnated with an ionic aminoglycoside, gentamycin, is described. The process employs hydrophobic ion pairing to solubilize gentamycin in a solvent compatible with PLA, followed by precipitation with a compressed antisolvent (supercritical carbon dioxide). The resulting precipitate is a homogeneous dispersion of the ion-paired drug in PLA microspheres. The microspheres are approximately 1 microm in diameter and can be compressed into beads (3-6 mm in diameter) strung on surgical sutures for implantation. The bead strings exhibit no significant change in release kinetics upon sterilization with a hydrogen peroxide plasma (Ster-Rad). The kinetics of gentamycin release from the PLA beads are consistent with a matrix-controlled diffusion mechanism. While nonbiodegradable poly(methyl methacrylate) (PMMA) beads initially release gentamycin in a similar manner, the drug release from PMMA ceases after 8 or 9 weeks, while the PLA beads continue to release drug for over 4 months. Moreover, only 10% of the gentamycin is released from the PMMA beads, while PLA beads release more than 60% of their load, if serum is present in the release medium. The PLA system displays improved release kinetics relative to PMMA, is biodegradable, is unaltered by gas sterilization, can be used for a range of antibiotics, and can be manipulated without disintegration. These are all desirable properties for an implantable drug delivery system for the prevention or treatment of osteomyelitis.

Carcinoma in a breast fibroadenoma.

A 48-year-old woman presented with a long history of a lump in the breast, which was clinically diagnosed as a fibroadenoma. Cytologic examination of fluid aspirated intraoperatively demonstrated groups of carcinoma cells. Excision of the mass showed that this discrepancy was due to carcinoma within a fibro adenoma. Reports of such an occurrence are few, and none, to the best of our knowledge, has previously documented the cytologic findings.

Extremozymes: expanding the limits of biocatalysis.

The study of enzymes isolated from organisms inhabiting unconventional ecosystems has led to the realization that biocatalysis need not be constrained to mild conditions and can be considered at pH's, temperatures, pressures, ionic and solvent environments long thought to be destructive to biomolecules. Parallel to this, it has been demonstrated that even conventional enzymes will catalyze reactions in solvents other than water. However, the intrinsic basis for biological function under extreme conditions is only starting to be addressed, as are associated applications. This was the focus of a recent NSF/NIST-sponsored workshop on extremozymes. Given the information acquired from the study of extremozymes, modification of enzymes to improve their ranges of stability and activity remains a possibility. Ultimately, by expanding the range of conditions suitable for enzyme function, new opportunities to use biocatalysis will be created.