The persistence of Legionella pneumophila in non-sterile, sterile and artificial hard waters and their growth pattern on tap washer fittings

Simultaneous experiments were performed with sterilized and non-sterile water and an artificial hard water. After seeding with an environmental isolate of Legionella pneumophila numbers in the sterile land hard water decreased rapidly and colonization of various tap washer fittings failed to take place. Adhesion and growth of an environmental isolate of L. pneumophila to washers in non-sterile tap water was followed over a 4-month period with fluorescein-labelled antibody and by scanning electron microscopy. After adherence the individual cells appeared to divide to form chains which spread over the surfaces. Organisms other than legionellas were also present and a complex colonization matt was formed which was embedded in a protective coat of slime and debris. The numbers of L. pneumophila recovered from the water were highest between 4 and 7 weeks but they could still be cultivated after 4 months.     

Verapamil counteracts the masking of long-lasting potentiation of hippocampal population spike produced by 2-amino-5-phosphonovalerate

In transversely sectioned rat hippocampal slices the effects of 2-amino-5-phosphonovalerate (APV), presumably a selective N-methyl-DL-aspartate antagonist, were examined on the development of long-lasting potentiation (LLP) of the CA1 population spike produced by Schaffer collateral tetanization (100 Hz, 1 s). APF (100 nA, 5 min), when applied 150-200 micron away from the CA1 cell bodies (apical dendritic area), had no significant effect if the population spike was evoked at 0.1 Hz, but produced a depression of the population spike if a 100 Hz tetanus was given during its iontophoresis. This depressant effect of APV was counteracted by verapamil (0.33 microM, 3 min) and LLP induced by the 100 Hz tetanus was unmasked. It is suggested that APV does not antagonize LLP, but only masks it by allowing CA++ influx into CA1 neurones that induces a depression. The results also raise doubts as to the selectivity of APV as an amino acid antagonist.     

Somatic hybrids between Solanum brevidens and Solanum tuberosum: Expression of a late blight resistance gene and potato leaf roll resistance

Hexaploid somatic hybrids resulting from mesophyll protoplast fusions between Solanum brevidens Phil., PI 218228, and Solanum tuberosum L., PI 203900 were tested for late blight resistance using two races of Phytophthora infestans Monte., de Bary. The S. tuberosum parent was a late blight differential possessing the R4 gene which confers resistance to race 0. The S. brevidens parent is resistant to potato leaf roll virus. Inoculations with both compatible (race 1.3.4.5) and incompatible (race 0) races of P. infestans clearly demonstrated the expression of the late blight resistance gene in all of the hybrid progeny tested. Most of the hybrids tested were also resistant to potato leaf roll virus (PLRV), indicating that the S. brevidens genes for PLRV resistance were present and expressed.     

Bacillus thuringiensis and control of plant pests

Summary The feasibility ofB. thuringiensis as an economic insect control agent is dependent upon various factors. Consideration of three situations. whereB. thuringiensis could be used illustrates the interaction of these factors, and their contribution to determining the success or failure of the organism as an insecticide. Apple crops suffer from attack by a complex of insect pests, many of which are very susceptible to theB. thuringiensis toxin. However chemical control methods are preferred by growers in order to satisfy the consumers' demand for top quality blemish-free fruit. By contrast cotton growers are beset with problems of pest resistance to chemical insecticides. This problem is tackled by using pest management strategies which orchestrate all possible methods of control. Such a situation is ideally suited toB. thuringiensis products, but as yet they have proven to be of inadequate efficacy. The softwood forest industry is an example whereB. thuringiensis is both needed and effective. The pest complex is relatively simple, and there is public concern about the health risks of chemical sprays. Recent developments of high performance formulations means thatB. thuringiensis is an economic, cost-effective, biological alternative to chemical control for forestry pests.

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Resumen La posibilidad de utilizarB. thuringiensis para el control de insectos de una forma económica depende de varios factores. La consideración de tres situaciones distintas en las queB. thuringiensis puede ser utilizado ilustra la interacción de estos factores y su contribución al éxito o fracaso del organismo como insecticida. El cultivo de manzanas sufre el ataque de una serie de plagas de insectos, muchas de las cuales son susceptibles a la toxina deB. thuringiensis. Sin embargo, los agricultores prefieren utilizar métodos de control químico a fin de satisfacer la demanda del consumidor por un fruto de primera calidad sin marcas ni señales. Por el contrario, los cultivadores de algodon estan sensibilizados a los problemas de la aparición de plagas resistentes a los insecticidas químicos. Este problema se ha abordado utilizando sistemas de control integrado que aglutinan todos los posibles métodos de control. Esta situación parece la ideal para el uso de productos a base deB. thuringiensis, pero hasta ahora estos han resultado ser poco eficaces. La industria de la madera blanda constituye en ejemplo en el cualB. thuringiensis es a la vez necesario y efectivo. Las plagas en cuestión forman un sistema relativamente simple y los efectos de los sprays químicos sobre la salud pública son causa de preocupación. Los desarrollos recientes en formulaciones de elevada eficacia convierten aB. thuringiensis en una alternativa biológica económicamente factible frente al control químico de plagas forestales.

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Résumé La possibilité d'utiliser économiquementB. thuringiensis dans la lutte contre les insectes nuisibles dépend de facteurs divers. L'examen de trois situations dans lesquellesB. thuringiensis peut être utilisé illustre l'interaction de ces facteurs et leur contribution dans le succès ou l'échec de cet organisme comme insecticide. Les récoltes de pommes sont menacées par les attaques d'un mélange complexe d'insectes, dont beaucoup sont sensibles à la toxine deB. thuringiensis. Cependant, les horticulteurs préfèrent les méthodes chimiques à cause de l'exigence des consommateurs pour des fruits de haute qualité et sans aucune tache. Par contre, les cultivateurs de coton sont obsédés par les problèmes de résistance aux insecticides chimiques. Ce problème est affronté par des stratégies faisant appel à tous les moyens de lutte possibles. C'est une situation convenant bien aux produits qui contiennentB. thuringiensis, mais jusqu'ici ces produits se sont montrés inefficaces. L'industrie du bois tendre forestier est toutefois un exemple de cas oùB. thuringiensis est à la fois nécessaire et efficace. En effet, le complexe des insectes agressifs est relativement simple et il existe en outre une prévention du public contre les risques sanitaires des pulvérisations de produits chimiques. La mise au point récente de formulations à haute performance démontre queB. thuringiensis est, pour la lutte contre les insectes nuisibles des forêts, une alternative économique et bon-marché.

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Invited paper presented at the VII International Conference on the Global Impacts of Applied Microbiology, Helsiki, 12–16 August 1985. Session 8     

Transfer of exogenous cholesterol to microsomes of hepatocytes investigated with [3H]desmosterol tracer.

The molybdenum centre of spinach (Spinacia oleracea) nitrate reductase has been investigated by e.p.r. spectroscopy of molybdenum(V) in reduced forms of the enzyme. The resting enzyme gives no signals attributable to Mo(V). However, on reduction with NADH, Mo(V) signals appeared at relatively short reaction times but decreased again on prolonged exposure to excess of the substrate as the enzyme was further reduced. On brief treatment of such samples with nitrate, Mo(V) signals reappeared but disappeared again on longer exposure to excess nitrate as the enzyme became fully reoxidized. Detailed investigation of the signals carried out in both 1H2O and 2H2O revealed the presence of two signal-giving species, referred to as 'signal A' and 'signal B', analogous to corresponding signals from nitrate reductase from Escherichia coli and from liver sulphite oxidase. Signal A has gav. 1.9767 and shows coupling to a single proton, exchangeable with the solvent, with A(1H)av. 1.3mT, whereas signal B shows no more than weak coupling to protons. Investigation of interconversion between the two species indicated that decreasing the pH from 8.0 to 6.7 had little effect, but that signal A was favoured by the presence of Cl-. This suggests, by analogy with recent work on sulphite oxidase by Bray, Gutteridge, Lamy & Wilkinson [Biochem. J. (1983) 211, 227-236] that Cl- is a ligand of molybdenum in the species giving signal A.     

On the relationship of ultrastructural and cytochemical features to color in mammalian skeletal muscle

Summary The semitendinosus muscle of the albino rat is divided grossly into two clearly distinguishable parallel longitudinal bands, one red (anterior) and the other white (posterior). By using mitochondrial content as a criterion for distinguishing fiber types, it is demonstrated that the red portion of the muscle is composed predominantly of red (52%) and intermediate (40%) fibers, while the white portion consists primarily of white fibers (82%). Red fibers have the smallest and white fibers have the largest average diameter. Ultrastructural characteristics of the three fiber types resemble closely those previously described for the rat diaphragm. Red fibers are rich in large mitochondria with abundant cristae, and possess the widest Z lines. In red fibers, the H-band region of the sarcoplasmic reticulum consists of an elaborate network of narrow tubules. In white fibers, mitochondria are smaller, less numerous, and have fewer cristae; Z lines are about half as wide as in red fibers. In the H-band region of the sarcoplasmic reticulum there is a more compact arrangement of broad more or less parallel tubules. Intermediate fibers are similar to red fibers except that their diameters are larger; mitochondria are somewhat smaller and cristae are less abundant; the width of the Z lines is close to that of white fibers. The consistent difference in Z line width establishes this dimension as an important criterion for distinguishing fiber types and facilitates ultrastructural identification, especially of the intermediate fiber.The clear relationship between color of the semitendinosus and cytological features of its component fibers supports the use of the terms red, white, and intermediate as simple and valid designations for fiber types in mammalian skeletal muscle. Measurement of the cross-sectional area contributed by each fiber type to the total area indicates that both red and intermediate fibers may contribute to redness in mammalian skeletal muscle.An early portion of this work was carried out with MissSharon Whelan (Mrs.Bernard Weiss). The author acknowledges the important contribution of Mr.Richard Stearns through his skillful work on the photographic illustrations and the technical assistance of MissAnn Campbell and Mrs.Joan Normington. — This study was supported by Grant No. HD 01026-04 from the United States Public Health Service.     

Species preservations in an optimal harvest model with random prices

In this paper, we consider an optimal harvest model in which the objective is to maximize the expected return. The unit price of biomass is assumed constant until a random time when the price increases by a given amount. Furthermore, due to obvious environmental protection requirements, it is assumed that the fishery population is bounded from below for all time so as to reduce the danger of species extinction. Clearly, this problem is an optimal control problem in which a random parameter is involved. However, due to its special structure, it is shown that the problem is convertible into a deterministic optimal control problem and hence is solvable by an existing optimal control software package, MISER. The practical implication of several computed results obtained by this approach is discussed. They are also compared with other related results in the literature.     

High frequency adventitious shoot regeneration from excised leaves ofPaulownia spp. cultured in vitro

High frequency, direct regeneration of shoots was induced in leaf cultures ofPaulownia tomentosa, P. fortunei x P. tomentosa andP. kawakamii. The optimum culture medium for the leaf explants derived from shoot cultures was Murashige-Skoog (MS) medium supplemented with 10 M indole-3-acetic acid and 50 M benzyladenine. Up to 40 shoots were obtained over a 4 month culture period from each leaf explant. Rooting occurred spontaneously in the shoots that were about 1 cm tall when subcultured on phytohormone-free MS medium. The plantlets could be transplanted successfully. Some of the transplantedP. tomentosa plantlets flowered in the greenhouse one year after transplanting. The protocol is suitable not only for rapid multiplication of the various species ofPaulownia, but also for analytical studies associated with adventitious shoot regeneration.     

Thermostability enhancement of polyethylene terephthalate degrading PETase using self- and nonself-ligating protein scaffolding approaches

Polyethylene terephthalate (PET) hydrolase enzymes show promise for enzymatic PET degradation and green recycling of single-use PET vessels representing a major source of global pollution. Their full potential can be unlocked with enzyme engineering to render activities on recalcitrant PET substrates commensurate with cost-effective recycling at scale. Thermostability is a highly desirable property in industrial enzymes, often imparting increased robustness and significantly reducing quantities required. To date, most engineered PET hydrolases show improved thermostability over their parental enzymes. Here, we report engineered thermostable variants of Ideonella sakaiensis PET hydrolase enzyme (IsPETase) developed using two scaffolding strategies. The first employed SpyCatcher-SpyTag technology to covalently cyclize IsPETase, resulting in increased thermostability that was concomitant with reduced turnover of PET substrates compared to native IsPETase. The second approach using a GFP-nanobody fusion protein (vGFP) as a scaffold yielded a construct with a melting temperature of 80°C. This was further increased to 85°C when a thermostable PETase variant (FAST PETase) was scaffolded into vGFP, the highest reported so far for an engineered PET hydrolase derived from IsPETase. Thermostability enhancement using the vGFP scaffold did not compromise activity on PET compared to IsPETase. These contrasting results highlight potential topological and dynamic constraints imposed by scaffold choice as determinants of enzyme activity.