The dielectric permittivity at radio frequencies and the bruggeman probe: novel techniques for the on-line determination of biomass concentrations in plant cell cultures

A novel technique is described for the measurement of the volume fraction of biomass in a suspension by the simultaneous measurement of the conductivity of a suspension containing cells and of the medium in which the cells are suspended. The presence of non-conducting particulate matter in a suspension will cause the conductivity of a suspension to be decreased relative to that of the medium in which the particles are suspended. A simple equation (the Bruggeman equation) describes the relationship between the volume fraction of non-conducting particulate matter and the decrease in conductivity. The accuracy of this method for the determination of the biomass concentration of plant cells (Festuca arundinacea) in culture was shown. The method was successfully applied to the on-line determination of biomass concentrations during the growth of F. arundinacea cultures, and gave good agreement with biomass levels as determined from measurements of the radio-frequency dielectric permittivity of such cultures.     

In vivo pH measurement in the xylem of broad-leaved trees using ion-sensitive field-effect transistors

Summary A new method of in vivo pH determination in the xylem of broad-leaved trees using ion-sensitive field effect transistors is developed and its suitability for use is studied. In the first few hours after the sensor had been implanted in the xylem signals could be detected which were generated in response to mechanical damage; particularly strong signal changes are detectable in Populus balsamifera L., Tilia cordata Mill, and Aesculus hippocastanum L. The pH values of the xylem sap extracted from branches corresponded to the values measured by the in vivo method only at certain times. Due to sensor drift the measuring accuracy of long-term experiments lasting up to 3 weeks is restricted. The in vivo measurement of pH in the xylem of poplar branches revealed the ability of the living xylem to buffer the pH of the sap to its own characteristic value.Dedicated to Prof. Dr. O. L. Lange to his 65th birthday     

Analysis of pollen-tube growth in cultured maize silks

Summary In vitro pollen-tube growth in maize was studied using an in vitro pollination system. In the cut-silk method, ovaries with silks were placed on medium in vitro, whereafter the silk was cut and the upper part of the silk was pollinated. Pollen tubes were not able to bridge the space between the two silk parts. Even when silk parts were tightly connected, pollen tubes still were not able to pass the cut ends and reach the lower silk part. Pollen-tube growth rates and the direction of tube growth were not influenced by the presence or absence of an ovary. Prepollination did not have any influence on pollen-tube growth rate. Measurements of pollen-tube growth rate also showed that there was no population effect, i.e. growth rate was not stimulated by pollination with an excess of pollen grains. We found that the direction in which maize pollen grew was determined only by the positioning of the silk hairs.     

The expression of PS integrins in Drosophila melanogaster imaginal disc cell lines

Summary Drosophila imaginal disc cell lines show a characteristic pattern of aggregation in culture, which appears to be due to cell-cell rather than cell-substrate interactions. We have examined the distribution of PS integrins in wing and leg cell lines, and find that these integrin homologues are expressed preferentially in aggregates. Cell sheets, small cell clumps and chains of cells express antigen at points of cell-cell contact only.     

Whole-embryo culture of Drosophila : development of embryonic tissues in vitro

Summary We have devised techniques to culture whole, dissected embryos of Drosophila melanogaster. We examine multiple aspects of the morphological and physiological development of the epidermis, musculature, nervous system, and internal organs in this cultured preparation, and show that in vitro development closely parallels normal embryogenesis. These techniques permit a wide range of experimental manipulations during embryogenesis and allow us to extend observations through late embryonic stages, after cuticle deposition. Applications of this technique are presented.     

Production of blastospores by three strains of metarhizium anisopliae (metch.) sorokin in submerged culture

Studies on blastospore production in different liquid media were conducted with three strains of Metarhizium anisopliae var. anisopliae (M. a.) derived from various countries (M. a. 43: Austria, M. a. 57: Brazil, M. a. 97: Philippines). Variation of six fermentation parameters (cornsteep products, carbohydrates, pH values, temperature, Tween 80, and polyethyleneglycol (PEG) 200) disclosed that the three strains of M. anisopliae differed in their growth pattern and physiology. In standard medium and in all tests, M. a. 57 produced the highest number of blastospores invariably amounting to > 10⁸ per ml, while mycelial pellets were never formed. The preferred carbohydrates were glucose and fructose. Blastospore production of M. a. 43 was increased by growth at 30°C, at pH 6.5 or by addition of 5% PEG 200. However, it was impaired by different concentrations of Tween 80 or higher concentrations of PEG 200 (10–15%). M. a. 97 produced most blastospores at 30°C, and the strain preferred basic (pH 8.0) as well as acid (pH 4.5) media. Blastospore production was increased by the addition of 5% PEG 200 or 0.4–1.2% Tween 80. Moreover, PEG 200 suppressed pellet formation effectively. Altogether, our results showed that for optimal blastospore production of Metarhizium anisopliae, suitable strain‐specific parameters have to be evaluated.     

Biodiversity amongst microorganisms and its relevance

Concluding remarks from the joint IUBS/IUMS workshop on Biodiversity amongst microorganisms and its relevance held in Amsterdam on 7–8 September 1991. An international microbial ecology programme can be justified in its own right now that appropriate investigative tools have been developed. Microorganisms influence global change, and indicate global health and environmental quality. At the same time, an inventory of the world's microbial species and their properties is required, together with associated culture collections and genomes. Sampling methods need to be standardized, both for species and functions. Extreme environments are a particularly rich source of microbial genomes, and endangered habitats should be sampled as a matter of priority. Cataloguing and conserving the world's microbial biodiversity is justifiable and scientifically important.     

Modulations of the epithelial phenotype and functional activity of cultured bovine tracheal gland cells: dependence on the culture medium and passage number.

Bovine tracheal gland (BTG) cells in culture show an epithelial-fibroblastoid transition after several passages. To investigate these BTG cell phenotype changes, we studied the effects of both the culture medium and passage number on the expression of epithelial cytoskeletal proteins and glandular serous cell markers. We also analyzed the intracellular cAMP level in the basal state and after adrenergic stimulation. Three culture media were used: 1) serum-free defined medium (SFDM); 2) medium supplemented with 2% Ultroser G; and 3) medium supplemented with 10% fetal calf serum (FCS). Using immunofluorescence microscopy, we showed that, in the first 4 passages whatever the culture conditions, BTG cells expressed immunoreactivities to cytokeratin filaments and desmoplakins I and II, whereas vimentin filaments were not detected. After four passages, BTG cells cultured in 10% FCS or 2% Ultroser G became progressively fibroblastoid and showed immunoreactivities to both vimentin and cytokeratin intermediate filaments. No immunoreactivity to vimentin filaments was observed on BTG cells cultured in a SFDM. Using biochemical analysis, we showed that basal levels of cAMP in cultured BTG cells and lysozyme secretion by these cells vary according to the culture medium and passage number. It was higher in BTG cells cultured in a SFDM compared to that recovered from cells cultured in medium supplemented with Ultroser G or FCS. Whatever the culture medium, BTG cells responded to stimulation by isoproterenol. However, the results of stimulation in a SFDM were higher than in Ultroser G or FCS supplemented medium. We conclude that the BTG epithelial cell organization and the regulation of biosynthesis of secretory proteins by these cells in culture depend on both the culture medium and passage number.(ABSTRACT TRUNCATED AT 250 WORDS)     

Onychomycosis caused by Scopulariopsis brumptii

Scopulariopsis brumptii was isolated from nail lesions in left hand of a 42 year-old-farmer. The direct microscopic examination of the nail samples revealed light brown, septate, branched fungal hyphae along with thick-walled spherical cells. The histopathological examination showed involvement of internal phase of the nail plate. Amongst the antimycotics tested against S. brumptii In vitro oxiconazole was found to be the most active with MIC value of 10 g/ml^–1. This report documents the first instance of onychomycosis caused by S. brumptii.     

Ammonia rhythm in Microcystis firma studied by in vivo 15N and 31P NMR spectroscopy

Cultures of the cyanobacterium Microcystis firma show rhythmic uptake and release of ammonia under conditions of carbon limitation. The massive removal of ammonia from the medium during the first light phase has little impact on the intracellular pH: a pH shift of less than 0.2 U towards the alkaline can be measured by in vivo ³¹P NMR. Furthermore, the energy status of the cells remains regulated. In vivo ¹⁵N NMR of M. firma, cultivated either with labelled nitrate or ammonia as the sole nitrogen source, reveals only gradual differences in the pool of free amino acids. Additionally both cultivation types show -aminobutyric acid, acid amides and yet unassigned secondary metabolites as nitrogen storing compounds. Investigating the incorporation of nitrogen under carbon limitation, however, only the amide nitrogen of glutamine is found permanently labelled in situ. While transamination reactions are blocked, nitrate reduction to ammonia can still proceed. Cation exchange processes in the cell wall are considered regarding the ammonia disappearance in the first phase, and the control of ammonia uptake is discussed with respect to the avoidance of intracellular toxification.Abbreviations GABA -aminobutyric acid - GOGAT glutamate synthase - GS glutamine synthetase - MDP methylene diphosphonate - MOPSO 3-(N-morpholino)-2-hydroxy-propanesulfonic acid - NDPS nucieoside diphosphosugars - NOE nuclear Overhauser effect - NMR nuclear magnetic resonance For convenience, the term ammonia is used throughout to denote ammonia or ammonium ion when there is no good evidence as to which chemical species is involved