An Emerging Approach for Parallel Quantification of Intracellular Protozoan Parasites and Host Cell Characterization Using TissueFAXS Cytometry

Characterization of host-pathogen interactions is a fundamental approach in microbiological and immunological oriented disciplines. It is commonly accepted that host cells start to change their phenotype after engulfing pathogens. Techniques such as real time PCR or ELISA were used to characterize the genes encoding proteins that are associated either with pathogen elimination or immune escape mechanisms. Most of such studies were performed in vitro using primary host cells or cell lines. Consequently, the data generated with such approaches reflect the global RNA expression or protein amount recovered from all cells in culture. This is justified when all host cells harbor an equal amount of pathogens under experimental conditions. However, the uptake of pathogens by phagocytic cells is not synchronized. Consequently, there are host cells incorporating different amounts of pathogens that might result in distinct pathogen-induced protein biosynthesis. Therefore, we established a technique able to detect and quantify the number of pathogens in the corresponding host cells using immunofluorescence-based high throughput analysis. Paired with multicolor staining of molecules of interest it is now possible to analyze the infection profile of host cell populations and the corresponding phenotype of the host cells as a result of parasite load.     

Four-year growth dynamics of beech-spruce model ecosystems under CO2 enrichment on two different forest soils

To elucidate how atmospheric CO₂ enrichment, enhanced nutrient supply and soil quality interact to affect regrowth of temperate forests, young Fagus sylvatica and Picea abies trees were grown together in large model ecosystems. Identical communities were established on a nutrient-poor acidic and on a more fertile calcareous soil and tree growth, leaf area index, fine root density and soil respiration monitored over four complete growing seasons. Biomass responses to CO₂ enrichment and enhanced N supply at the end of the experiment reflected compound interest effects of growth stimulation during the first two to three seasons rather than persistent stimulation over the whole duration of the experiment. Whereas biomass of Picea was enhanced in elevated CO₂ on both soils, Fagus responded negatively to CO₂ on acidic but positively on calcareous soil. Biomass of both species profited from enhanced N supply on the poor acidic soil only. Leaf area index on both soils was greater in high N supply as a consequence of a stimulation early in the experiment, but was unaffected by CO₂ enrichment. Fine root density on acidic soil was increased in high N supply, but this did not stimulate soil respiration rate. In contrast, elevated CO₂ stimulated both fine root density and soil CO₂ efflux on calcareous soil, especially towards the end of the experiment. Our experiment suggests that future species dominance in beech-spruce forests is likely to change in response to CO₂ enrichment, but this response is subject to complex interactions with environmental factors other than CO₂, particularly soil type.     

Influence of oxygenated sterol compounds on phase transitions in model membranes. A study by differential scanning calorimetry

A marked influence of oxygenated sterol compounds (OSC: 7 alpha-, 7 beta-, and 25-hydroxycholesterol and 7-ketocholestanol) on the reversible gel to liquid-crystalline phase transition behavior of cholesterol-free and cholesterol-containing model membranes is evidenced by high-sensitivity differential scanning calorimetry. Liposomes of dipalmitoylphosphatidylcholine (DPPC) were chosen as model membranes. Each of the investigated OSC exerts an individual influence on the phase transition of DPPC liposomes, which expresses itself in the temperature range, the corresponding enthalpy, and the peak shape of calorimetric curves. The onset temperature of the phase transition is lowered in the following range of effectiveness: 7 beta-hydroxycholesterol greater than 7 alpha-hydroxycholesterol greater than 7-ketocholestanol greater than cholesterol. The mutual presence of cholesterol and of OSC leads to the following order: 7 alpha-hydroxycholesterol approximately equal to 7 beta-hydroxycholesterol greater than 7-ketocholestanol greater than cholesterol (without OSC) greater than 25-hydroxycholesterol. The enthalpy of the phase transition is decreased with increasing content of cholesterol, 7 alpha- or 7 beta-hydroxycholesterol, or 7-ketocholestanol. At a concentration of about 10 mol % of the latter three OSC, the corresponding enthalpy value of the transition is lowered from 9.1 kcal/mol for pure DPPC to about 7.5 kcal/mol, whereas 10 mol % cholesterol lowers the enthalpy value to 7.0 kcal/mol.(ABSTRACT TRUNCATED AT 250 WORDS)     

Growth characteristics of a thermotolerant methylotrophic Bacillus sp. (NCIB 12522) in batch culture

Summary This contribution deals with problems associated with the culture of a thermotolerant methylotrophic Bacillus sp. The results reported clearly demonstrate why conventional enrichment/isolation procedures have, in the past, failed to allow such microbes to assert themselves. The catastrophic effect of carbon substrate (methanol) exhaustion on such cultures is clearly evidenced, but the effects of other nutrient exhaustion or limitations are demonstrated to be markedly less stringent. The failure of such cultures to complete the sporulation process when growing on methanol has important consequences with respect to their survival characteristics.     

PspA adopts an ESCRT-III-like fold and remodels bacterial membranes

1. Download : Download high-res image (323KB)
2. Download : Download full-size image