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.     

Morphometry, densitometry and pattern analysis of plastic-embedded histologic material from urothelial cell carcinoma of the bladder.

An image analysis method of grading histologic sections of bladder carcinoma was tested. The method was new in four respects. First, for fixation of the biopsies a coagulant fixative was used. Second, 2-microns plastic sections were used to ensure the reproducibility of nuclear imaging. Third, a new stereologic approach was used for calculation of the nuclear volume and DNA content. Fourth, for the classification rule the morphometric, densitometric and texture features were used in concert. The IBAS 2000 instrument was used for the measurements. Texture analysis of the chromatin patterns was performed using Markovian texture features. Using discriminant analysis, of 22 parameters, 2 morphometric, 2 densitometric and 3 texture features were selected for the classification rule. With them, 89% of the bladder carcinomas were correctly classified into the three grades. All grade III tumors were classified correctly. Among the features tested, the densitometry of the DNA had the highest F values. All of the grade III tumors and 45% of the grade II tumor group had DNA histograms indicating aneuploidy. This study showed that plastic-embedded material is well suited to morphometry and densitometry and can be used for quantitative grading of bladder carcinoma.     

When the Background Matters: Using Scenarios from Integrated Assessment Models in Prospective Life Cycle Assessment

Prospective life cycle assessment (LCA) needs to deal with the large epistemological uncertainty about the future to support more robust future environmental impact assessments of technologies. This study proposes a novel approach that systematically changes the background processes in a prospective LCA based on scenarios of an integrated assessment model (IAM), the IMAGE model. Consistent worldwide scenarios from IMAGE are evaluated in the life cycle inventory using ecoinvent v3.3. To test the approach, only the electricity sector was changed in a prospective LCA of an internal combustion engine vehicle (ICEV) and an electric vehicle (EV) using six baseline and mitigation climate scenarios until 2050. This case study shows that changes in the electricity background can be very important for the environmental impacts of EV. Also, the approach demonstrates that the relative environmental performance of EV and ICEV over time is more complex and multifaceted than previously assumed. Uncertainty due to future developments manifests in different impacts depending on the product (EV or ICEV), the impact category, and the scenario and year considered. More robust prospective LCAs can be achieved, particularly for emerging technologies, by expanding this approach to other economic sectors beyond electricity background changes and mobility applications as well as by including uncertainty and changes in foreground parameters. A more systematic and structured composition of future inventory databases driven by IAM scenarios helps to acknowledge epistemological uncertainty and to increase the temporal consistency of foreground and background systems in LCAs of emerging technologies.     

Shading and Watering as a Tool to Mitigate the Impacts of Climate Change in Sea Turtle Nests

Increasing sand temperatures resulting from climate change may negatively impact sea turtle nests by altering sex ratios and decreasing reproductive output. We analyzed the effect of nest shading and watering on sand temperatures as climate mitigation strategies in a beach hatchery at Playa Grande, Costa Rica. We set up plots and placed thermocouples at depths of 45cm and 75cm. Half of the plots were shaded and half were exposed to the sun. Within these exposure treatments, we applied three watering treatments over one month, replicating local climatic conditions experienced in this area. We also examined gravimetric water content of sand by collecting sand samples the day before watering began, the day after watering was complete, and one month after completion. Shading had the largest impact on sand temperature, followed by watering and depth. All watering treatments lowered sand temperature, but the effect varied with depth. Temperatures in plots that received water returned to control levels within 10 days after watering stopped. Water content increased at both depths in the two highest water treatments, and 30 days after the end of water application remained higher than plots with low water. While the impacts of watering on sand temperature dissipate rapidly after the end of application, the impacts on water content are much more lasting. Although less effective at lowering sand temperatures than shading, watering may benefit sea turtle clutches by offsetting negative impacts of low levels of rain in particularly dry areas. Prior to implementing such strategies, the natural conditions at the location of interest (e.g. clutch depth, environmental conditions, and beach characteristics) and natural hatchling sex ratios should be taken into consideration. These results provide insight into the effectiveness of nest shading and watering as climate mitigation techniques and illustrate important points of consideration in the crafting of such strategies.