Identification of bacteria associated with Dinoflagellates (Dinophyceae) Alexandrium spp. using tyramide signal amplification fluorescent in situ hybridization and confocal microscopy

In the marine environment, phytoplankton and bacterioplankton can be physically associated. Such association has recently been hypothesized to be involved in the toxicity of the dinoflagellate genus Alexandrium. However, the methods, which have been used so far to identify, localize, and quantify bacteria associated with phytoplankton, are either destructive, time consuming, or lack precision. In the present study we combined tyramide signal amplification–fluorescent in situ hybridization (TSA-FISH) with confocal microscopy to determine the physical association of dinoflagellate cells with bacteria. Dinoflagellate attached microflora was successfully identified with TSA-FISH, whereas FISH using monolabeled probes failed to detect bacteria, because of the dinoflagellate autofluorescence. Bacteria attached to entire dinoflagellates were further localized and distinguished from those attached to empty theca, by using calcofluor and DAPI, two fluorochromes that stain dinoflagellate theca and DNA, respectively. The contribution of specific bacterial taxa of attached microflora was assessed by double hybridization. Endocytoplasmic and endonuclear bacteria were successfully identified in the nonthecate dinoflagellate Gyrodinium instriatum. In contrast, intracellular bacteria were not observed in either toxic or nontoxic strains of Alexandrium spp. Finally, the method was successfully tested on natural phytoplankton assemblages, suggesting that this combination of techniques could prove a useful tool for the simultaneous identification, localization, and quantification of bacteria physically associated with dinoflagellates and more generally with phytoplankton.

Identification of image attributes that are most affected with changes in displayed image size

This paper describes an investigation of changes in image appearance when images are viewed at different image sizes on a high-end LCD device. Two digital image capturing devices of different overall image quality were used for recording identical natural scenes with a variety of pictorial contents. From each capturing device, a total of sixty four captured scenes, including architecture, nature, portraits, still and moving objects and artworks under various illumination conditions and recorded noise level were selected. The test set included some images where camera shake was purposefully introduced. An achromatic version of the image set that contained only lightness information was obtained by processing the captured images in CIELAB space. Rank order experiments were carried out to determine which image attribute(s) were most affected when the displayed image size was altered. These evaluations were carried out for both chromatic and achromatic versions of the stimuli. For the achromatic stimuli, attributes such as contrast, brightness, sharpness and noisiness were rank-ordered by the observers in terms of the degree of change. The same attributes, as well as hue and colourfulness, were investigated for the chromatic versions of the stimuli. Results showed that sharpness and contrast were the two most affected attributes with changes in displayed image size. The ranking of the remaining attributes varied with image content and illumination conditions. Further, experiments were carried out to link original scene content to the attributes that changed mostly with changes in image size.

Identification of a Novel Recycling Sequence in the C-tail of FPR2/ALX Receptor: Association with Cell Protection from Apoptosis

Formyl-peptide receptor type 2 (FPR2; also called ALX because it is the receptor for lipoxin A4) sustains a variety of biological responses relevant to the development and control of inflammation, yet the cellular regulation of this G-protein-coupled receptor remains unexplored. Here we report that, in response to peptide agonist activation, FPR2/ALX undergoes β-arrestin-mediated endocytosis followed by rapid recycling to the plasma membrane. We identify a transplantable recycling sequence that is both necessary and sufficient for efficient receptor recycling. Furthermore, removal of this C-terminal recycling sequence alters the endocytic fate of FPR2/ALX and evokes pro-apoptotic effects in response to agonist activation. This study demonstrates the importance of endocytic recycling in the anti-apoptotic properties of FPR2/ALX and identifies the molecular determinant required for modulation of this process fundamental for the control of inflammation.