Using dBASE to write PENNAME, a computer key to common Penicillium species

dBASE III Plus™ computer software includes a high level computer language as well as a database management program. dBASE possesses advantages for key construction, including simple entry of raw data, effective user interaction in the selection of outputs, and ready comparison of included species with unknowns. This paper describes the use of dBASE in the construction of PENNAME, a new computer key to common Penicillium species.     

Calcium binding and induction of conidiation in protoplasts of Penicillium cyclopium

Cell wall-free protoplasts of P. cyclopium could regenerate a cell wall and form mycelia in liquid culture with high rates of viability. When calcium was added to the medium, protoplasts displayed biphasic accumulation with an immediate metabolism-independent adsorption phase, followed by slow metabolism-dependent uptake.Exposure of the protoplasts to Ca²⁺ for periods of 2 min, followed by incubation in calcium-free medium for 24 hours, was sufficient to induce conidiation with morphogenetic events parallel to those found in cultures containing calcium throughout the incubation period, and similar to those reported in cultures inoculated from conidia.The conidiation event caused by short exposure to calcium could be reversed, within 2 hours of Ca²⁺ addition, by a brief treatment with the specific calcium chelating agent BAPTA (100 M), which removed 65 to 75% of the total cell calcium.The results implicate the membrane-bound calcium fraction in the process of conidiation induction.     

Preparation of right-side-out plasma membrane vesicles from Penicillium cyclopium: a critical assessment of markers.

A plasma membrane fraction was obtained by the combined use of differential centrifugation and aqueous polymer two-phase partitioning techniques. Vanadate-inhibited ATPase and glucan synthase activities were highly enriched in this fraction, although the presence of ATPase activity which was not inhibited by vanadate, nitrate, molybdate, anyimycin A or azide was also detected. Other intracellular membrane marker activities were present at very low or undetectable levels. A further separation step using Percoll density gradient centrifugation resulted in the separation of a fraction which exclusively contained vanadate-inhibited ATPase activity, and was enriched with silicotungstic-acid-staining membrane material. Latency tests performed on the plasma membrane markers showed that the membrane vesicles were in the right-side-out orientation.     

Potential Sequential Exposures to 2-Butoxyethanol After Use of a Hard-Surface Cleaner

Professional workers and consumers frequently use hard-surface cleaning products and these products may contain glycol ethers (GEs), such as 2-Butoxyethanol (2-BE). Governmental agencies have set exposure limits for some chemicals used in cleaning products and these exposure limits have been used as guides to protect human health. The study objectives were to determine realistic inhalation exposures for professional workers performing multiple, sequential cleaning tasks and compare the exposures to the acute reference exposure level (REL), which California established for 2-BE. The ConsExpo model was acceptable for evaluating exposure based on a comparison of its predictions to experimentally measured 2-BE vapor concentrations from hard-surface cleaning. The typical worker exposure was predicted for a cleaning scenario consisting of three bathrooms and three kitchens (or three bedrooms) in a 1-h period. This exposure scenario would not be expected to result in significant health consequences because the predicted exposure was much lower than the REL. The predicted chronic and aggregate exposures were also acceptable. This analysis identified two important variables that affect inhalation exposure: cleaners should be used with adequate ventilation and wet wiping towels should be properly disposed so that they are not a source of continuing exposure.     

Iron Is a Sensitive Biomarker for Inflammation in Multiple Sclerosis Lesions

MRI phase imaging in multiple sclerosis (MS) patients and in autopsy tissue have demonstrated the presence of iron depositions in white matter lesions.The accumulation of iron in some but not all lesions suggests a specific, potentially disease-relevant process, however; its pathophysiological significance remains unknown.Here, we explore the role of lesional iron in multiple sclerosis using multiple approaches: immunohistochemical examination of autoptic MS tissue, an in vitro model of iron-uptake in human cultured macrophages and ultra-highfield phase imaging of highly active and of secondary progressive MS patients.Using Perls'' stain and immunohistochemistry, iron was detected in MS tissue sections predominantly in non-phagocytosing macrophages/microglia at the edge of established, demyelinated lesions. Moreover, iron-containing macrophages but not myelin-laden macrophages expressed markers of proinflammatory (M1) polarization.Similarly, in human macrophage cultures, iron was preferentially taken up by non-phagocytosing, M1-polarized macrophages and induced M1 (super) polarization. Iron uptake was minimal in myelin-laden macrophages and active myelin phagocytosis led to depletion of intracellular iron.Finally, we demonstrated in MS patients using GRE phase imaging with ultra-highfield MRI that phase hypointense lesions were significantly more prevalent in patients with active relapsing than with secondary progressive MS.Taken together, our data provide a basis to interpret iron-sensitive GRE phase imaging in MS patients: iron is present in non-phagocytosing, M1-polarized microglia/macrophages at the rim of chronic active white matter demyelinating lesions. Phase imaging may therefore visualize specific, chronic proinflammatory activity in established MS lesions and thus provide important clinical information on disease status and treatment efficacy in MS patients.     

Nitrosative stress inhibits the aminophospholipid translocase resulting in phosphatidylserine externalization and macrophage engulfment: implications for the resolution of inflammation

Macrophage recognition of apoptotic cells depends on externalization of phosphatidylserine (PS), which is normally maintained within the cytosolic leaflet of the plasma membrane by aminophospholipid translocase (APLT). APLT is sensitive to redox modifications of its -SH groups. Because activated macrophages produce reactive oxygen and nitrogen species, we hypothesized that macrophages can directly participate in apoptotic cell clearance by S-nitrosylation/oxidation and inhibition of APLT causing PS externalization. Here we report that exposure of target HL-60 cells to nitrosative stress inhibited APLT, induced PS externalization, and enhanced recognition and elimination of "nitrosatively" modified cells by RAW 264.7 macrophages. Using S-nitroso-L-cysteine-ethyl ester (SNCEE) and S-nitrosoglutathione (GSNO) that cause intracellular and extracellular trans-nitrosylation of proteins, respectively, we found that SNCEE (but not GSNO) caused significant S-nitrosylation/oxidation of thiols in HL-60 cells. SNCEE also strongly inhibited APLT, activated scramblase, and caused PS externalization. However, SNCEE did not induce caspase activation or nuclear condensation/fragmentation suggesting that PS externalization was dissociated from the common apoptotic pathway. Dithiothreitol reversed SNCEE-induced S-nitrosylation, APLT inhibition, and PS externalization. SNCEE but not GSNO stimulated phagocytosis of HL-60 cells. Moreover, phagocytosis of target cells by lipopolysaccharide-stimulated macrophages was significantly suppressed by an NO. scavenger, DAF-2. Thus, macrophage-induced nitrosylation/oxidation plays an important role in cell clearance, and hence in the resolution of inflammation.     

Mikrobiologie in der Schule

In the project “The hidden world of bacteria” students from six classes from high schools around Salzburg work for two years together with staff members of the Research Department for Limnology, Mondsee (University of Innsbruck). The aim is to isolate new bacterial strains from surrounding inland waters and to describe them as new species or higher taxonomic units.