No abstract

Ellipsoidal Chlorella-like species are very common in all kinds of aquatic and terrestrial habitats, and often identified as Chlorella saccharophila or C. ellipsoidea. However, the taxonomic status of these species remains unclear, because they are not related to the type species of the genus, Chlorella vulgaris. In this study, 23 strains isolated from different habitats, were investigated using a polyphasic approach, i.e. morphology and reproduction, ecophysiology, and combined SSU and ITS rDNA sequences. Phylogenetic analyses clearly demonstrated that these isolates formed a monophyletic lineage within the green algal class Trebouxiophyceae. All strains were characterized by ellipsoidal cell shape, unequal autospores during reproduction, and parietal chloroplasts, as well as by the biochemical capability to synthesize and accumulate the rather unusual polyol, ribitol. Although ribitol is a typical stress metabolite involved in osmotic acclimation, it can also be used as a chemotaxonomic marker. Comparative growth measurements under different temperature regimes indicated similar optimum growth temperatures and maximum growth rates in all studied Chlorella-like species. However, these were different from those of C. vulgaris. We therefore propose to transfer all Chlorella-like strains related to Chlorella saccharophila and C. ellipsoidea to the genus Chloroidium Nadson and to emend its diagnosis. We propose four new combinations: Chloroidium saccharophilum comb. nov., Chloroidium ellipsoideum comb. nov., Chloroidium angusto-ellipsoideum comb. nov. and Chloroidium engadinensis comb. nov. In contrast, Chlorella ellipsoidea sensu Puncˇocha´rova´, which has other morphological and ecophysiological characters, should be assigned to the genus Pseudochlorella (P. pringsheimii comb. nov.).

A UV-absorbing mycosporine-like amino acid (324 nm-MAA), so far only known from the green macroalgal genus Prasiola (Trebouxiophyceae), was also identified in other morphologically diverse green algae closely related to Prasiola spp. in 18S rDNA phylogenies. Using HPLC, a second UVabsorbing compound was found only in Myrmecia incisa Reisigal among all studied strains. This substance showed an absorption maximum at 322 nm and hence was designated as putative 322 nm-MAA. Preliminary UV-exposure experiments indicated that all species containing one or the other MAA showed a strong accumulation of the respective compound, thus supporting their function as putative UV sunscreen. Both UV-absorbing substances were only identified in the studied members of the Trebouxiophyceae but were absent in members of the Ulvophyceae and Chlorophyceae. When mapped on an 18S rDNA phylogeny, the distribution of 324 nm-MAA was found to be scattered within the Trebouxiophyceae but was consistent with a distribution that follows phylogenetic patterns rather than ecological adaptations. The 324 nm-MAA was also detected in two phylogenetically related species from freshwater as well as from subaerial habitats, Watanabea reniformis Hanagata et al. and isolate UR7/5, which were phylogenetically independent of Prasiola and its closer allies. MAAs were absent in another Trebouxiophyceae clade comprising lichen photobionts (Coccomyxa pringsheimii Jaag) as well as freshwater picoplanktonic algae (Choricystis minor (Skuja) Fott). The data presented suggest a chemotaxonomic value of the 324 nm-MAA in green algal taxonomy. To address the paraphyly of the genus Myrmecia Printz as presently circumscribed, we propose the new combination Lobosphaera incisa.

A new method of respiration rate measurement based on oxygen luminescence quenching in sensor spots was evaluated for the first time for aquatic bacterial communities. The commonly used Winkler and Clark electrode methods to quantify oxygen concentration both require long incubation times, and the latter additionally causes signal drift due to oxygen consumption at the cathode. The sensor spots proved to be advantageous over those methods in terms of precise and quick oxygen measurements in natural bacterial communities, guaranteeing a respiration rate estimate during a time interval short enough to neglect variations in organism composition, abundance, and activity. Furthermore, no signal drift occurs during measurements, and respiration rate measurements are reliable even at low temperatures and low oxygen consumption rates. Both a natural bacterioplankton sample and a bacterial isolate from a eutrophic river were evaluated in order to optimize the new method for aquatic microorganisms. A minimum abundance of 2.2 ؋ 10 6 respiring cells ml

We applied different types of fluorescent markers to natural bacterioplankton from different aquatic systems to investigate microscopically the percentage of viable bacteria. To characterise viable bacteria, cell-specific respiration was measured by cyanoditolyltetrazolium chloride (CTC) reduction. Membrane integrity was investigated with 3 'dead cell' stains (SYTOX ® Green, propidium iodide and ethidium homodimer-2). Cellular enzyme activity was detected by artificial substrate analogs with a high cell retention (CellTracker™ Green CMFDA for cellular esterase and 7-amino-4-chloromethylcoumarin L-leucine amide, hydrochloride [CMAC-Leu] for cellular peptidase). The percentage of impermeable, i.e. morphologically intact, cells accounted for 22 to 81% of the total cell number at all locations. Although up to 48% of all bacteria were respiring, they averaged between 10 and 14% in freshwater, estuarine waters and in the Baltic Sea. The portion of esterase-positive cells correlated significantly with the concentrations of dissolved (DOC) and particulate organic carbon (POC) as well as with chlorophyll a (chl a) content. Cellular esterase was shown by this labelling technique in only 9% of freshwater, 12% of estuarine and 5% of Baltic Sea bacteria, . The percentages of bacteria with cellular peptidase were even lower with 6, 5 and 3%, respectively. The different amounts of intact and respiring bacteria as well as those with cellular hydrolytic enzyme activities require not only correct operational definitions of active and viable bacteria, but also the appropriate choice of fluorescent markers regarding the goals of investigation. Fluorescent labels for cellular hydrolytic enzymes will also provide a new tool to localise active cells in aggregates or on sediment particles, where, besides the respiration of organic carbon, hydrolysis of organic substances is an important conversion process.

The effects of artificial ultraviolet radiation [UVR; 8 W m(-2) ultraviolet-A (UVA), 0.4 W m(-2) ultraviolet-B (UVB)] on photosynthetic performance, growth and the capability to synthesise mycosporine-like amino acids (MAAs) was investigated in the aeroterrestrial green algae Stichococcus sp. and Chlorella luteoviridis forming biofilms on building facades, and compared with the responses of two green algae, from soil (Myrmecia incisa) and brackish water (Desmodesmus subspicatus). All species exhibited decreasing quantum efficiency (Fv/Fm) after 1-3 days exposure to UVR. After 8-12 days treatment, however, all aeroterrestrial isolates exhibited full recovery under UVA and UVA/B. In contrast, D. subspicatus showed only 80% recovery after treatment with UVB. While Stichococcus sp. and C. luteoviridis exhibited a broad tolerance in growth under all radiation conditions tested, M. incisa showed a significant decrease in growth rate after exposure to UVA and UVA/B. Similarly D. subspicatus grew with a reduced rate under UVA, but UVA/B led to full inhibition. Using HPLC, an UV-absorbing MAA (324 nm-MAA) was identified in Stichococcus sp. and C. luteoviridis. While M. incisa contained a specific 322 nm-MAA, D. subspicatus lacked any trace of such compounds. UV-exposure experiments indicated that all MAA-containing species are capable of synthesizing and accumulating these compounds, thus supporting their function as an UV-sunscreen. All data well explain the conspicuous ecological success of aeroterrestrial green algae in biofilms on facades. Biosynthesis and accumulation of MAAs under UVR seem to result in a reduced UV-sensitivity of growth and photosynthesis, which consequently may enhance survival in the environmentally harsh habitat.

The River Warnow is the drinking water source for the city of Rostock. Its eutrophic status is accompanied by high amounts of bacteria, which may reach up to 24 x 10(6) cells mL(-1) as recorded during a seasonal study in 2002. Because the river is eutrophic and also heavily loaded with organic matter, this burden is a problem for drinking water purification, as it must be removed completely to not trigger new bacterial growth in the pipeline network. Therefore, restoration measures in the river have to be planned, and bacteria have to be favored as decomposers. That includes the investigation of the physiological state of bacteria in situ. Viable and active cells in the lower reaches of River Warnow were estimated using a broad set of methods. Intact bacteria were investigated by the LIVE/DEAD BacLight bacterial viability kit, containing a mixture of permeant and impermeant nucleic acid stains. Cells with ribosomes were visualized by fluorescence in situ hybridization with the EUB338 oligonucleotide probe. Intact cells and ribosome-containing bacteria represented 24% of total numbers stained by 4'6,-diamidino-2-phenylindole (DAPI) or 66 and 62%, respectively, in relation to all bacteria visualized by the LIVE/DEAD kit. Both fractions were considered as viable, although the fraction of RIB + bacteria is most likely underestimated by the protocol applied. 5-Cyano-2,3-ditolyltetrazolium chloride (CTC) was applied to mark respiring bacteria. The esterase substrate CellTracker Green 5-chloromethylfluorescein diacetate showed cells with intracellular hydrolytic activity. Whereas 1.5% of DAPI-stained bacteria were observed as respiring, 3.8% exhibited intracellular hydrolytic activity on average. If these active fractions were calculated as the percentages of intact cells, much higher fractions of 5.4% were respiring and 16% hydrolytic. Temperature was a main factor influencing total and viable cell numbers simultaneously. The results confirm that there are different states of viable and active cells in natural bacterioplankton communities. However, it remains unclear why fractions of viable and active cells were rather low in this eutrophic river in comparison to similar waters. We recommend to carefully address cells as viable in contrast to nonviable, i.e., dead. As viable cells may be active or inactive with respect to many different activities, e.g., substrate uptake, respiration, hydrolysis, and cell deviation, it is necessary to choose the method to visualize active cells according to the question to be answered.

The photosynthetic performance of a microalgal biofilm colonizing a building facade was investigated between February and July 2004, with an emphasis on changing water availability and air humidity. The fluorimetric measurements of the quantum efficiency (F (v)/F (m)) indicated diurnal activity patterns. At most sampling dates the algal biofilm photosynthesized particularly in the morning and substantially less in the afternoon. As long as liquid water was present, the microalgae exhibited at least some degree of photosynthesis. However, F (v)/F (m) values never exceeded 0.4, pointing to slight photoinhibition or damage of the cells. Dried cells without photosynthesis could recover within minutes after artificial moistening. Three microalgal strains were isolated from aeroterrestrial biofilms and established as unialgal cultures. Their photosynthesis and growth were characterized under different air humidities and temperatures. Photosynthesis and growth of strain ROS 55/3 (Stichococcus sp.) showed similar patterns with decreasing relative air humidity. Positive growth and optimum photosynthesis were recorded at 100% relative air humidity. At air humidities below 93%, both processes were strongly inhibited. All studied strains grew between 1 and 30 degrees Celsius with optimum rates at 20-23 degrees Celsius, indicating eurythermal features. The data indicate that liquid water or 100% air humidity are the prerequisite for optimum photosynthesis and growth of aeroterrestrial microalgae. However, when dried and consequently inactive, these microorganisms can recover quickly if water is suddenly available, e.g., after rain events. These physiological capabilities explain well the ecological success of aeroterrestrial microalgae in occupying many man-made substrata such as building facades and roof tiles in urban areas.