Effects of a cyanobacterial extract containing-anatoxin-a(s) on the cardiac rhythm of Leurolestes circunvagans

This work presents the effects of an anatoxin-a(s)-containing extract on a cockroach semi-isolated heart preparation and the results supporting the extract s biological activity on acetylcholinesterase (purified from ell). The presence of the toxin in cyanobacterial strains Anabaena spiroides (ITEP-024, ITEP-025 and ITEP-026) isolated from the Tapacurá reservoir in Pernambuco, Brazil, was confirmed by means of liquid chromatography coupled to an ion-trap mass spectrometer. The anticholinesterase activity was assessed biochemically by the Ellman test and was confirmed by measuring the cockroach s heart rate. The concentration of the extract containing the tested anatoxin-a(s) (antx-a(s)) (10, 16 and 100 μg.μL-1) inhibited the eel acetylcholinesterase (AChE) by more than 90%. The cockroach cardiac frequency increased by a maximum of about 20% within 29 min after the addition of 2.5x10³ μg of extract containing antxa (s).g-1 bw (n=9, p<0.05). Our results strongly indicate that antx-a(s) is capable of exerting biological effects on cockroach, indicating that more research might be conducted to determine its role in the environment, especially on insects.

Three-dimensional electron microscopy of myriapod hemocyanin, planorbid snail acetylcholine-binding protein and cyanobacterial Vipp1

Three-dimensional electron microscopy (3-D EM) provides a framework for the analysis of large protein quaternary structures. The advantage over the generally higher resolving meth- od of X-ray crystallography is the embedding of the proteins in their physiological environ- ment. However, results of the two methods can be combined to obtain superior structural information. In this work, three different protein types – (i) Myriapod hemocyanin, (ii) vesi- cle-inducing protein in plastids 1 (Vipp1) and (iii) acetylcholine-binding protein (AChBP) – were structurally analyzed by 2-D and 3-D EM and, where possible, functionally interpreted.rnMyriapod hemocyanins have been previously shown to be 6x6-meric assemblies that, in case of Scutigera coleoptrata hemocyanin (ScoHc), show two 3x6-mer planes whith a stag- gering angle of approximately 60°. Here, previously observed structural differences between oxy- and deoxy-ScoHc could be substantiated. A 4° rotation between hexamers of two dif- ferent 3x6-mer planes was measured, which originates at the most central inter-hexamer in- terface. Further information about allosteric behaviour in myriapod hemocyanin was gained by analyzing Polydesmus angustus hemocyanin (PanHc), which shows a stable 3x6-mer and divergent histidine patterns in the inter-hexamer interfaces when compared to ScoHc. Both findings would conclusively explain the very different oxygen binding properties of chilopod and diplopod hemocyanin.rnVipp1 is a protein found in cyanobacteria and higher plants which is essential for thyla- koid membrane function and forms highly variable ring-shaped structures. In the course of this study, the first 3-D analysis of Vipp1 was conducted and yielded reconstructions of six differently sized Vipp1 rings from negatively stained images at resolutions between 20 to 30 Å. Furthermore, mutational analyses identified specific N-terminal amino acids that are essential for ring formation. On the basis of these analyses and previously published results, a hypothetical model of the Vipp1 tertiary and quaternary structure was generated.rnAChBP is a water-soluble protein in the hemolymph of mollusks. It is a structural and functional homologue of the ligand-binding domain of nicotinic acetylcholine receptors. For the freshwater snail Biomphalaria glabrata, we previously described two types of AChBP (BgAChBP1 and BgAChBP2). In this work, a 6 Å 3-D reconstruction of native BgAChBP is presented, which shows a dodecahedral assembly that is unprecedented for an AChBP. Single particle analysis of recombinantely expressed BgAChBP types led to preliminary results show- ing a dodecahedral assembly of BgAChBP1 and a dipentameric assembly of BgAChBP2. This indicates divergent biological functions of the two types.

Mats of psychrophilic thiotrophic bacteria associated with cold seeps of the Barents Sea

This study focused on the bacterial diversity associated with microbial mats of deep-sea cold seeps at the Norwegian continental margin. Study sites included the Storegga and Nyegga areas as well as the Håkon Mosby mud volcano, where the mats occurred at temperatures permanently close to the freezing point of seawater. Two visually different mat types, i.e. small gray mats and extensive white mats, were studied with the aim to determine the identity of the mat-forming sulfide oxidizers, and to investigate which environmental factors (e.g. sulfate reduction and methane oxidation rates) shown here could explain the observed diversity. Sequence data have been submitted to the EMBL database under accession No. FR847864-FR847887 (giant sulfur bacteria), No. FR827864 (Menez Gwen filament; see Supplementary Material) and No. FR875365-FR877509 (except FR875905; remaining partial sequences).

Microsensor measurements in photosynthetic microbial mats

Based on combined microsensor measurements of irradiance, temperature and O2, we compared light energy budgets in photosynthetic microbial mats, with a special focus on the efficiency of light energy conservation by photosynthesis. The euphotic zones in the three studied mats differed in their phototrophic community structure, pigment concentrations and thickness. In all mats, < 1% of the absorbed light energy was conserved via photosynthesis at high incident irradiance, while the rest was dissipated as heat. Under light-limiting conditions, the photosynthetic efficiency reached a maximum, which varied among the studied mats between 4.5% and 16.2% and was significantly lower than the theoretical maximum of 27.7%. The maximum efficiency correlated linearly with the light attenuation coefficient and photopigment concentration in the euphotic zone. Higher photosynthetic efficiency was found in mats with a thinner and more densely populated euphotic zone. Microbial mats exhibit a lower photosynthetic efficiency compared with ecosystems with a more open canopy-like organization of photosynthetic elements, where light propagation is not hindered to the same extent by photosynthetically inactive components; such components contributed about 40-80% to light absorption in the investigated microbial mats, which is in a similar range as in oceanic planktonic systems.

Biogeochemistry (speciation and isotopes of S and C) in bottom sediments, water and bacterial mats from the White Sea

This publication presents results of microbiological and biogeochemical studies in the White Sea. Material was obtained during a series of expeditions in 1999-2002. The studies were carried out in the open part of the White Sea, in the Onega, Dvina and Kandalaksha Bays, as well as in the intertidal zone of the Kandalaksha Bay. Quantitative characteristics of activity of microbial processes in waters and bottom sediments of the White Sea were obtained. The total number of bacteria was equal to 150000-800000 cells/ml, and intensity of dark CO2 assimilation was equal to 0.9-17 µg C/l/day. Bacterial sulfate reduction was equal to 3-150 mg S/m**2/day, and methane formation and oxidation was equal to 13-6840 and 20-14650 µl CH4/m**2/day, respectively. Extremely high values of intensity of all principal microbial processes were found in intertidal sediments rich in organic matter: under decomposing macrophytes, in local pits at the lower intertidal boundary, and in the mouth of a freshwater brook. Average hydrogen sulfide production in highly productive intertidal sediments was 1950-4300 mg S/m**2/day, methane production was 0.5-8.7 ml CH4/m**2/day, and intensity of methane oxidation was up to 17.5 ml CH4/m**2/day. Calculations performed with account for areas occupied by microlandscapes of increased productivity showed that diurnal production of H2S and CH4 per 1 km**2 of the intertidal zone (August) was estimated as 60.8-202 kg S/km**2/day and 192-300 l CH4/km**2/day, respectively.

Pigment, carbohydrate and protein content of microbial mats from northern Victoria Land lake sediments

The composition of algal pigments and extracellular polymeric substances (EPS) was determined in microbial mats from two lakes in Victoria Land (Continental Antarctica) with different lithology and environmental features. The aim was to expand knowledge of benthic autotrophic communities in Antarctic lacustrine ecosystems, providing reference data for future assessment of possible changes in environmental conditions and freshwater communities. The results of chemical analyses were supported by microscopy observations. Pigment profiles showed that filamentous cyanobacteria are dominant in both lakes. Samples from the water body at Edmonson Point had greater biodiversity, fewer pigments and lower EPS ratios than those from the lake at Kar Plateau. Differences in mat composition and in pigment and EPS profile between the two lakes are discussed in terms of local environmental conditions such as lithology, ice-cover and UV radiation. The present study suggests that a chemical approach could be useful in the study of benthic communities in Antarctic lakes and their variations in space and time.

Chloroflexus spp. and Sulfurihydrogenibium spp. in microbial mats of the Nakabusa hot spring, Japan

Microbial mats develop in a wide range of aquatic habitats, such as geothermal hot springs, hypersaline ponds, marine cold seeps or hydrothermal vents. The Nakabusa hot spring is located in the Nagano Prefecture, Japan (36.3875N, 137.75E), dense olive-green microbial mats develop in regions where the slightly alkaline, sulfidic effluent has cooled to 65°C. The microbial community of such mats was analyzed by focusing on the diversity, as well as the in situ distribution and function of bacteria involved in sulfur cycling. Microbial mat samples were kept in sterile plastic tubes (for molecular analysis) or glass bottles completely filled with hot spring water to avoid oxidation. Samples were transferred to the laboratory on ice and used for physiological experiments within 8h. Quantification of cell biovolumes was carried out based on images of mat sections hybridized with Sulfurihydrogenibium- and Chloroflexi-specific probes, and stained with DAPI. In situ hybridizations (CARD-FISH) of thin matsections showed a heterogeneous vertical distribution of Sulfurihydrogenibium and Chloroflexus. Sulfurihydrogenibium dominated near the mat surface (50% of the total mat biovolume), while Chloroflexus dominated in deeper layers (up to 64% of the total mat biovolume).

Biogeochemical investigation of microbial mats from the hypoxic region of the Crimean Shelf (Black Sea)

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998. High-resolution ex situ sulfide and pH microprofiles, were assessed only for station MSM15/1_492_PUC1. "in mat 1, 2 and 3" refers to 3 different profiles in 3 different spots of the microbial mat, whereas "outside mat", a profile outside the microbial mat.