Klára Řeháková

Morphological and molecular characterization of selected desert soil cyanobacteria: three species new to science including Mojavia pulchra gen. et sp. nov.

K. Řeháková, J.R. Johansen, D.A. Casamatta, L. Xuesong and J. Vincent. 2007. Morphological and molecular characterization of selected desert soil cyanobacteria: three species new to science including Mojavia pulchra gen. et sp.nov. Phycologia 46: 481–502. DOI: 10.2216/06-92.1 Four Nostocacean species from desert soils of the western United States, including the phycobiont of the lichen Collema tenax, were studied. Our strains could be forced into morphospecies previously described from Europe, but phylogenetic analysis indicated that they belonged in separate, distinct, and previously undescribed taxa. Partial 16S rRNA sequences of the strains CM1-VF10, CM1-VF14, CNP-AK1 and JT2-VF2 were determined and aligned with published Nostoc sequences from GenBank and our lab, as well as other Nostocales. All aligned sequences were analysed using parsimony, distance, and maximum likelihood methods, and trees based on three separate data sets were generated. Full 16S-23S internal transcribed spacer (ITS) regions were also characterized for our strains, and secondary structures of the ITS region were compared among these and N. commune and N. punctiforme. Intragenomic variability was documented among ITS regions in different operons for these taxa. One of the four strains (JT2-VF2) is distinct from Nostoc by both morphological and molecular criteria and is described as Mojavia pulchra gen. et sp. nov. Two other strains (CM1-VF10 and CM1-VF14) are described as Nostoc indistinguendum sp. nov. and Nostoc desertorum sp. nov., respectively. According to both morphological and molecular characteristics, the phycobiont of C. tenax is not N. commune, N. sphaericum or N. punctiforme as variously suggested in the lichenological literature, and the older name for this taxon, Nostoc lichenoides, is consequently validated in this paper.

POLYPHASIC CHARACTERIZATION OF THREE STRAINS OF ANABAENA RENIFORMIS AND APHANIZOMENON APHANIZOMENOIDES (CYANOBACTERIA) AND THEIR RECLASSIFICATION TO SPHAEROSPERMUM GEN. NOV. (INCL. ANABAENA KISSELEVIANA)(1).

Occurrences of rare cyanobacteria Anabaena reniformis Lemmerm. and Aphanizomenon aphanizomenoides (Forti) Horecká et Komárek were recently detected at several localities in the Czech Republic. Two monoclonal strains of An. reniformis and one strain of Aph. aphanizomenoides were isolated from distant localities and different sampling years. They were characterized by a combination of morphological, genetic, and biochemical approaches. For the first time, partial 16S rRNA gene sequences were obtained for these morphospecies. Based on this gene, all of these strains clustered separately from other planktonic Anabaena and Aphanizomenon strains. They appeared in a cluster with Cylindrospermopsis Seenaya et Subba Raju and Raphidiopsis F. E. Fritsch et M. F. Rich, clustered closely together with two An. kisseleviana Elenkin strains available from GenBank. A new generic entity was defined (Sphaerospermum gen. nov., with the type species S. reniforme, based on the traditional species An. reniformis). These results contribute significantly to the knowledge base about genetic heterogeneity among planktonic Anabaena–like and Aphanizomenon–like morphospecies. Accordingly, the subgenus Dolichospermum, previously proposed for the group of planktonic Anabaena, should be revaluated. Secondary metabolite profiles of the An. reniformis and Aph. aphanizomenoides strains differed considerably from 17 other planktonic Anabaena strains of eight morphospecies isolated from Czech water bodies. Production of puwainaphycin A was found in both of the An. reniformis strains. Despite the relatively short phylogenetic distance from Cylidrospermopsis, the production of cylindrospermopsin was not detected in any of our strains.

Testing the Stress-Gradient Hypothesis at the Roof of the World: Effects of the Cushion Plant Thylacospermum caespitosum on Species Assemblages

Many cushion plants ameliorate the harsh environment they inhabit in alpine ecosystems and act as nurse plants, with significantly more species growing within their canopy than outside. These facilitative interactions seem to increase with the abiotic stress, thus supporting the stress-gradient hypothesis. We tested this prediction by exploring the association pattern of vascular plants with the dominant cushion plant Thylacospermum caespitosum (Caryophyllaceae) in the arid Trans-Himalaya, where vascular plants occur at one of the highest worldwide elevational limits. We compared plant composition between 1112 pair-plots placed both inside cushions and in surrounding open areas, in communities from cold steppes to subnival zones along two elevational gradients (East Karakoram: 4850–5250 m and Little Tibet: 5350–5850 m). We used PERMANOVA to assess differences in species composition, Friedman-based permutation tests to determine individual species habitat preferences, species-area curves to assess whether interactions are size-dependent and competitive intensity and importance indices to evaluate plant-plant interactions. No indications for net facilitation were found along the elevation gradients. The open areas were not only richer in species, but not a single species preferred to grow exclusively inside cushions, while 39–60% of 56 species detected had a significant preference for the habitat outside cushions. Across the entire elevation range of T. caespitosum, the number and abundance of species were greater outside cushions, suggesting that competitive rather than facilitative interactions prevail. This was supported by lower soil nutrient contents inside cushions, indicating a resource preemption, and little thermal amelioration at the extreme end of the elevational gradient. We attribute the negative associations to competition for limited resources, a strong environmental filter in arid high-mountain environment selecting the stress-tolerant species that do not rely on help from other plants during their life cycle and to the fact the cushions do not provide a better microhabitat to grow in.

The effect of extreme rainfall on summer succession and vertical distribution of phytoplankton in a lacustrine part of a eutrophic reservoir

Abstract.During the summer of 2006, phytoplankton succession and vertical distribution were studied in the eutrophic Římov Reservoir (Czech Republic). Disturbances in the form of dramatic summer storms had a crucial effect on phytoplankton succession and its vertical distribution. Two extreme rainfalls substantially increased flushing rate, yielding a fairly short retention time within the reservoir that disrupted established thermal stratification. The first storm initiated the development of summer phytoplankton, while the second storm reversed the phytoplankton succession to an earlier stage. The measurement of vertical profiles of chlorophyll a showed that the phytoplankton were heterogeneously distributed in the water column and formed subsurface diatom maxima (SDM) over most of the summer. Biogenic silica deposition studied by PDMPO (2-(4-pyridyl)-5{[4-dimethylaminoethyl-aminocarbamyl)-methoxy] phenyl}oxazole) was measured to compare diatom growth rates over the season and between surface and depth of SDM. Diatoms deposited 20 times more silica at the surface than at the SDM depth, although only a half of diatom population was involved in the silica deposition. Maximum rates of silica deposition were measured in samples just after both summer storms events. When diatoms dominated the phytoplankton, the amounts of available Si and P in water were significantly correlated (r2 = 0.81, p < 0.001, df = 42). Outliers from the linear relationship were found only during a period when substantial picocyanobacterial population developed in the reservoir. Due to their lack of Si requirements, ambient Si concentration markedly increased while P was depleted to growth limiting levels. Temporal variations in Si:P ratio therefore illustrate the importance of resource competition in phytoplankton seasonal succession.

Polyphasic characterization of eight planktonic Anabaena strains (Cyanobacteria) with reference to the variability of 61 Anabaena populations observed in the field

The plasticity of morphological features used for single morphospecies identification was studied under varied experimental conditions (temperature, light, nitrogen, phosphorus) in eight planktonic Anabaena strains. The strains represented all of the morphospecies with coiled trichomes commonly occurring in Central Europe (two strains of A. mendotae & A. sigmoidea complex, two A. lemmermannii strains, two A. flos-aquae strains, and two strains of A. circinalis & A. crassa complex). Significant effects of the growth conditions on vegetative cell dimensions were observed in seven strains, and P concentration was the main influencing factor in most cases (six strains). Significant effect of an environmental factor (P) on akinete morphology was found in only one strain. Experimentally assessed temperature and light growth optima were specific for each strain and were not consistent with the taxonomic affiliation of the strains. Morphologies of the Anabaena strains studied were compared with the field morphologies of 61 Anabaena populations of eight morphospecies observed in the Czech Republic. The range of morphological variability of single strains under the experimental conditions spanned the total variability of the populations of relevant morphospecies observed in the field. Delimitations and proper descriptions of the morphospecies are discussed in the light of partial 16S rRNA gene sequences of the studied strains.

Morphological variability in selected heterocystous cyanobacterial strains as a response to varied temperature, light intensity and medium composition.

The effect of temperature, light and nutrient composition on morphological traits was determined in seven nostocacean cyanobacteria (Anabaena planctonica, A. sphaerica var. conoidea, A. spiroides, Aphanizomenon gracile, Nostoc sp., Scytonema sp., and Tolypothrix sp.). Their morphological variability was high but only some of the features showed changes reflecting varied growth conditions. The frequency of heterocyst occurrence decreased with increasing nitrogen concentration. Within the range studied, the effect of temperature on heterocyst frequency of Tolypothrix sp. and planktonic Anabaena strains could be fitted by a normal curve with a clear optimum while linear correlation was found in Aphanizomenon gracile. T-and S-type branching was observed in both Scytonema sp. and Tolypothrix sp. strains. T-type branching was found to be markedly dependent on nitrogen concentration. The abundance of necridic cells of Tolypothrix sp. increased linearly with temperature and light intensity. Regularity of trichome coiling of A. spiroides depended on culture medium, suggesting that nutrient composition may be the main controlling factor. In contrast, the effect of the experimental conditions on the dimensions of vegetative cells and heterocysts was weak. Their variability was markedly higher within each experimental treatment than between treatments.

Cushions of Thylacospermum caespitosum (Caryophyllaceae) do not facilitate other plants under extreme altitude and dry conditions in the north-west Himalayas

BACKGROUND Cushion plants are commonly considered as keystone nurse species that ameliorate the harsh conditions they inhabit in alpine ecosystems, thus facilitating other species and increasing alpine plant biodiversity. A literature search resulted in 25 key studies showing overwhelming facilitative effects of different cushion plants and hypothesizing greater facilitation with increased environmental severity (i.e. higher altitude and/or lower rainfall). At the same time, emerging ecological theory alongside the cushion-specific literature suggests that facilitation might not always occur under extreme environmental conditions, and especially under high altitude and dryness. METHODS To assess these hypotheses, possible nursing effects of Thylacospermum caespitosum (Caryophyllaceae) were examined at extremely high altitude (5900 m a.s.l.) and in dry conditions (precipitation <100 mm year(-1)) in Eastern Ladakh, Trans-Himalaya. This is, by far, the highest site, and the second driest, at which the effects of cushions have been studied so far. KEY RESULTS In accordance with the theoretical predictions, no nursing effects of T. caespitosum on other alpine plants were detected. The number and abundance of species were greater outside cushions than within and on the edge of cushions. None of the 13 species detected was positively associated with cushions, while nine of them were negatively associated. Plant diversity increased with the size of the area sampled outside cushions, but no species-area relationship was found within cushions. CONCLUSIONS The results support the emerging theoretical prediction of restricted facilitative effects under extreme combinations of cold and dryness, integrating these ideas in the context of the ecology of cushion plants. This evidence suggests that cases of missing strong facilitation are likely to be found in other extreme alpine conditions.

Community structure of soil phototrophs along environmental gradients in arid Himalaya

The well-developed biological soil crusts cover up to 40% of the soil surface in the alpine and subnival zones of the Tibetan Plateau, accounting for a vast area of Asia. We investigated the diversity and biomass of the phototrophic part (Cyanobacteria) of the microbial community inhabiting biological soil crusts and uncrusted soils in their surroundings on the elevation gradient of 5200-5900 m a.s.l. The influence of soil physico-chemical properties on phototrophs was studied. The ability of high-altitude phototrophs to fix molecular nitrogen was also determined under laboratory conditions. The biological soil crust phototroph community did not differ from that living in uncrusted soil in terms of the species composition, but the biomass is three-to-five times higher. An increasing trend in the cyanobacterial biomass from the biological soil crusts with elevation was observed, with the genera Nostoc spp., Microcoleus vaginatus and Phormidium spp. contributing to this increase. Based on the laboratory experiments, the highest nitrogenase activity was recorded in the middle elevations, and the rate of nitrogen fixation was not correlated with the cyanobacterial biomass.

Methanogens at the top of the world: occurrence and potential activity of methanogens in newly deglaciated soils in high-altitude cold deserts in the Western Himalayas.

Methanogens typically occur in reduced anoxic environments. However, in recent studies it has been shown that many aerated upland soils, including desert soils also host active methanogens. Here we show that soil samples from high-altitude cold deserts in the western Himalayas (Ladakh, India) produce CH4 after incubation as slurry under anoxic conditions at rates comparable to those of hot desert soils. Samples of matured soil from three different vegetation belts (arid, steppe, and subnival) were compared with younger soils originating from frontal and lateral moraines of receding glaciers. While methanogenic rates were higher in the samples from matured soils, CH4 was also produced in the samples from the recently deglaciated moraines. In both young and matured soils, those covered by a biological soil crust (biocrust) were more active than their bare counterparts. Isotopic analysis showed that in both cases CH4 was initially produced from H2/CO2 but later mostly from acetate. Analysis of the archaeal community in the in situ soil samples revealed a clear dominance of sequences related to Thaumarchaeota, while the methanogenic community comprised only a minor fraction of the archaeal community. Similar to other aerated soils, the methanogenic community was comprised almost solely of the genera Methanosarcina and Methanocella, and possibly also Methanobacterium in some cases. Nevertheless, ~103 gdw−1 soil methanogens were already present in the young moraine soil together with cyanobacteria. Our results demonstrate that Methanosarcina and Methanocella not only tolerate atmospheric oxygen but are also able to survive in these harsh cold environments. Their occurrence in newly deglaciated soils shows that they are early colonizers of desert soils, similar to cyanobacteria, and may play a role in the development of desert biocrusts.

POLYPHASIC CHARACTERIZATION OF DOLICHOSPERMUM SPP. AND SPHAEROSPERMOPSIS SPP. (NOSTOCALES, CYANOBACTERIA): MORPHOLOGY, 16S rRNA GENE SEQUENCES AND FATTY ACID AND SECONDARY METABOLITE PROFILES(1).

The genera Dolichospermum (Ralfs ex Bornet et Flahault) Wacklin, L. Hoffm. et Komárek and Sphaerospermopsis Zapomělová, Jezberová, Hrouzek, Hisem, K. Řeháková et Komárk.‐Legn. represent a highly diversified group of planktonic cyanobacteria that have been recently separated from the traditional genus Anabaena Bory ex Bornet et Flahault. In this study, morphological diversity, phylogeny of the 16S rRNA gene, production of fatty acids, and secondary metabolite profiles were evaluated in 33 strains of 14 morphospecies isolated from the Czech Republic. Clustering of the strains based on 16S rRNA gene sequences corresponded to wider groups of species in terms of morphology. The overall secondary metabolite and fatty acid profiles, however, were not correlated to each other and neither were they correlated to the 16S rRNA phylogeny nor the morphology of the strains. Nevertheless, a minor part of the detected secondary metabolites (19% of all compounds) was present only in close relatives and can be thus considered as autapomorphic features.