Seasonal Dynamics of Zygnema (Zygnematophyceae) Mats from the Austrian Alps.

Filamentous green algae of the genus Zygnema are an essential part of hydro-terrestrial ecosystems. Despite several studies on their resistance to natural stresses, little is known about the composition of their assemblages and the changes they undergo over time. Two sites at altitudes above 2200 m a.s.l. in the Austrian Alps were selected for a 2-year observation period and sampled five times. Molecular phylogenetic analysis of the 152 isolated strains of Zygnema sp. was performed based on the rbcL and trnG sequences. Seven genotypes were found at these sites during the samplings, but their proportion varied throughout the seasons. The site with a more stable water regime also had a more stable representation of genotypes, in contrast to the site with fluctuating water availability. The mats formed resistant pre-akinetes at the end of the season with reduced photosynthetic activity. Contrary to expectations, the mats were not exposed to extremely cold temperatures in winter due to snow cover. Some genotypes have been previously observed at this site, indicating that the population composition is stable. This work highlights the importance of resistant pre-akinetes in surviving winter conditions, the ability of algae to re-establish mats, and the need to address the hidden diversity of the genus Zygnema.

The conjugating green alga Zygnema sp. (Zygnematophyceae) from the Arctic shows high frost tolerance in mature cells (pre-akinetes).

Green algae of the genus Zygnema form extensive mats and produce large amounts of biomass in shallow freshwater habitats. Environmental stresses including freezing may perturb these mats, which usually have only annual character. To estimate the limits of survival at subzero temperatures, freezing resistance of young Zygnema sp. (strain MP2011Skan) cells and pre-akinetes was investigated. Young, 2-week-old cultures were exposed to temperatures of 0 to - 14 °C at 2-K steps, whereas 8-month-old cultures were frozen from - 10 to - 70 °C at 10-K intervals. Cell viability after freezing was determined by 0.1% auramine O vital fluorescence staining and measurements of the effective quantum yield of photosystem II (ФPSII). At - 8 °C, the young vegetative cells were unable to recover from severe frost damage. But temperatures even slightly below zero (- 2 °C) negatively affected the cells' physiology. Single pre-akinetes could survive even at - 70 °C, but their LT50 value was - 26.2 °C. Severe freezing cytorrhysis was observed via cryo-microscopy at - 10 °C, a temperature found to be lethal for young cells. The ultrastructure of young cells appeared unchanged at - 2 °C, but severe damage to biomembranes and formation of small foamy vacuoles was observed at - 10 °C. Pre-akinetes did not show ultrastructural changes at - 20 °C; however, vacuolization increased, and gas bubbles appeared at - 70 °C. Our results demonstrate that the formation of pre-akinetes increases freezing resistance. This adaptation is crucial for surviving the harsh temperature conditions prevailing in the High Arctic in winter and a key feature in seasonal dynamics of Zygnema sp.

Molecular and morphological diversity of Zygnema and Zygnemopsis (Zygnematophyceae, Streptophyta) from Svalbard (High Arctic).

Filamentous conjugating green microalgae (Zygnematophyceae, Streptophyta) belong to the most common primary producers in polar hydro-terrestrial environments such as meltwater streamlets and shallow pools. The mats formed by these organisms are mostly composed of sterile filaments with Zygnema morphology, but the extent of their diversity remains unknown. Traditional taxonomy of this group is based on reproductive morphology, but sexual reproduction (conjugation and formation of resistant zygospores) is very rare in extreme conditions. In the present study we gave the first record of zygospore formation in Svalbard field samples, and identified conjugating filaments as Zygnemopsis lamellata and Zygnema cf. calosporum. We applied molecular phylogeny to study genetic diversity of sterile Zygnema filaments from Svalbard in the High Arctic. Based on analysis of 143 rbcL sequences, we revealed a surprisingly high molecular diversity: 12 Arctic Zygnema genotypes and one Zygnemopsis genotype were found. In addition, we characterized individual Arctic genotypes based on cell width and chloroplast morphology using light and confocal laser scanning microscopy. Our findings highlight the importance of a molecular approach when working with sterile filamentous Zygnematophyceae, as hidden diversity might be very beneficial for adaptation to harsh environmental conditions, and experimental results could be misinterpreted when hidden diversity is neglected.

Arctic, Antarctic, and temperate green algae Zygnema spp. under UV-B stress: vegetative cells perform better than pre-akinetes.

Species of Zygnema form macroscopically visible mats in polar and temperate terrestrial habitats, where they are exposed to environmental stresses. Three previously characterized isolates (Arctic Zygnema sp. B, Antarctic Zygnema sp. C, and temperate Zygnema sp. S) were tested for their tolerance to experimental UV radiation. Samples of young vegetative cells (1 month old) and pre-akinetes (6 months old) were exposed to photosynthetically active radiation (PAR, 400-700 nm, 400 µmol photons m-2 s-1) in combination with experimental UV-A (315-400 nm, 5.7 W m-2, no UV-B), designated as PA, or UV-A (10.1 W m-2) + UV-B (280-315 nm, 1.0 W m-2), designated as PAB. The experimental period lasted for 74 h; the radiation period was 16 h PAR/UV-A per day, or with additional UV-B for 14 h per day. The effective quantum yield, generally lower in pre-akinetes, was mostly reduced during the UV treatment, and recovery was significantly higher in young vegetative cells vs. pre-akinetes during the experiment. Analysis of the deepoxidation state of the xanthophyll-cycle pigments revealed a statistically significant (p < 0.05) increase in Zygnema spp. C and S. The content of UV-absorbing phenolic compounds was significantly higher (p < 0.05) in young vegetative cells compared to pre-akinetes. In young vegetative Zygnema sp. S, these phenolic compounds significantly increased (p < 0.05) upon PA and PAB. Transmission electron microscopy showed an intact ultrastructure with massive starch accumulations at the pyrenoids under PA and PAB. A possible increase in electron-dense bodies in PAB-treated cells and the occurrence of cubic membranes in the chloroplasts are likely protection strategies. Metabolite profiling by non-targeted RP-UHPLC-qToF-MS allowed a clear separation of the strains, but could not detect changes due to the PA and PAB treatments. Six hundred seventeen distinct molecular masses were detected, of which around 200 could be annotated from databases. These results indicate that young vegetative cells can adapt better to the experimental UV-B stress than pre-akinetes.