Lars Wörmer

Multi-scale strategies for the monitoring of freshwater cyanobacteria: reducing the sources of uncertainty.

Cyanobacterial blooms are a frequent phenomenon in eutrophic freshwaters worldwide and are considered potential hazards to ecosystems and human health. Monitoring strategies based on conventional sampling often fail to cover the marked spatial and temporal variations in cyanobacterial distribution and fluctuating toxin concentrations inherent to cyanobacterial blooms. To deal with these problems, we employed a multi-scale approach for the study of a massive Microcystis bloom in Tajo River (Spain) utilizing 1) remote sensing techniques, 2) conventional water sampling and 3) analysis of chemotypical subpopulations. Tajo River at the study area is influenced by high temperatures waters diverted upstream from a nuclear power plant, the presence of a dam downstream and a high nutrient load, which provide optimal conditions for massive cyanobacterial proliferation. MERIS imagery revealed high Chl-a concentrations that rarely fell below 20 μg L(-1) and moderate spatiotemporal variations throughout the study period (March-November 2009). Although the phytoplanktonic community was generally dominated by Microcystis, sampling points highly differed in cyanobacterial abundance and community composition. Microcystin (MC) concentrations were highly heterogeneous, varying up to 3 orders of magnitude among sampling points, exceeding in some cases WHO guideline values for drinking and also for recreational waters. The analysis of single colonies by MALDI-TOF MS revealed differences in the proportion of MC-producing colonies among points. The proportion of toxic colonies showed a highly significant linear correlation with total MC: biovolume ratio (r(2) = 0.9; p < 0.001), evidencing that the variability in toxin concentrations can be efficiently addressed by simple analysis of subpopulations. We propose implementing a multi-scale monitoring strategy that allows covering the spatiotemporal heterogeneities in both cyanobacterial distribution (remote sensing) and MC concentrations (subpopulation analysis) and thereby reduce the main sources of uncertainty in the assessment of the risks associated to bloom events.

Novel cardiolipins from uncultured methane-metabolizing archaea.

Novel cardiolipins from Archaea were detected by screening the intact polar lipid (IPL) composition of microbial communities associated with methane seepage in deep-sea sediments from the Pakistan margin by high-performance liquid chromatography electrospray ionization mass spectrometry. A series of tentatively identified cardiolipin analogues (dimeric phospholipids or bisphosphatidylglycerol, BPG) represented 0.5% to 5% of total archaeal IPLs. These molecules are similar to the recently described cardiolipin analogues with four phytanyl chains from extreme halophilic archaea. It is worth noting that cardiolipin analogues from the seep archaeal communities are composed of four isoprenoidal chains, which may contain differences in chain length (20 and 25 carbon atoms) and degrees of unsaturation and the presence of a hydroxyl group. Two novel diether lipids, structurally related to the BPGs, are described and interpreted as degradation products of archaeal cardiolipin analogues. Since archaeal communities in seep sediments are dominated by anaerobic methanotrophs, our observations have implications for characterizing structural components of archaeal membranes, in which BPGs are presumed to contribute to modulation of cell permeability properties. Whether BPGs facilitate interspecies interaction in syntrophic methanotrophic consortia remains to be tested.

Methanothermobacter thermautotrophicus modulates its membrane lipids in response to hydrogen and nutrient availability

Methanothermobacter thermautotrophicus strain ΔH is a model hydrogenotrophic methanogen, for which extensive biochemical information, including the complete genome sequence, is available. Nevertheless, at the cell membrane lipid level, little is known about the responses of this archaeon to environmental stimuli. In this study, the lipid composition of M. thermautotrophicus was characterized to verify how this archaeon modulates its cell membrane components during growth phases and in response to hydrogen depletion and nutrient limitation (potassium and phosphate). As opposed to the higher abundance of phospholipids in the stationary phase of control experiments, cell membranes under nutrient, and energy stress were dominated by glycolipids that likely provided a more effective barrier against ion leakage. We also identified particular lipid regulatory mechanisms in M. thermautotrophicus, which included the accumulation of polyprenols under hydrogen-limited conditions and an increased content of sodiated adducts of lipids in nutrient-limited cells. These findings suggest that M. thermautotrophicus intensely modulates its cell membrane lipid composition to cope with energy and nutrient availability in dynamic environments.

Ultra-high-resolution paleoenvironmental records via direct laser-based analysis of lipid biomarkers in sediment core samples

Significance Lipid biomarkers in geological samples are important informants regarding past environments and ecosystems. Conventional biomarker analysis is labor intensive and requires relatively large sediment or rock samples; temporal resolution is consequently low. Here, we present an approach that has the potential to revolutionize paleoenvironmental biomarker research; it avoids wet-chemical sample preparation and enables analysis of biomarkers directly on sediment cores at submillimeter spatial resolution. Our initial application to a sediment core deposited during the Holocene climate optimum in the Mediterranean Sea reveals a new view of how small-scale variations in lipid distribution are integrated into commonly reported signals obtained by conventional analysis and demonstrates a strong influence of the ∼200-y de Vries solar cycle on sea-surface temperatures and planktonic archaeal ecology. Marine microorganisms adapt to their habitat by structural modification of their membrane lipids. This concept is the basis of numerous molecular proxies used for paleoenvironmental reconstruction. Archaeal tetraether lipids from ubiquitous marine planktonic archaea are particularly abundant, well preserved in the sedimentary record and used in several molecular proxies. We here introduce the direct, extraction-free analysis of these compounds in intact sediment core sections using laser desorption ionization (LDI) coupled to Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). LDI FTICR-MS can detect the target lipids in single submillimeter-sized spots on sediment sections, equivalent to a sample mass in the nanogram range, and could thus pave the way for biomarker-based reconstruction of past environments and ecosystems at subannual to decadal resolution. We demonstrate that ratios of selected archaeal tetraethers acquired by LDI FTICR-MS are highly correlated with values obtained by conventional liquid chromatography/MS protocols. The ratio of the major archaeal lipids, caldarchaeol and crenarchaeol, analyzed in a 6.2-cm intact section of Mediterranean sapropel S1 at 250-µm resolution (∼4-y temporal resolution), provides an unprecedented view of the fine-scale patchiness of sedimentary biomarker distributions and the processes involved in proxy signal formation. Temporal variations of this lipid ratio indicate a strong influence of the ∼200-y de Vries solar cycle on reconstructed sea surface temperatures with possible amplitudes of several degrees, and suggest signal amplification by a complex interplay of ecological and environmental factors. Laser-based biomarker analysis of geological samples has the potential to revolutionize molecular stratigraphic studies of paleoenvironments.

Transitory microbial habitat in the hyperarid Atacama Desert

Significance It has remained an unresolved question whether microorganisms recovered from the most arid environments on Earth are thriving under such extreme conditions or are just dead or dying vestiges of viable cells fortuitously deposited by atmospheric processes. Based on multiple lines of evidence, we show that indigenous microbial communities are present and temporally active even in the hyperarid soils of the Atacama Desert (Chile). Following extremely rare precipitation events in the driest parts of this desert, where rainfall often occurs only once per decade, we were able to detect episodic incidences of biological activity. Our findings expand the range of hyperarid environments temporarily habitable for terrestrial life, which by extension also applies to other planetary bodies like Mars. Traces of life are nearly ubiquitous on Earth. However, a central unresolved question is whether these traces always indicate an active microbial community or whether, in extreme environments, such as hyperarid deserts, they instead reflect just dormant or dead cells. Although microbial biomass and diversity decrease with increasing aridity in the Atacama Desert, we provide multiple lines of evidence for the presence of an at times metabolically active, microbial community in one of the driest places on Earth. We base this observation on four major lines of evidence: (i) a physico-chemical characterization of the soil habitability after an exceptional rain event, (ii) identified biomolecules indicative of potentially active cells [e.g., presence of ATP, phospholipid fatty acids (PLFAs), metabolites, and enzymatic activity], (iii) measurements of in situ replication rates of genomes of uncultivated bacteria reconstructed from selected samples, and (iv) microbial community patterns specific to soil parameters and depths. We infer that the microbial populations have undergone selection and adaptation in response to their specific soil microenvironment and in particular to the degree of aridity. Collectively, our results highlight that even the hyperarid Atacama Desert can provide a habitable environment for microorganisms that allows them to become metabolically active following an episodic increase in moisture and that once it decreases, so does the activity of the microbiota. These results have implications for the prospect of life on other planets such as Mars, which has transitioned from an earlier wetter environment to today’s extreme hyperaridity.

Important roles for membrane lipids in haloarchaeal bioenergetics.

Recent advances in lipidomic analysis in combination with various physiological experiments set the stage for deciphering the structure-function of haloarchaeal membrane lipids. Here we focused primarily on changes in lipid composition of Haloferax volcanii, but also performed a comparative analysis with four other haloarchaeal species (Halobacterium salinarum, Halorubrum lacusprofundi, Halorubrum sodomense and Haloplanus natans) all representing distinctive cell morphologies and behaviors (i.e., rod shape vs. pleomorphic behavior). Common to all five haloarchaea, our data reveal an extraordinary high level of menaquinone, reaching up to 72% of the total lipids. This ubiquity suggests that menaquinones may function beyond their ordinary role as electron and proton transporter, acting simultaneously as ion permeability barriers and as powerful shield against oxidative stress. In addition, we aimed at understanding the role of cations interacting with the characteristic negatively charged surface of haloarchaeal membranes. We propose for instance that by bridging the negative charges of adjacent anionic phospholipids, Mg2+ acts as surrogate for cardiolipin, a molecule that is known to control curvature stress of membranes. This study further provides a bioenergetic perspective as to how haloarchaea evolved following oxygenation of Earths atmosphere. The success of the aerobic lifestyle of haloarchaea includes multiple membrane-based strategies that successfully balance the need for a robust bilayer structure with the need for high rates of electron transport - collectively representing the molecular basis to inhabit hypersaline water bodies around the planet.

A highly asynchronous developmental program triggered during germination of dormant akinetes of filamentous diazotrophic cyanobacteria

Germination of akinetes of filamentous heterocyst-forming cyanobacteria of the order Nostocales is an essential process that ensures survival and recolonization after long periods of unfavorable conditions, as desiccation, cold and low light. We studied the morphological, physiological and metabolic changes that occur during germination of akinetes in two model species of cell differentiation, Anabaena variabilis ATCC 29413 and Nostoc punctiforme ATCC 29133, which live in different habitats. We characterized the akinete envelopes and showed their similarity to envelopes of N2-fixing heterocysts. Akinete germination started inside the envelopes and was dependent on light intensity but independent of nitrogen supply. During the germination of A. variabilis akinetes, cell division and heterocyst differentiation were highly accelerated. The energy for cell division was initially supplied by respiration of glycogen and subsequently by photosynthesis. By contrast, during germination of N. punctiforme akinetes, cell division and heterocyst differentiation were slow. During the initial 15-20 h, N. punctiforme akinetes increased in volume and some burst. Only then did intact akinetes start to divide and fully germinate, possibly fueled by nutrients released from dead akinetes. The different strategies used by these different cyanobacteria allow successful germination of dormant cells and recolonization under favorable conditions.

The ABC Transporter Components HgdB and HgdC are Important for Glycolipid Layer Composition and Function of Heterocysts in Anabaena sp. PCC 7120

Anabaena sp. PCC 7120 is a filamentous cyanobacterium able to fix atmospheric nitrogen in semi-regularly spaced heterocysts. For correct heterocyst function, a special cell envelope consisting of a glycolipid layer and a polysaccharide layer is essential. We investigated the role of the genes hgdB and hgdC, encoding domains of a putative ABC transporter, in heterocyst maturation. We investigated the subcellular localization of the fusion protein HgdC-GFP and followed the differential expression of the hgdB and hgdC genes during heterocyst maturation. Using a single recombination approach, we created a mutant in hgdB gene and studied its phenotype by microscopy and analytical chromatography. Although heterocysts are formed in the mutant, the structure of the glycolipid layer is aberrant and also contains an atypical ratio of the two major glycolipids. As shown by a pull-down assay, HgdB interacts with the outer membrane protein TolC, which indicates a function as a type 1 secretion system. We show that the hgdB-hgdC genes are essential for the creation of micro-oxic conditions by influencing the correct composition of the glycolipid layer for heterocyst function. Our observations confirm the significance of the hgdB-hgdC gene cluster and shed light on a novel mode of regulation of heterocyst envelope formation.