The natural ecology of Saccharomyces yeasts

There are some organisms on Earth that have an especially close relationship with humankind, and one of them is Saccharomyces cerevisiae. This species, which is also known as the baker´s or brewer´s yeast, has been used for thousands of years, and almost all around the world, for the production of alcoholic beverages and bread. Today, we owe even more to this organism than wine, as it became one of the best studied model organisms in biology and is widely used in different scientific disciplines like Cell Biology, Biochemistry, Genetics and Molecular Biology. But surprisingly little is known about its natural ecology. The habitat of wild, undomesticated Saccharomyces yeasts cannot be reliably defined and contrary hypotheses exist. There is some evidence that the yeast is adapted to live in sugar rich environments like fruits, but on the other hand there is also indication that wild Saccharomyces yeasts are associated with oak trees, especially their bark. In both environments, yeast is only found in a small proportion of samples and most studies additionally suffer from different biases making it impossible to conclude which, if either, environment is truly the natural habitat of the yeast. In this thesis the natural ecology of Saccharomyces spp. was studied, with an emphasis on the natural environment. The oak bark environment was analyzed by characterizing the associated microbial community using culturing and pyrosequencing methods. S. paradoxus, the wild and undomesticated relative of S. cerevisiae, could indeed grow on nutrients present in oak bark but is only a rare member of the microbial community. Studying the influence of co-isolated oak bark microorganisms on S. paradoxus’ growth and survival in natural oak bark medium revealed a wide range of effects strongly dependent on temperature. Further experimental analysis of the interactions between the yeast and two of the bacteria gave insights into the diversity and complexity of natural microbial interactions. While one Pseudomonas spp. killed the yeast, another bacterium; Mucilaginibacter spp. promoted its growth. Saccharomyces’ metabolic specialty of fermenting under aerobic conditions (Crabtree effect) is taken as evidence that the yeast is adapted to sugary fruit environments. To test this, a Crabtree positive (“fermenter”) and its isogenic Crabtree negative mutant strain (“respirer”) were competed in laboratory media and grape juice, confirming that the Crabtree effect provides a benefit under resource competition in lab medium. Unexpectedly, this benefit was absent in the more natural grape juice, although adding natural microbial competitors restored the benefit of fermentation over respiration, perhaps by interference competition. Finally, the yeast / oak association could be confirmed by an intense sampling study, but Saccharomyces was found to be much more abundant in oak leaf litter than on bark. Oak leaf litter provides a stable habitat over the year from which yeast can be easily isolated and studied. This is a useful discovery for the ecology and evolutionary history of Saccharomyces yeasts, with great promise for future studies

A systematic forest survey showing an association of Saccharomyces paradoxus with oak leaf litter.

Although we understand the genetics of the laboratory model yeast Saccharomyces cerevisiae very well, we know little about the natural ecology and environment that shaped its genome. Most isolates of Saccharomyces paradoxus, the wild relative of S. cerevisiae, come from oak trees, but it is not known whether this is because oak is their primary habitat. We surveyed leaf litter in a forest in Northern Germany and found a strong correlation between isolation success of wild Saccharomyces and the proximity of the nearest oak. We compared the four most common tree genera and found Saccharomyces most frequently in oak litter. Interestingly, we show that Saccharomyces is much more abundant in oak leaf litter than on oak bark, suggesting that it grows in litter or soil rather than on the surfaces of oaks themselves. The distribution and abundance of Saccharomyces over the course of a year shows that oak leaf litter provides a stable habitat for the yeast, although there was significant tree-to-tree variation. Taken together, our results suggest that leaf litter rather than tree surfaces provide the better habitat for wild Saccharomyces, with oak being the preferred tree genus. 99.5% of all strains (633/636) isolated were S. paradoxus

Measuring microbial fitness in a field reciprocal transplant experiment

Microbial fitness is easy to measure in the laboratory, but difficult to measure in the field. Laboratory fitness assays make use of controlled conditions and genetically modified organisms, neither of which are available in the field. Among other applications, fitness assays can help researchers detect adaptation to different habitats or locations. We designed a competitive fitness assay to detect adaptation of Saccharomyces paradoxus isolates to the habitat they were isolated from (oak or larch leaf litter). The assay accurately measures relative fitness by tracking genotype frequency changes in the field using digital droplet PCR (DDPCR). We expected locally adapted S. paradoxus strains to increase in frequency over time when growing on the leaf litter type from which they were isolated. The DDPCR assay successfully detected fitness differences among S. paradoxus strains, but did not find a tendency for strains to be adapted to the habitat they were isolated from. Instead, we found that the natural alleles of the hexose transport gene we used to distinguish S. paradoxus strains had significant effects on fitness. The origin of a strain also affected its fitness: strains isolated from oak litter were generally fitter than strains from larch litter. Our results suggest that dispersal limitation and genetic drift shape S. paradoxus populations in the forest more than local selection does, although further research is needed to confirm this. Tracking genotype frequency changes using DDPCR is a practical and accurate microbial fitness assay for natural environments

The feedback between selection and demography shapes genomic diversity during coevolution

Species interactions and coevolution are integral to ecological communities, but we lack empirical information on when and how these interactions generate and purge genetic diversity. Using genomic time series data from host-virus experiments, we found that coevolution occurs through consecutive selective sweeps in both species, with temporal consistency across replicates. Sweeps were accompanied by phenotypic change (resistance or infectivity increases) and expansions in population size. In the host, population expansion enabled rapid generation of genetic diversity in accordance with neutral processes. Viral molecular evolution was, in contrast, confined to few genes, all putative targets of selection. This study demonstrates that molecular evolution during species interactions is shaped by both eco-evolutionary feedback dynamics and interspecific differences in how genetic diversity is generated and maintained

Recurrent Recruitment Manoeuvres Improve Lung Mechanics and Minimize Lung Injury during Mechanical Ventilation of Healthy Mice

INTRODUCTION: Mechanical ventilation (MV) of mice is increasingly required in experimental studies, but the conditions that allow stable ventilation of mice over several hours have not yet been fully defined. In addition, most previous studies documented vital parameters and lung mechanics only incompletely. The aim of the present study was to establish experimental conditions that keep these parameters within their physiological range over a period of 6 h. For this purpose, we also examined the effects of frequent short recruitment manoeuvres (RM) in healthy mice. METHODS: Mice were ventilated at low tidal volume V(T) = 8 mL/kg or high tidal volume V(T) = 16 mL/kg and a positive end-expiratory pressure (PEEP) of 2 or 6 cm H(2)O. RM were performed every 5 min, 60 min or not at all. Lung mechanics were followed by the forced oscillation technique. Blood pressure (BP), electrocardiogram (ECG), heart frequency (HF), oxygen saturation and body temperature were monitored. Blood gases, neutrophil-recruitment, microvascular permeability and pro-inflammatory cytokines in bronchoalveolar lavage (BAL) and blood serum as well as histopathology of the lung were examined. RESULTS: MV with repetitive RM every 5 min resulted in stable respiratory mechanics. Ventilation without RM worsened lung mechanics due to alveolar collapse, leading to impaired gas exchange. HF and BP were affected by anaesthesia, but not by ventilation. Microvascular permeability was highest in atelectatic lungs, whereas neutrophil-recruitment and structural changes were strongest in lungs ventilated with high tidal volume. The cytokines IL-6 and KC, but neither TNF nor IP-10, were elevated in the BAL and serum of all ventilated mice and were reduced by recurrent RM. Lung mechanics, oxygenation and pulmonary inflammation were improved by increased PEEP. CONCLUSIONS: Recurrent RM maintain lung mechanics in their physiological range during low tidal volume ventilation of healthy mice by preventing atelectasis and reduce the development of pulmonary inflammation

A clade uniting the green algae Mesostigma viride and Chlorokybus atmophyticus represents the deepest branch of the Streptophyta in chloroplast genome-based phylogenies

BACKGROUND: The Viridiplantae comprise two major phyla: the Streptophyta, containing the charophycean green algae and all land plants, and the Chlorophyta, containing the remaining green algae. Despite recent progress in unravelling phylogenetic relationships among major green plant lineages, problematic nodes still remain in the green tree of life. One of the major issues concerns the scaly biflagellate Mesostigma viride, which is either regarded as representing the earliest divergence of the Streptophyta or a separate lineage that diverged before the Chlorophyta and Streptophyta. Phylogenies based on chloroplast and mitochondrial genomes support the latter view. Because some green plant lineages are not represented in these phylogenies, sparse taxon sampling has been suspected to yield misleading topologies. Here, we describe the complete chloroplast DNA (cpDNA) sequence of the early-diverging charophycean alga Chlorokybus atmophyticus and present chloroplast genome-based phylogenies with an expanded taxon sampling. RESULTS: The 152,254 bp Chlorokybus cpDNA closely resembles its Mesostigma homologue at the gene content and gene order levels. Using various methods of phylogenetic inference, we analyzed amino acid and nucleotide data sets that were derived from 45 protein-coding genes common to the cpDNAs of 37 green algal/land plant taxa and eight non-green algae. Unexpectedly, all best trees recovered a robust clade uniting Chlorokybus and Mesostigma. In protein trees, this clade was sister to all streptophytes and chlorophytes and this placement received moderate support. In contrast, gene trees provided unequivocal support to the notion that the Mesostigma + Chlorokybus clade represents the earliest-diverging branch of the Streptophyta. Independent analyses of structural data (gene content and/or gene order) and of subsets of amino acid data progressively enriched in slow-evolving sites led us to conclude that the latter topology reflects the true organismal relationships. CONCLUSION: In disclosing a sister relationship between the Mesostigmatales and Chlorokybales, our study resolves the long-standing debate about the nature of the unicellular flagellated ancestors of land plants and alters significantly our concepts regarding the evolution of streptophyte algae. Moreover, in predicting a richer chloroplast gene repertoire than previously inferred for the common ancestor of all streptophytes, our study has contributed to a better understanding of chloroplast genome evolution in the Viridiplantae

Complex systems and the technology of variability analysis

Characteristic patterns of variation over time, namely rhythms, represent a defining feature of complex systems, one that is synonymous with life. Despite the intrinsic dynamic, interdependent and nonlinear relationships of their parts, complex biological systems exhibit robust systemic stability. Applied to critical care, it is the systemic properties of the host response to a physiological insult that manifest as health or illness and determine outcome in our patients. Variability analysis provides a novel technology with which to evaluate the overall properties of a complex system. This review highlights the means by which we scientifically measure variation, including analyses of overall variation (time domain analysis, frequency distribution, spectral power), frequency contribution (spectral analysis), scale invariant (fractal) behaviour (detrended fluctuation and power law analysis) and regularity (approximate and multiscale entropy). Each technique is presented with a definition, interpretation, clinical application, advantages, limitations and summary of its calculation. The ubiquitous association between altered variability and illness is highlighted, followed by an analysis of how variability analysis may significantly improve prognostication of severity of illness and guide therapeutic intervention in critically ill patients

Practical guidelines for rigor and reproducibility in preclinical and clinical studies on cardioprotection

The potential for ischemic preconditioning to reduce infarct size was first recognized more than 30 years ago. Despite extension of the concept to ischemic postconditioning and remote ischemic conditioning and literally thousands of experimental studies in various species and models which identified a multitude of signaling steps, so far there is only a single and very recent study, which has unequivocally translated cardioprotection to improved clinical outcome as the primary endpoint in patients. Many potential reasons for this disappointing lack of clinical translation of cardioprotection have been proposed, including lack of rigor and reproducibility in preclinical studies, and poor design and conduct of clinical trials. There is, however, universal agreement that robust preclinical data are a mandatory prerequisite to initiate a meaningful clinical trial. In this context, it is disconcerting that the CAESAR consortium (Consortium for preclinicAl assESsment of cARdioprotective therapies) in a highly standardized multi-center approach of preclinical studies identified only ischemic preconditioning, but not nitrite or sildenafil, when given as adjunct to reperfusion, to reduce infarct size. However, ischemic preconditioning—due to its very nature—can only be used in elective interventions, and not in acute myocardial infarction. Therefore, better strategies to identify robust and reproducible strategies of cardioprotection, which can subsequently be tested in clinical trials must be developed. We refer to the recent guidelines for experimental models of myocardial ischemia and infarction, and aim to provide now practical guidelines to ensure rigor and reproducibility in preclinical and clinical studies on cardioprotection. In line with the above guideline, we define rigor as standardized state-of-the-art design, conduct and reporting of a study, which is then a prerequisite for reproducibility, i.e. replication of results by another laboratory when performing exactly the same experiment

The interaction of Saccharomyces paradoxus with its natural competitors on oak bark

The natural history of the model yeast Saccharomyces cerevisiae is poorly understood and confounded by domestication. In nature, S. cerevisiae and its undomesticated relative S. paradoxus, are usually found on the bark of oak trees, a habitat very different from wine or other human fermentations. It is unclear whether the oak trees are really the primary habitat for wild yeast, or whether this apparent association is due to biased sampling. We use culturing and high-throughput environmental sequencing to show that S. paradoxus is a very rare member of the oak bark microbial community. We find that S. paradoxus can grow well on sterile medium made from oak bark, but that its growth is strongly suppressed when the other members of the community are present. We purified a set of twelve common fungal and bacterial species from the oak bark community, and tested how each affected the growth of S. paradoxus in direct competition on oak bark medium at summer and winter temperatures, identifying both positive and negative interactions. One Pseudomonas species produces a diffusible toxin that suppresses S. paradoxus almost as effectively as either the whole set of twelve species together, or the complete community present in non-sterilized oak medium. Conversely, one of the twelve species, Mucilaginibacter sp., had the opposite effect and promoted S. paradoxus growth at low temperatures. We conclude that, in its natural oak tree habitat, S. paradoxus is a rare species whose success depends on the much more abundant microbial species surrounding it