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Abstract book 1 We acknowledge the following Institutions/Companies for their support: Fundação para a Ciência e Tecnologia – FCT Reitoria da Universidade do Porto Faculdade de Ciências da Universidade do Porto Instituto de Ciências Biomédicas Abel Salazar da Universidade do Porto Centro Interdisciplinar de Investigação Marinha e Ambiental – CIIMAR Sociedade Portuguesa de Microscopia Câmara Municipal do Porto Enzifarma - - Diagnóstica e Farmacêutica, S.A. VWR International- Material de Laboratório, Lda. Nzytech – genes and enzymes Sogrape Vinhos 2 Organizing Committee: Olga Maria Lage, FCUP, Univ Porto Alexandre Lobo da Cunha, ICBAS, Univ Porto Damien P. Devos, CABD, UPO, Sevilla Program Day 1 – Wednesday, 10 th May 9:00 – 9:45 Registration 9:45 – 10:00 Opening Session 10:00 - 10:30 Warming up talk - Damien P. Devos - The Paradigms They Are a-Changin’: Past, present and future of your favorite bacteria 10:30 – 11:00 Coffee break 11:00 – 12:00 Keynote speaker - Christian Jogler - Planctomycetes after the paradigm shift: more exciting than ever! 12:00 - 12:30 Lise Øvreås - Disentangling the extant of Planctomycetes diversity from numerous habitats 12:30 – 14:00 Lunch 14:00 – 15:00 Keynote speaker – Matthias Horn - Chlamydiae in the environment - changing our perspective on the C in the PVC Superphylum 15:00 – 15:30 Astrid Collingro - Marine chlamydiae are motile - evidence from single cell genomes 15:30 – 16:00 Sandra Wiegand - Diversity driven cultivation and genome sequencing of 80 slow growing Planctomycetes reveal their potential for small molecule production 16:00 – 16:30 Coffee break 16:30- 17:00 Rita Calisto - Anticancer activity in Planctomycetes 17:00 – 17:30 Olga M. Lage - Planctomycetes: links to the environment 18:00 Welcome reception 3 Day 2 - Thursday, 11th May 9:30 – 10:30 Gemmata Keynote speaker – Siv Andersson - Comparative genomics of 10:30 – 11:00 Coffee break 11:00 – 11:20 Timo Kohn - So close, yet so far – Closest related Planctomycetes can differ substantially in their genome organizations 11:20 – 11:50 Carlos Santana, Elena Rivas and Sean Stettner - Hopanoid versus sterol synthesis in PVCs 11:50- 12:10 Julia Endresen Storesund - Planctomycetes distribution along chemical gradient in the meromictic Lake Sælenvannet 12:10 – 12:30 Milton S. da Costa - The Accumulation of Compatible solutes in Plantomycetes 12:30 – 14:00 Lunch 14:00 – 15:00 Keynote speaker – Clara Belzer - Verrucomicrobia and the Intestinal Microbiome: The case of Akkermansia muciniphila 15:00 – 15:30 Laura van Niftrik - The energy-conserving prokaryotic organelle of anammox Planctomycetes 15:30 - 16:00 Michael Y. Galperin - Bacterial Membrane Energetics and Signaling from the First Cells to Chlamydia and Kuenenia 16:00 – 16:30 Coffee break 16:30 – 18:00 Enzifarma FCM workshop – Alexandre Salvador – Flow Cytometry applied to bacterial studies 19:30 20:30 Porto City tour Dinner by the river Day 3 - Friday, 12th May 9:30 – 10:30 Keynote speaker – Svetlana Dedysh - Planctomycetes in wetlands: diversity and ‘omics’-based insights into ecological functions 10:30 – 11:00 Coffee break 11:00 – 11:30 Anastasia A. Ivanova - Metatranscriptome-based insight into hydrolytic potential of peat-inhabiting Planctomycetes 11:30 – 12:00 Irina S. Kulichevskaya - Abundance and diversity of Planctomycetes in lichen-dominated sub-Arctic ecosystems of northwestern 12:00 – 12:30 Ice Cave, Northern Norway Eirik Færøy Sæbø - Novel Planctomycetes from Svarthamar 12:30 – 12:45 Closing session 12:45 – 14:00 Lunch 4 Keynote lectures 5 Comparative Genomics of Gemmata Species Siv G.E. Andersson, Mayank Mahajan, Benjamin Yee and John Fuerst Department of Molecular Evolution, Biomedical Center, Science for Life Laboratory, Uppsala University, Uppsala, Sweden Many of the pioneering studies on the Planctomycetes have used Gemmata obscuriglobus as the experimental model organism. These studies have revealed many unique cellular traits, such as a highly complex intracellular network of membranes, cell division by budding, spatial separation of transcription from translation and the ability to take up proteins and complex sugars from the environment. Despite the importance of G. obscuriglobus as a model organism for studies of compartmentalized cell plans, no closed genome is available for this species. The 9 Mb draft genome which has been used for inferences of the proteome contains more than 900 gaps. The unfinished status of the genome has been problematic for the interpretations of the results obtained thus far for many reasons. We will present the closed genome of G. obscuriglobus along with the genomes of two closely related strains as well as the genome of a recently described species of a related, but novel genus. First, a phylogeny of their internal relationships will be presented, onto which we have mapped the flux of protein families. The comparative analyses indicate a gradual expansion in gene content by duplication and divergence at most of the ancestral nodes within this bacterial clade. We also noted remarkable differences in the occurrence of mobile elements among these genomes, with a massive proliferation of transposons in G. obscuriglobus. Our analyses of the evolution of potentially new gene functions through expansions, losses and rearrangements of protein domains in the proteomes of these species will be discussed, along with our interpretations of why such dramatic changes have occurred. 6 Verrucomicrobia and the Intestinal Microbiome: The case of Akkermansia muciniphila Clara Belzer Laboratory of Microbiology, Wageningen University, The Netherlands The intestinal microbiome is considered as an important modulator of host health. The Verrucomicrobia is one of the most abundant microbial phyla in the human microbiota, making up on average 1-4% in the colon. To date, Akkermansia spp. is the only representatives of the Verrucomicrobia colonizing the intestine. Akkermansia spp. have adapted to the intestinal environment by specializing in the degradation of host mucin, which it may use as a sole source of carbon and nitrogen. The mucinases encoded in the genome of Akkermansia spp. are mostly obtained via horizontal gene transfer from Bifidobacterium spp. and Bacteroides spp.. In vitro work with Akkermansia muciniphila indicated that the organism is auxotrophic for growth on an amino acid highly abundant in the peptide core of the mucus protein. On top of this A. muciniphila is adapted to its host through the ability to use oxygen, diffusing from host epithelial cells, for higher growth yields. A. muciniphila has a direct effect on the microbial community structure at the mucosal layer. Mucus degradation and fermentation by A. muciniphila result in the liberation of oligosaccharides and subsequent production of acetate, which becomes directly available to microorganisms in the vicinity of the intestinal mucosa. Co-culturing experiments of A. muciniphila with the non-mucus degrading butyrate-producing microbes; Anaerostipes caccae, Eubacterium hallii or Faecalibacterium prausnitzii resulted in syntrophic growth and production of butyrate. The metabolic interactions of A. muciniphila and butyrigens provide evidence for a colonic butyrate production pathway that is dependent on host produced glycans and independent of dietary carbohydrates. Apart from microbial ecology A. muciniphila is described to directly effect host response. A. muciniphila possesses anti-inflammatory properties, and has been correlated to protection against inflammation in diseases such as, type 2 diabetes mellitus, and obesity. In fact, A. muciniphila treatment can reverse fat gain, serum lipopolysaccharide (LPS) levels, gut barrier function, and insulin resistance in mice. Immune stimulatory capacities of A. muciniphila lay partially within its extraordinary outer-membrane structure. Single purified outer-membrane proteins of A. muciniphila stimulate host immune response through TLR2 (mainly IL6, Il8, Il10, INFg) and increase trans-epithelial resistance of host cells. On top of this our results have indicated that pasteurized cells and the outer membrane protein Amuc_1100 can be used as an oral treatment against fat mas gain and insulin resistance in mice. In conclusion Akkermansia spp. are a extraordinary group within the Verrucomicrobia. They are the only Verrucomicrobia adapted to the host intestinal environment. Within the mucosa Akkermansia spp. directly stimulates host response and a beneficial microbial community. In light of these findings, future work will gain a better mechanistic understanding and hopefully the actual role of A. muciniphila in health and disease. 7 Planctomycetes in wetlands: diversity and ‘omics’-based insights into ecological functions Svetlana N. Dedysh Winogradsky Institute of Microbiology, Research Center of Biotechnology RAS, Moscow, Russia Wetlands are ecosystems in which the water table is permanently or periodically close to the soil surface. These water-saturated terrestrial environments, such as boreal Sphagnum-dominated peat bogs and lichen-dominated tundra peatlands, are abundantly colonized by members of the phylum Planctomycetes, which inhabit both oxic and anoxic peat layers. Examination of peat material by fluorescence in situ hybridization commonly reveals numerous (up to 107 cells per gram of wet peat) cells of planctomycetes, which are arranged in chains or shapeless cell aggregates and are mostly attached to the particles of non-decomposed organic material. 16S rRNA gene fragments from the Planctomycetes comprise 5-13% of total 16S rRNA gene reads retrieved using high-throughput sequencing techniques from peat samples. Most 16S rRNA gene reads representing this phylum in Sphagnum- derived peat commonly affiliate with the family Isosphaeraceae. By contrast, assemblages of planctomycetes in lichen-dominated peatlands of tundra have a high proportion of representatives affiliated with the Phycisphaera-related group WD2101 (order Tepidisphaerales). Members of the family Gemmataceae are also common in boreal and tundra wetlands. Although several peat-inhabiting planctomycetes have been obtained in pure cultures, little is known about their potential functions in the environment. In order to get insights into the metabolic potential of planctomycetes, we have assessed their activity response to amendment of native peat with cellulose, xylan, pectin and chitin, which are the major biopolymers in Sphagnum-derived peat. The analysis of RNA pool via barcoded Illumina sequencing revealed that some groups within uncultivated planctomycetes positively responded to the amendment of peat with various polysaccharides, suggesting the presence of hydrolytic capabilities in these bacteria. In particular, the strongest substrate-induced response was detected on chitin for Gemmata- and Phycisphaera-like planctomycetes. Given that routine laboratory tests used for assessing hydrolytic capabilities are complicated by slow growth rates of peat-inhabiting planctomycetes, the comparative genomic approach was applied to unveil the hidden potential of these bacteria. The genomes of two recently described planctomycetes, Paludisphaera borealis PX4T (family Isosphaeraceae) and Fimbriiglobus ruber SP5T (family Gemmataceae), were sequenced and analyzed. The genomes encode wide repertoires of carbohydrate-active enzymes (CAZymes) and large numbers of unclassified putative glycoside hydrolases, suggesting that peat-inhabiting planctomycetes possess extremely high but partly hidden glycolytic potential and have the ability to utilize a wide range of natural carbohydrates and glycoconjugates. Notably, the CAZyme repertoire in planctomycetes significantly differs from those in well-studied hydrolytic bacteria. We were also able to demonstrate the presence of chitinolytic capability in Fimbriiglobus ruber SP5T, which was suggested by the results of metatranscriptomic study and the genome analysis. This is the first member of the order Planctomycetales with confirmed chitinolytic capability. In summary, the results of our studies suggest participation of peat-inhabiting planctomycetes in degradation of plant-derived polymers, exoskeletons of peat-inhabiting arthropods as well as exopolysaccharides produced by other bacteria. 8 Chlamydiae in the environment - changing our perspective on the C in the PVC Superphylum Matthias Horn Division of Microbial Ecology, Department of Microbiology and Ecosystem Science, University of Vienna, Austria The Chlamydiae, originally considered a small group of closely related bacteria infecting humans and animals, are much more diverse than recognized previously. A picture is beginning to emerge, in which the Chlamydiae represent the probably most ancient lineage of intracellular bacteria; they have adapted to the infection of eukaryotic host cells very early during evolution, likely during interplay with ancient protists. Extant Chlamydiae comprise both pathogens of humans and ubiquitous symbionts of amoebae. The discovery of chlamydiae in the environment not only fundamentally changed our perception of chlamydial diversity; their analysis also provided new perspectives on evolution and biology of this important group of intracellular microbes. In this talk I will summarize recent findings about their unique developmental cycle, their cell biology, and molecular evolution. 9 Planctomycetes after the paradigm shift: more exciting than ever! Christian Jogler Faculty of Science, Radboud University Nijmegen, The Netherlands Since their discovery in 1924, Planctomycetes seemed to bluer the pro- / eukaryotic dichotomy. Their unusual FtsZ-less cell division -mostly through polar budding- their lack of peptidoglycan and their complex subcellular membrane systems clearly set them apart from all other bacteria. Ultimately, even a nucleus-like structure with nuclear pores and separation of transcription and translation was suggested. With the postulation of endocytosis-like uptake of macromolecules the major eukaryotic hallmark trait required for gaining an endosymbiont was described for Planctomycetes. Even membrane-coat-like proteins -as in eukaryotes- seemed to be required for vesicle formation to facilitate endocytosis. With good reason, Planctomycetes were envisioned as potential ancestors of both, bacteria and eukaryotes and their traits appeared indeed beyond the bacterium. However, with the advent of genetic tools, advanced genome research methods and high resolution imaging systems for light- (dSTORM and SRSIM) and electron microscopy (FIB-SEM and CET), multiple laboratories provided strong arguments for the Gram-negative cell architecture of Planctomycetes. Employing such methods, my group recently demonstrated that Planctomycetes in general lack additional membrane surrounded compartments, but that their cytoplasmic membrane can create invaginations to store complex carbon substrates in the enlarged periplasmic space. We further found that at least the suspected MC protein is not required for macro-molecule uptake and that a process different from endocytosis is employed to incorporate such molecules. Thus, we reject the hypothesis of the planctomycetal ancestral relationship with eukaryotes and suggest seeing Planctomycetes as maverick Gramnegative bacteria instead. However, in the past the clear majority of high impact publications on Planctomycetes focused on such eukaryote-like traits and their potential involvement in eukaryogenesis. Thus, the findings of others and us will cause a major paradigm shift in the field of planctomycetal research. One might even ask if Planctomycetes are important to study after all? In my talk, I will present further evidence supporting this paradigm shift. I will focus on what is left of all these planctomycetal curiosities and which novel fields of future research such as the production of antibiotics or the planctomycetal role in global carbon cycles just emerged. In the light of these recent findings -without any doubt- Planctomycetes are even more exciting than ever! 10 Oral presentations 11 Anticancer activity in Planctomycetes Rita Calisto1, Eirik Færøy Sæbø2, Lise Øvreås2, Olga M. Lage1,3 and Lars Herfindal4 1 Department of Biology, Faculty of Sciences, University of Porto, Portugal 2 Department of Biology, University of Bergen, Norway 3 Interdisciplinary Centre of Marine and Environmental Research, University of Porto, Portugal 4 Centre for Pharmacy, Department of Clinical Science, University of Bergen, Norway Planctomycetes has recently been recognized as important candidates for possessing antibiotic and antifungal activities. They are together with Actinobacteria and Myxobacteria, known for their bioactive potential and specific characteristics such as large genomes and complex life cycles. These characteristics make Planctomycetes good candidates for production of bioactive molecules. Previous in silico genome mining analyses from several Planctomycetes revealed the presence of genes related to various pathways for the production of several bioactive molecules, including some antitumor compounds like epothilone. Besides this potential, no study has up-to-now addressed the anti-cancer proprieties of Planctomycetes. In fact, this study is the first to approach the planctomycetes’ capacity to induce apoptosis and decrease cell growth of cancer cell lines. In this study, twenty-six Planctomycetes strains were screened for their ability to induce apoptosis and diminish cell growth on an acute myeloid leukaemia cell line and prostatic cancer cell line. This was achieved by focusing on the solubility of the compounds to see if they were present on the organic or aqueous phase of the cell extracts. Acute myeloid leukaemia (AML) and Prostate cancer cell lines were chosen because AML is the one of the most aggressive forms of leukaemia and, frequently, presents chemoresistance to the treatments, and prostate cancer is one of the most common cancers and with higher mortality rate, especially in western countries. Normal rat kidney epithelial cell line (NRK) was used as control. Assessment of cytotoxic activity was performed by two different methods: first by the metabolic capacity of the cell culture, using the cell proliferation reagent WST-1, and thereafter by analysing the percentage of cell presenting apoptotic nuclei, through microscopy. Our findings indicate that Planctomycetes are producers of anticancer compounds and of toxins for human cells, with seventeen extracts/strains affecting at least one of the cancer cell lines, with intermediate or high toxicity. The AML (Molm-13) cells were in general more sensitive towards the extracts compared to the prostate cancer cells. Moreover, we identified that the anti-cancer activity induced a delayed cell death, not present in any cell culture after 24 hours. However, after 72 hours, several extracts had induced cell death in all cell lines. This screening also demonstrates that the production of compounds is probably related with the phylogeny of the bacteria, as planctomycetes from the same genera affects the same type of cells. Strains affiliating to Roseimaritima ulvae, Rhodopirellula lusitana and Rhodopirellula sp. only affected the AML cell line, while Rhodopirellula baltica affected the AML cells, the prostatic cancer cells and the control normal cells. Contrarily, Rubinisphaera brasiliensis did not seem to affect any type of cells. More consistent were the results regarding the solubility of the produced compounds as all the positive results were obtain from organic extracts. Finally, the extracts that provoked a higher cell death (over 70% of dead cells) still affected the cells even after being diluted 4 times. This study provides the first insight on the effects of the compounds produced by Planctomycetes on human cells, especially cancer cell lines, demonstrating their amazing potential for biotechnological application. 12 Marine chlamydiae are motile - evidence from single cell genomes Astrid Collingro1, Stephan Köstlbacher1, Mark Mußmann1, Ramunas Stepanauskas2, Steven J. Hallam3,4, Matthias Horn1 1 Department of Microbial and Ecosystems Science, Division of Microbial Ecology, University of Vienna, 1090 Vienna, Austria; 2 Bigelow Laboratory for Ocean Sciences, East Boothbay, ME 04544,USA; 3 Department of Microbiology and Immunology, University of British Columbia, Vancouver, BC, Canada; 4 Genome Science and Technology Program, University of British Columbia, Vancouver, BCV6T 1Z3, Canada The past two decades revealed an enormous diversity within the chlamydiae. Molecular data provide evidence for the existence of at least 180 chlamydial families beyond the human and animal pathogenic Chlamydiaceae. Many of these divergent chlamydiae are of marine origin and besides limited molecular data no information about their lifestyle and biology is available so far. In order to gain insights into the biology of these yet unexplored chlamydiae, and to understand their genomic diversity and similarity to known chlamydiae, we recovered and analyzed three chlamydial single-cell amplified genome (SAG) sequences from marine environments. Single microbial cells from water samples from the Saanich Inlet (Canada) and the Northern Sea (Germany), respectively, were sorted by high-speed fluorescence activated cell sorting (FACS). Genomic DNA of these cells was amplified by multiple displacement amplification (MDA), and chlamydial SAGs were identified by 16S rRNA gene sequencing. Three chlamydial SAGs were subsequently sequenced on MiSeq and NextSeq (Illumina) instruments. Sequence reads were quality checked, assembled with SPAdes, and annotated with ConsPred. Phylogenetic analyses were performed with MrBayes or RAxML. Approximately 41-50% of the genome of each SAG could be retrieved, indicating whole genome sizes of 2.2-2.6 Mbp for these organisms. According to phylogenies of 16S rRNA and other single copy marker genes, the SAGs represent deeply branching marine chlamydiae. Their predicted metabolic capabilities are consistent with those reported for other chlamydiae, including reduced amino acid, vitamin, and cofactor biosynthetic pathways. The presence of type III secretion genes, ATP/ADPtranslocases, CPAF, Euo and other chlamydia-specific genes are pointing towards an intracellular lifestyle with the typical biphasic developmental cycle in these chlamydiae. Surprisingly, all three SAGs harbor genes for chemotaxis and assembly of flagella. Phylogenetic analysis of flagellar genes suggests a common chlamydial origin of this trait subsequently lost by all other known chlamydiae. Analysis of the first SAGs of divergent and so far uncultured marine chlamydiae suggests an intracellular lifestyle by employing characteristic mechanisms for host interaction known from other chlamydiae. In addition these chlamydiae have the genetic repertoire for chemotaxis and motility, which they might use to trace or attach to their hosts. Together this indicates that albeit the conservation of the genes involved in the basic chlamydial lifestyle, genomes of divergent lineages bear surprises and are more flexible than previously thought. 13 The Accumulation of Compatible solutes in Plantomycetes Milton S. da Costa, Centro de Neurociências e Biologia Celular, Universidade de Coimbra, 3001-401 Coimbra, Portugal. All microorganisms must adjust, within intrinsic limits, to alterations in the water activity of the environment. The majority of microorganisms adjust osmotically by the selective accumulation of small molecular weight organic compounds, such as sugars and sugar derivatives, polyols, amino acids and amino acid derivatives. Some compatible solutes, namely trehalose, glutamate and glycine betaine are common compatible solutes of both bacteria and archaea. Other compatible solutes such as glucosylglycerate and mannosylglyceramide are very rare and some are restricted to one known organism. The bacteria of the phylum Planctomycetaeota (Planctomycetes) represent a unique group of organisms with unique morphologies. Many of these organisms are of marine origin and should thus accumulate compatible solutes to cope with increases in the salinity of the environment. We have examined the compatible solute pools of three planctomycetes to grasp the diversity of compatible solutes that accumulate in the organisms under salt stress and the biosynthetic pathways involved in their synthesis. The species Rhodopirellula baltica accumulates sucrose, glutamate, trehalose and the extremely rare mannosylglucosylglyerate (MGG) found previously in only two species of thermophilic Petrotoga of the order Thermotogales). The synthetic pathway of the latter compatible solute was also determined. The relationship between nitrogen availability and osmotic adjustment showed that MGG accumulated during growth under low nitrogen levels in the medium. To extend these results we also examined the biosynthesis and the accumulation of compatible solutes in two other planctomyces, namely Rubinisphaera brasiliensis and Gimesia maris. The former accumulated primarily α-glutamate, sucrose, ectoine and hydroxyectoine. But G. maris also accumulated glucosylglycerate under low nitrogen levels in the medium. The synthesis and regulation of the accumulation of glucosylglycerate will be discussed. 14 The Paradigms They Are a-Changin’: Past, present and future of your favorite bacteria. Damien P. Devos Centro Andaluz de Biología del Desarrollo CABD Universidad Pablo de Olavide-CSIC, Sevilla, Spain These are wonderful times for PVC research! The field is booming, erroneous interpretations have been corrected and we are now facing a bright future. Let’s hope the funding agencies are listening! In this talk, I will revisit the past and current efforts in the analyses of the bacterial Planctomycetes-Verrucomicrobia-Chalmydiae (PVC) superphylum. I will present an historical perspective on this disparate group of bacteria. I will go back to the early days of PVC research and follow the evolution of the analyses performed and interpretations presented. I will then explore the increasing number and diversity of PVC researches observed today. I will summarize the recent novelties described in the field and how this modified our perception and interpretations of these bacteria. Eventually, I will close by presenting a few directions that the PVC field and its happy community might take in the future. 15 Bacterial Membrane Energetics and Signaling from the First Cells to Chlamydia and Kuenenia Michael Y. Galperin NCBI, NLM, National Institutes of Health, Bethesda, Maryland, USA The Planctomycetes-Verrucomicrobia-Chlamydiae superphylum is an early-branching bacterial lineage that could provide valuable clues on the evolution of bacteria. Recent studies of prokaryotic genome sequences brought about a significant progress in understanding of the general physiology of bacteria and their metabolic pathways. Components of the membrane energetics machinery and signal transduction pathways attracted much less attention, owing largely to the multi-subunit nature of the membrane energy-transducing and signaling enzymes and their diversity. We have used the Clusters of Orthologous Groups of proteins (COG) system for a comparative analysis of the PVC bacteria and their ability to utilize protonmotive force and/or sodium-motive force. This study placed the membrane bioenergetics of PVC members squarely in the bacterial domain but revealed presence of distinct mechanisms of energy conservation in different lineages, reflecting their likely diversification from the ancestral sodium-dependent bioenergetics [1-3]. Free-living PVC members often encode both H+- and Na+-translocating ion pumps and H+- and Na+-translocating ATP synthases. In contrast, chlamydia show clear signs of loss of membrane enzymes in the course of the adaptation to the parasitic lifestyle [4,5]. The signaling mechanisms of PVC members also show dramatic diversity with planctomycetes and verrucomicrobia having some of the most complex signal transduction machineries among all bacteria, which include two-component signaling, chemotaxis, Ser/Thr-protein kinases and protein phosphatases, as well as c-di-GMP- and c-diAMP-mediated signal transduction [6,7]. Chlamydia encode greatly streamlined versions of these systems, which, however, still include two-component, Ser/Thr protein phosphorylation, and c-di-AMP-mediated signaling. The relatively narrow bioenergetics and signaling capabilities of chlamydial cells open possibilities for designing potential lineage-specific antibiotics. References 1. Mulkidjanian AY et al. (2008) Biol. Direct 3: 13. PMID: 18380897 2: Mulkidjanian AY et al. (2008) Biochim. Biophys. Acta 1777: 985-992. PMID: 18485887 3. Mulkidjanian AY et al. (2012) Proc. Natl. Acad. Sci. USA 109: E821-E830. PMID: 20693418 4. Häse CC et al. (2001) Microbiol. Mol. Biol. Rev. 65: 353-370. PMID: 11528000. 5. Dibrov P et al. (2004) J. Mol. Microbiol. Biotechnol. 8: 1-6. PMID: 15741735. 6. Galperin MY (2005) BMC Microbiol. 5:35. PMID: 15955239. 7. Galperin MY et al. (2010) Mol. BioSystems 6: 721-728. PMID: 20237650. 16 Metatranscriptome-based insight into hydrolytic potential of peatinhabiting Planctomycetes Anastasia A. Ivanova1, Carl-Eric Wegner3, Werner Liesack2 and Svetlana N. Dedysh1 1 Winogradsky Institute of Microbiology, Research Center of Biotechnology RAS, Moscow, Russia; 2 Max Planck Institute for Terrestrial Microbiology, Marburg, Germany; 3 Friedrich Schiller University Jena, Institute of Ecology, Aquatic Geomicrobiology, Jena, Germany Members of the phylum Planctomycetes are common inhabitants of northern Sphagnumdominated wetlands. The evidence is accumulating that some members of this phylum can be involved in degradation of organic matter in these ecosystems but the experimental data remain scarce due to the low number of characterized representatives from this bacterial group. Here, we aimed to get insights into the metabolic potential of planctomycetes by monitoring their activity response to amendment of native peat with cellulose, xylan, pectin and chitin, which are the major biopolymers in Sphagnum-derived peat. Peat sampled from a Sphagnum peat bog in northern Russia was amended with 500 mg/L of the polysaccharides mentioned above and incubated for 6 weeks. Insights into community changes caused by substrate availability were gained by the analysis of rRNA and mRNA pools via barcoded Illumina sequencing. A total of 1,638,305 rRNA and mRNA sequences affiliated with the Planctomycetes were retrieved in our study. The majority (~60%) of 16S rRNA sequences could not be assigned to taxonomically characterized organisms. Community shifts within the Planctomycetes were rather distinct depending on the added carbon source. The strongest substrate-induced response was detected on chitin. The two groups with increased transcript pools were Gemmata- and Phycisphaera-like planctomycetes. Among uncultivated members of the Planctomycetaceae, two abundant transcript pools were retrieved from pectin-amended samples and belonged to Pirellula-like bacteria. Taken together, metatranscriptome analysis revealed various planctomycetes-like populations to respond positively to the amendment with polysaccharides, providing evidence for hydrolytic capabilities in these bacteria. 17 So close, yet so far – Closest related Planctomycetes can differ substantially in their genome organizations Timo Kohn1, Sandra. Wiegand1, Patrick Rast2, Manfred Rohde3, Franz Brümmer4, RalfWalter Müller4, Mareike Jogler2 and Christian Jogler1 1 Radboud University, Nijmegen, Netherlands 2 DSMZ, Braunschweig, Germany 3 HZI, Braunschweig, Germany 4 University of Stuttgart, IBBS, Stuttgart, Germany Since decades, the 16S rRNA gene is the gold standard for the classification of bacteria. But its resolution, when it comes to close related strains, is restricted. This limitation can be overcome with the advances in genome sequencing and the comparison of multiple marker genes or whole genomes as such. Here we show a novel Planctomycete, isolated from a freshwater sponge, that shows 99% 16S sequence identity with the type strain Planctopirus limnophila DSM3776T, but differs considerably when the whole genomes are compared. Based on 16S rRNA sequence identity, this isolate would represent a novel strain, while Average Nucleotide Identity (ANI) proves that this isolate represents a novel species. Compared to the type strain, this strain has two different genes encoding for the 16S rRNA and shows differences in the predicted secondary metabolite gene clusters. Therefore, evaluating the “novelty” of a strain based on 16S rRNA identity, may leave novel talented producers undetected. 18 Abundance and diversity of Planctomycetes in lichen-dominated subArctic ecosystems of northwestern Siberia Irina S. Kulichevskaya, Anastasia I. Ivanova and Svetlana N. Dedysh Winogradsky Institute of Microbiology, Research Center of Biotechnology of the Russian Academy of Sciences, Moscow, Russia Members of the bacterial phylum Planctomycetes inhabit a wide range of aquatic and terrestrial environments with diverse environmental conditions. Yet, most cultured and taxonomically characterized representatives of this phylum are mesophiles. At the same time, planctomycetes are commonly detected in various low-temperature ecosystems by molecular surveys. This study was initiated in order to assess the abundance and diversity of planctomycetes in wetlands and upland soils within the zone of forested tundra and discontinuous permafrost in northwest Siberia with a ground vegetation cover composed of reindeer lichens (genera Cladonia and Cetraria). The microbial communities of two lichen-dominated ecosystems typical of the sub-arctic zone of northwestern Siberia, that is a forested tundra soil and a shallow acidic peatland, were examined in our study. As revealed by molecular analyses, soil and peat layers just beneath the lichen cover were abundantly colonized by bacteria from the phylum Planctomycetes. Highest abundance of planctomycetes detected by fluorescence in situ hybridization was in the range 2.2–2.7 х 107 cells per gram of wet weight. 16S rRNA gene fragments from the Planctomycetes comprised 8–13% of total 16S rRNA gene reads retrieved using Illumina pair-end sequencing from the soil and peat samples. Lichen-associated assemblages of planctomycetes displayed unexpectedly high diversity, with a total of 89,662 reads representing 1723 operational taxonomic units determined at 97% sequence identity. The soil of forested tundra was dominated by uncultivated members of the family Planctomycetaceae (53–71% of total Planctomycetes-like reads), while sequences affiliated with the Phycisphaera-related group WD2101 (recently assigned to the order Tepidisphaerales) were most abundant in peat (28– 51% of total reads). Representatives of the Isosphaera–Singulisphaera group (14–28% of total reads) and the lineages defined by the genera Gemmata (1–4%) and Planctopirus– Rubinisphaera (1–3%) were present in both habitats. Two isolates of Singulisphaera-like bacteria, strains P12 and P515, were isolated from a peatland and a forested tundra soil, respectively. The isolates shared identical 16S rRNA gene sequences, which exhibit only 93–94% sequence similarity to 16S rRNA gene sequences from members of the genus Singulisphaera and 91–92% sequence similarity to members of the genus Paludisphaera. Strains P12 and P515 displayed good tolerance of low temperatures (4– 15oC) and were capable of growth on a number of polysaccharides, including lichenan, a characteristic component of lichen-derived phytomass. Based on the characteristics determined in our study, we propose to classify the novel planctomycetes as representing a novel genus and species, ‘Tundrisphaera lichenicola’ gen. nov., sp. nov. Thus, our study provided the first proof of high planctomycete abundance and diversity in cold, sub-arctic tundra ecosystems. 19 Planctomycetes: links to the environment Olga M. Lage1,2, Maria da Conceição Marinho1, Carlos Ramos1, Teresa Carvalho1 and Sara C. Antunes1,2 1 2 Department of Biology, Faculty of Sciences, University of Porto, Portugal Interdisciplinary Centre of Marine and Environmental Research, University of Porto, Portugal Planctomycetes have been found in a broad range of habitats but their precise role in such environments is not well known. Their physiological capacities make them suitable for sulfated hydrocarbon degradation playing an important role in mineralization or for the removal of ammonium and nitrite in the anammox process. The constant accumulation of toxic compounds such as pharmaceutical drugs can affect the base of the trophic chain, reducing an important food source to organisms in higher trophic levels. In this work, we explore different ecological aspects of planctomycetes namely (1) their adequacy to serve as food source for other organisms in higher trophic levels; (2) their capacity for hydrocarbon degradation; (3) the impact of pharmaceuticals in their viability. Our studies address thematics still scarcely explored. 20 Disentangling the extant of Planctomycetes diversity from numerous habitats Julia Storesund, Eirik Færøy Sæbø and Lise Øvreås Department of Biology, University of Bergen, Norway The phylogenetic positions of 30 isolates that morphologically resemble members of the family Planctomycetaea were determined by sequencing analyses of the 16S rRNA gene. The isolates were distributed amongst seven genera, where two represents completely novel genera. Seven isolates were assigned to the genera Rhodopirellula, 5 to genera Blastopirellula, 7 to the genera Gimesia, 7 to the genera Rubinisphaeara whereas the five last isolates were found to be only distantly related to previously described Planctomycetes. The collection of these unique Planctomycetes strains were obtained from different extreme environments, including deep-sea iron hydroxide deposits (Atlantic Mid Ocean spreading Ridge and the South Pacific spreading Ridge), 700 years old glacier cave ice, biofilm on kelp from polar waters around Svalbard, and water masses from a meromictic lake displaying a strong chemical gradient. Morphological characters and internal membrane structures of the isolates generally correlates with features described for the order Planctomycetales, but distinct differences were seen between the isolates. We anticipate that the Planctomycetes are reservoir for innovative knowledge both regarding the evolution of life and cellular compartmentalisation processes, which are translated into specialized and novel microbial communities that colonize various natural environments. The putative role of planctomycetal carbon remineralization is key for the ecology of many different environments, especially marine sediments and ice biomes. Given the slow growth of planctomycetes, some species have doubling time up to several weeks. The high abundance of planctomycetes in certain deep ocean environments might be driven by the presence of complex carbon sources in contrasting to oligotrophic surrounding water masses where they are rather scarcely present. Our collection also contains isolates from the biofilm of kelps where they have to face strong competition and are obliged to develop scavenging survival strategies like the production of antimicrobials. Functional analyses of closely related bacteria obtained from widely different environments, could improve our understanding of such mechanisms, and several of our bacterial strains obtained from different environments are documented as closely related. 21 Novel Planctomycetes from Svarthamar Ice Cave, Northern Norway Eirik Færøy Sæbø, Julia Endresen Storesund, Hilde Rief Armo, Stein Erik Lauritzen, Lise Øvreås Department of Biology, University of Bergen, Norway Ice caves are exceedingly conserved and quite unexposed extreme environments, representing a unique location for studying the impact of darkness, low temperatures and evolution in time on the organisms living there. The Svarthamar Ice cave in Norway is a two entrance, dynamic ice cave, possessing the largest Cave room in Scandinavia and is one of the lowest altitude ice caves in Norway. Radiocarbon dating of plant debris at the base of the ice mass indicates that the oldest part of the ice is more than 700 years old. So far only few studies on describing the microbiome of Ice cave have been performed, and to our knowledge no Planctomycetes isolates have been retrieved from these environments. As the entire chronosequence is exposed inside the cave we have been able to drill ice samples spanning this entire time span. The Ice samples were drilled under sterile conditions and the ice cores were transferred to sterile bottles and transported back to the laboratory in frozen conditions. The samples were then thawed in the dark and most of the water was collected onto Sterivex filters for DNA and RNA extractions. DNA and cDNA was amplified using primers targeting the 16S rRNA gene and sequenced using illumina MiSeg approach. 5 ml of the water were used for two parallel incubations using Planctomycetes specific medium (M30), at 10°C and at room temperature. Enrichments were carefully monitored by examination in light microscope, and when cells with characteristics resembling that of Planctomycetes were observed, these were plated onto solid media (gelrite) for purification. From the descriptive microbial community analyses 6% of the total reads was found to affiliate to Planctomycetes. From the isolation procedure, three novel Planctomycetes was found in 2 different samples, two of the isolates were found in the same sample, incubated at 10°C, and one isolate from the sample incubated at room temperature. One of the isolates formed colonies that were solid and had a rubberlike consistency, whilst the two other isolates formed colonies with a more creamy consistency. Light microscopy revealed that two of the isolates shared oval and egg-shaped cell structure, whilst the other had cells with shapes of a more spherical nature. All isolates showed clear signs of budding reproduction and formed characteristic rosette structures. The unique Planctomycete 16S rRNA gene sequences identified in this study were phylogenetically closely related to sequences from clone sequences from treatment plants in other published studies. The closest cultivable isolate is Pirellula sp. Schlesner strain 302 isolated from chalk mine in Germany (90% 16S sequence similarity). This study reveals new insight to diversity of Planctomycetes and suggests the presence of a novel, cryotolerant genus from glacial environments. 22 Flow Cytometry applied to bacterial studies Alexandre Salvador Enzifarma S.A Flow Cytometry (FCM) is a powerful technic used to analyze particles in a suspension. The analyses can go from a simple comparison on the morphological characteristics of the particles, to their level of autofluorescence. In many cases, the levels of autofluorecence are not enough to distinguish between particles of interest, and morphology is also difficult because of the presence of non-biological particles. In these cases, using dyes to stain cells or cell specific characteristics, cells can be useful to distinguish and correlate parameters. In fact, Flow Cytometry allows multiparametric measurements. Several parameters can be correlated at the same time, emphasizing the properties that we want to highlight. Besides identification, in isolated samples physiological status of the cells can also be addressed. Being able to stain specific structures or access metabolic status, FCM users can do real-time analyses, understanding the biological patterns and consequences of stimulus or inhibitors. Due to a high number of fluorescent probes or molecules that allow to access different cell parameters or stages, in a real research environment FCM became a powerful simple reliable and rapid tool. Because of these characteristics bacterial analyses became more popular. The most common assays involve the capacity to discriminate bacteria from other type of cells, viability assays and functional assays. The biggest question is always: what test to use, what fluorochromes? How to combine? What is the correct protocol? The share of experience and knowledge is highly important and fundamental in this developing area. 23 Hopanoid versus sterol synthesis in PVCs Carlos Santana1, Elena Rivas-Marin1, Sean Stettner2, E. Gottshall2, M Helling2, F Basile2, Naomi Ward2 and Damien Devos1 1 Centro Andaluz de Biología del Desarrollo CABD, Universidad Pablo de Olavide-CSIC, Sevilla, Spain 2 Department of Molecular Biology, Department of Chemistry, University of Wyoming, USA Membrane composition is central to the multiple roles of membranes in the cell. Hopanoids and sterols are, respectively, bacterial and eukaryotic polycyclic triterpenoids with important membrane-ordering function. The two classes of lipids are chemically, structurally, and functionally related. Few bacterial species have been reported to synthesize sterols, and the origins and functions of bacterial sterols are unknown. Most Planctomycetes synthesize hopanoids while Gemmata obscuriglobus is the only PVC member known to synthesize sterols. Here, we addressed this conundrum using bioinformatics, genetic, and chemical approaches. Bioinformatics analyses showed that Planctomycetes have undergone an expansion in the number of hopanoid cyclase genes. In addition, phylogenetic analyses of the sterol synthesis genes suggested that the assumption of lateral gene transfer for acquisition of these genes by G. obscuriglobus should be reconsidered. Given the essential nature of eukaryotic sterols, the logical next step was to investigate the necessity of sterol synthesis for bacterial life, using G. obscuriglobus as an experimental model. We found that sterols are essential for the growth of G. obscuriglobus, and that sterol depletion leads to aberrant membrane structures and budding defects. We also confirmed that chemical inhibition of sterol synthesis leads to dosedependent depletion of membrane sterols, and that both this effect and the related budding defect could be complemented by provision of exogenous lanosterol (the native G. obscuriglobus sterol). To the best of our knowledge, this is the first report of essential sterols in a prokaryotic species. Our work provides a foundation for pursuit of fundamental questions in evolutionary cell biology: Why have some bacteria acquired the ability to produce sterols? What are the advantages of sterols over hopanoids? Why have sterols, and not hopanoids, prevailed in eukaryotes? 24 Planctomycetes distribution along chemical gradient in the meromictic Lake Sælenvannet Julia Endresen Storesund1, Eva-Lena Nordmann1, Hilde Rief Armo1, Anders Lanzèn2 and Lise Øvreås1 1 Department of Biology, University of Bergen, Norway 2 Neiker-Technalia, Spain Lake Sælenvannet is a meromictic lake located south of Bergen, Norway. The 26 m deep lake has a small channel, which connects it via the Nordåsvannet to the open sea. The lake is permanently stratified into two layers. The upper water layer is a brackish layer with major input from of water runoff from the lake surroundings. The bottom layer consist of old saline water, with low or no oxygen concentrations. The interface between these two layers is referred to as “chemocline”. At the chemocline bacteria can have benefits from both layers such as oxygen from the upper layer and reduced sulphur from the bottom layer, and it is often regarded hot spot for prokaryotic activity. Samples throughout the water column were collected and microbial community profile analyses were done using 454 high throughput sequencing in 2012. Planctomycetes related sequences were found both in the oxic and anoxic parts of the lake, but showed an uneven distribution throughout the water column, with the highest relative abundances, 10%, in the saline anoxic layer at 15 m depth, 6 % at 5 meters depth, whereas the surface water and the area around the chemocline had less than 1% planctomycetes reads. Subsequently, samples for enrichment and isolation of novel Plantomycetes were collected from seven different depths of the lake in 2014. Ten novel isolates from four different genera affiliating to the Planctomycetaceae family were obtained in pure culture. Of these, two strains representing new species within the Rhodopirellula and Gimesia genera were isolated from the chemocline at seven and nine meters depth respectively. A second strain obtained from nine meters depth was closely related to Blastopirellula cremea. Seven isolates retrieved from seven different depths with extensively different salinities and chemical compositions showed identical 16S rRNA gene sequences, and likely represent a new species closely associated with the Rubinisphaera genus. The presence of this novel planctomycete in all water depths spanning the entire chemical gradient could indicate high phenotypic plasticity in this isolate or a very efficient survival strategy. Overall, our results indicate the presence of a diverse group of Planctomycetes present in Lake Sælenvannet, with a strong potential for novel adaptations to chemical stress factors. 25 The energy-conserving prokaryotic organelle of anammox Planctomycetes Laura van Niftrik Microbiology, Institute for Water & Wetland Research, Faculty of Science, Radboud University, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands Anammox bacteria perform anaerobic ammonium oxidation with nitrite as electron acceptor and nitrogen gas as end product. Intermediates of the anammox reaction are nitric oxide and the “rocket fuel” hydrazine. Anammox bacteria belong to the phylum Planctomycetes and are recognized as major players in the global nitrogen cycle. It is estimated that anammox bacteria are responsible for up to 50% of the nitrogen in the air that we breathe. In addition, anammox bacteria are extremely valuable for wastewater treatment where they are applied for the cost-effective and environment-friendly removal of nitrogen compounds. Anammox bacteria harbor a major intracellular compartment called the anammoxosome (Fig. 1) which is the location of the anammox reaction. Currently we are investigating how the substrates ammonium and nitrite are transported through the cell to the anammoxosome and how the anammox reaction is coupled to the anammoxosome membrane to conserve energy. To this end, we use metagenome, transcriptome and proteome analysis, subcellular (membrane) fractionation, biochemistry of purified proteins, activity assays and cryo-transmission electron microscopy. The aim is to unravel the molecular and biochemical pathway of anaerobic ammonium oxidation. Fig. 1. Transmission electron micrograph and schematic model of the cell plan of anammox bacteria. Scale bar: 200 nm. 26 Diversity driven cultivation and genome sequencing of 80 slow growing Planctomycetes reveal their potential for small molecule production Sandra Wiegand1, Mareike Jogler2, Anja Heuer2, Patrick Rast2, Anne-Kristin Kaster2, Olga M. Lage3, Lise Øvreås4, Laura van Niftrik1, John Vollmers2 and Christian Jogler1 1 Microbiology, Institute for Water & Wetland Research, Faculty of Science, Radboud University, Nijmegen, The Netherlands 2 Microbial Cell Biology and Genetics, Leibniz Institute DSMZ, Braunschweig, Germany 3 University of Porto, Porto, Portugal 4 University of Bergen, Bergen, Norway Members of the taxon Planctomycetes, that belongs to the PVC superphylum, are characterized by many unusual features. They show a life style switch during which the sessile mother cell releases a swimming daughter cell after FtsZ-independent binary fission or polar budding. Also, planctomycetal genomes encode numerous giant genes and bear the genomic potential for secondary metabolite production. As the whole clade is notably under-sampled with relatively few genomic sequences being publicly available, we focused our efforts on increasing the data basis for comparative studies. For this purpose, we sampled multiple aquatic habitats around the globe and isolated about 80 new planctomycetal strains from various biotic and abiotic surfaces. Planctomycetes were enriched by targeting the slow growing antibiotic resistant bacteria in combination with selective carbon sources. Most of these strains represent novel taxa up to the level of a new order, capturing an unpreceded degree of diversity. In total, we brought more novel Planctomycetes into pure culture than currently described species of this phylum exist. To allow detailed analyses we produced closed as well as high-quality draft genomes of the novel isolates. The exploration of the gained genomic information not only enables deep insights into the taxonomy of the phylum and its subtaxa, but also the definition of its core and pan genome. The latter increases our knowledge on the metabolic versatility and the environmental role of this phylum. Furthermore, we identified secondary metabolite related genes and further strengthened our hypothesis of Planctomycetes as talented producers. 27 Participants List Eduarda Almeida FCUP, University of Porto, Portugal [email protected] Centro Andaluz de Biología del Desarrollo CABD, Universidad Pablo de Olavide-CSIC, Spain [email protected] Siv Andersson Department of Cell and Molecular Biology, Uppsala University, Sweden [email protected] Clara Belzer Laboratory of Microbiology, Wageningen University, The Netherlands [email protected] Rita Calisto FCUP, University of Porto, Portugal [email protected] Teresa Carvalho FCUP, University of Porto, Portugal [email protected] Laura Claret Centro Andaluz de Biología del Desarrollo CABD, Universidad Pablo de Olavide-CSIC, Spain [email protected] Astrid Collingro Division of Microbial Ecology, Department of Microbiology and Ecosystem Science, University of Vienna, Austria [email protected] Milton da Costa Department of Life Sciences, University of Coimbra [email protected] Svetlana Dedysh Laboratory of Wetland Microbiology, Winogradsky Institute of Microbiology, Russia [email protected] Damien P. Devos Teresa Dias FCUP, University of Porto, Portugal [email protected] Michael Galperin NCBI, NLM, National Institutes of Health, USA [email protected] Ofélia Godinho Universidade de Aveiro, Portugal [email protected] Ana Patrícia Graça Synthetic Microbiology – Hans Knöll Institute, Germany [email protected] Matthias Horn Division of Microbial Ecology, Department of Microbiology and Ecosystem Science, University of Vienna, Austria [email protected] Anastasia Ivanova Winogradsky Institute of Microbiology, Research Center of Biotechnology RAS, Moscow, Russia [email protected] Christian Jogler Faculty of Science, Radboud University Nijmegen, The Netherlands [email protected] Timo Kohn Faculty of Science, Radboud University Nijmegen, The Netherlands [email protected] Irina Kulichevskaya Laboratory of Wetland Microbiology, Winogradsky Institute of Microbiology, Russia 28 [email protected] Eirik Færøy Sæbø Olga Maria Lage Department of Biology, University of Bergen, Norway [email protected] FCUP, University of Porto, Portugal [email protected] Alexandre Lobo da Cunha ICBAS, University of Porto, Portugal [email protected] Conceição Marinho FCUP, University of Porto, Portugal [email protected] Enrique Merino Universidad Nacional Autonoma de Mexico, Mexico [email protected] Alexandre Salvador Enzifarma, Portugal [email protected] Carlos Santana Molina Centro Andaluz de Biología del Desarrollo CABD, Universidad Pablo de Olavide-CSIC, Spain [email protected] José Diogo Santos FCUP, University of Porto, Portugal [email protected] Lise Øvreås Department of Biology, University of Bergen, Norway [email protected] Claudia Serra Mónia Pedrosa Enzifarma, Portugal [email protected] Sean Stettner Departments of Molecular Biology and Botany, Wyoming INBRE Bioinformatics Core, USA [email protected] Ana Mafalda Pinto FCUP and CIIMAR, University of Porto, Portugal [email protected] FCUP, University of Porto, Portugal [email protected] Carlos Ramos FCUP, University of Porto, Portugal Julia Endresen Storesund Department of Biology, University of Bergen, Norway [email protected] [email protected] Susana Ribeiro Enzifarma, Portugal [email protected] Elena Rivas Marin Centro Andaluz de Biología del Desarrollo CABD, Universidad Pablo de Olavide-CSIC, Spain [email protected] Laura van Niftrik Institute for Water & Wetland Research, Faculty of Science, Radboud University, The Netherlands [email protected] Naomi Louise Ward Departments of Molecular Biology and Botany, Wyoming INBRE Bioinformatics Core, USA [email protected] Inês Vitorino FCUP, University of Porto, Portugal [email protected] Sandra Wiegand Faculty of Science, Radboud University Nijmegen, The Netherlands [email protected] 29