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Staff Publications

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    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

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Comparative genomics, symbiotic capacities and high metabolic flexibility of the marine genus Pseudovibrio
Versluis, D. ; Nijsse, B. ; Mohamad, M.A.N. ; Koehorst, J.J. ; Wiese, Jutta ; Imhoff, Johannes F. ; Schaap, P.J. ; Passel, M.W.J. van; Smidt, H. ; Sipkema, D. - \ 2017
PRJEB20602 - ERP022771 - Pseudovibrio - comparative genomics
Pseudovibrio is a marine bacterial genus, members of which are predominantly isolated from sessile marine animals, and particularly sponges. It has been hypothesized that they form a mutualistic relationship with their hosts. Here, we studied their phylogeny highlighting genetic adaptations that may play a role in host colonization, by comparative genomics of 31 Pseudovibrio strains of which 25 were isolated from sponges.
Comparative genomics of the genus Desulfitobacterium
Kruse, Thomas ; Goris, Tobias ; Maillard, Julien ; Woyke, Tanja ; Lechner, Ute ; Vos, Willem de; Smidt, Hauke - \ 2017
FEMS Microbiology Ecology 93 (2017)12. - ISSN 0168-6496
comparative genomics - Desulfitobacterium spp. - genome - metabolic potential - organohalide respiration - rdhA
The Desulfitobacterium genus comprises anaerobic Gram-positive bacteria, of which the majority are facultative organohalide respirers. We here present the genomes of eight strains of Desulfitobacterium spp., including five strains of Desulfitobacterium hafniense, one strain each from D. dichloroeliminans and D. metallireducens, and one strain that had not been assigned to any species prior to this study. The newly sequenced genomes were compared with four previously published desulfitobacterial genomes. The average genome sizes are 5.5, 4.3 and 3.4 Mbp for D. hafniense, D. dehalogenans and D. dichloroeliminans/metallireducens, respectively. The genomes encode up to seven reductive dehalogenases, the genomes of both D. hafniense DP7 and D. metallireducens 853-15AT did not encode any reductive dehalogenase. The latter result was a surprise as D. metallireducens 853-15AT has been reported to carry out organohalide respiration. Unlike reported for the pceABCT gene cluster, the other reductive dehalogenase gene clusters do not show any signs of being genetically mobile. All analyzed desulfitobacterial genomes encode a complete cobalamin synthesis pathway. A menaquinone synthesis pathway was found in all strains except D. dichloroeliminans DCA1T. The detailed analysis of the genome sequence of 12 desulfitobacteria from four different species confirmed that this genus has an extremely large metabolic repertoire.
High-throughput comparative genomics for plant breeding and its application in the tomato clade
Aflitos, S.A. - \ 2015
University. Promotor(en): Hans de Jong, co-promotor(en): Dick de Ridder; Sander Peters. - Wageningen : Wageningen University - ISBN 9789462576131 - 198
solanum lycopersicum - tomatoes - plant breeding - comparative genomics - tomaten - plantenveredeling - vergelijkende genomica
A genetic linkage map of Sole (Solea solea): A tool for evolutionary and comparative analyses of exploited (flat)fishes
Diopere, E. ; Maes, G.E. ; Komen, J. ; Volckaert, F.A.M. ; Groenen, M. - \ 2014
PLoS One 9 (2014)12. - ISSN 1932-6203 - 16 p.
turbot scophthalmus-maximus - common sole - microsatellite markers - comparative genomics - marine fishes - populations - time - divergence - pleuronectiformes - senegalensis
Linkage maps based on markers derived from genes are essential evolutionary tools for commercial marine fish to help identify genomic regions associated with complex traits and subject to selective forces at play during exploitation or selective breeding. Additionally, they allow the use of genomic information from other related species for which more detailed information is available. Sole (solea solea L.) is a commercially important flatfish species in the North Sea, subject to overexploitation and showing evidence of fisheries-induced evolutionary changes in growth- and maturation-related traits. Sole would definitely benefit from a linkage map to better understand how evolution has shaped its genome structure. This study presents a linkage map of sole based on 423 single nucleotide polymorphisms derived from expressed sequence tags and 8 neutral microsatellite markers. The total map length is 1233.8 cM and consists of 38 linkage groups with a size varying between 0 to 92.1 cM. Being derived from expressed sequence tags allowed us to align the map with the genome of four model fish species, namely medaka (Oryzias latipes), Nile tilapia (Oreochromis niloticus), three-spined stickleback (Gasterosteus aculeatus) and green spotted pufferfish (Tetraodon nigroviridis). This comparison revealed multiple conserved syntenic regions with all four species, and suggested that the linkage groups represent 21 putative sole chromosomes. The map was also compared to the linkage map of turbot (Scophthalmus maximus), another commercially important flatfish species and closely related to sole. For all putative sole chromosomes (except one) a turbot homolog was detected, confirming the even higher degree of synteny between these two flatfish species.
Group VII Ethylene Response Factor diversification and regulation in four species from flood-prone environments.
Veen, H. van; Akman, M. ; Jamar, D.C.L. ; Vreugdenhil, D. ; Kooiker, M. ; Tienderen, P.H. van; Voesenek, L.A.C.J. ; Schranz, M.E. ; Sasidharan, R. - \ 2014
Plant, Cell & Environment 37 (2014)10. - ISSN 0140-7791 - p. 2421 - 2432.
end rule pathway - submergence tolerance - transcription factors - arabidopsis-thaliana - comparative genomics - rorippa-sylvestris - gene-expression - rice - hypoxia - plants
Flooding events negatively affect plant performance and survival. Flooding gradients thereby determine the dynamics in vegetation composition and species abundance. In adaptation to flooding, the group VII Ethylene Response Factor genes (ERF-VIIs) play pivotal roles in rice and Arabidopsis through regulation of anaerobic gene expression and antithetical survival strategies. We investigated if ERF-VIIs have a similar role in mediating survival strategies in eudicot species from flood-prone environments. Here, we studied the evolutionary origin and regulation of ERF-VII transcript abundance and the physiological responses in species from two genera of divergent taxonomic lineages (Rumex and Rorippa). Synteny analysis revealed that angiosperm ERF-VIIs arose from two ancestral loci and that subsequent diversification and duplication led to the present ERF-VII variation. We propose that subtle variation in the regulation of ERF-VII transcript abundance could explain variation in tolerance among Rorippa species. In Rumex, the main difference in flood tolerance correlated with the genetic variation in ERF-VII genes. Large transcriptional differences were found by comparing the two genera: darkness and dark submergence-induced Rumex ERF-VIIs, whereas HRE2 expression was increased in submerged Rorippa roots. We conclude that the involvement of ERF-VIIs in flooding tolerance developed in a phylogenetic-dependent manner, with subtle variations within taxonomic clades.
Filling the gap between sequence and function: a bioinformatics approach
Bargsten, J.W. - \ 2014
University. Promotor(en): Richard Visser, co-promotor(en): Jan-Peter (Jp) Nap. - Wageningen : Wageningen University - ISBN 9789462570764 - 170
bio-informatica - planten - genomica - nucleotidenvolgordes - functionele genomica - vergelijkende genomica - vergelijkende genetische kartering - genomen - genetische kartering - plantenveredeling - methodologie - bioinformatics - plants - genomics - nucleotide sequences - functional genomics - comparative genomics - comparative mapping - genomes - genetic mapping - plant breeding - methodology

The research presented in this thesis focuses on deriving function from sequence information, with the emphasis on plant sequence data. Unravelling the impact of genomic elements, in most cases genes, on the phenotype of an organism is a major challenge in biological research and modern plant breeding. An important part of this challenge is the (functional) annotation of such genomic elements. Currently, wet lab experiments may provide high quality, but they are laborious and costly. With the advent of next generation sequencing platforms, vast amounts of sequence data are generated. This data are used in connection with the available experimental data to derive function from a bioinformatics perspective.

The connection between sequence information and function was approached on the level of chromosome structure (chapter 2) and of gene families (chapter 3) using combinations of existing bioinformatics tools. The applicability of using interaction networks for function prediction was demonstrated by first markedly improving an existing method (chapter 4) and by exploring the role of network topology in function prediction (chapter 5). Taken together, the combination of methods and results presented indicate the potential as well as the current state-of-the-art of function prediction in (plant) bioinformatics.

Chapter 1 introduces the basis for the approaches used and developed in this thesis. This includes the concepts of genome annotation, comparative genomics, gene function prediction and the analysis of network topology for gene function prediction. A requirement for the study of any new organism is the sequencing and annotation of its genome. Current genome annotation is divided into structural identification and functional categorization of genomic elements. The de facto standard for categorizing functional annotation is provided by the Gene Ontology. The Gene Ontology is divided into three domains, molecular function, biological process and cellular component. Approaches to predict molecular function and biological process are outlined. Accurate function prediction generally relies on existing input data, often of experimental origin, that can be transferred to unannotated genomic elements. Plants often lack such input data, which poses a big challenge for current function prediction algorithms. In unravelling the function of genomic elements, comparative genomics is an important approach. Via the comparison of multiple genomes it gives insights into evolution, function as well as genomic structure and variation. Comparative genomics has become an essential toolkit for the analysis of newly sequenced organisms. Often bioinformatics methods need to be adapted to the specific needs of plant genome research. With a focus on the commercially important crop plants tomato and potato, specific requirements of plant bioinformatics, such as the high amount of repetitive elements and the lack of experimental data, are outlined.

In chapter 2, the structural homology of the long arm of chromosome 2 (2L) of tomato, potato and pepper is analyzed. Molecular organization and collinear junctions are delineated using multi-color BAC FISH analysis and comparative sequence alignment. We identify several large-scale rearrangements including inversions and segmental translocations that were not reported in previous comparative studies. Some of the structural rearrangements are specific for the tomato clade, and differentiate tomato from potato, pepper and other solanaceous species. There are many small-scale synteny perturbations, but local gene vicinity is largely preserved. The data suggests that long distance intra-chromosomal rearrangements and local gene rearrangements have evolved frequently during speciation in the Solanum genus, and that small changes are more prevalent than large-scale differences. The occurrence of transposable elements and other repeats near or at junction breaks may indicate repeat-mediated rearrangements. The ancestral 2L topology is reconstructed and the evolutionary events leading to the current topology are discussed.

In chapter 3, we analyze the Snf2 gene family. As part of large protein complexes, Snf2 family ATPases are responsible for energy supply during chromatin remodeling, but the precise mechanism of action of many of these proteins is largely unknown. They influence many processes in plants, such as the response to environmental stress. The analysis is the first comprehensive study of Snf2 family ATPases in plants. Some subfamilies of the Snf2 gene family are remarkably stable in number of genes per genome, whereas others show expansion and contraction in several plants. One of these subfamilies, the plant-specific DRD1 subfamily, is non-existent in lower eukaryote genomes, yet it developed into the largest Snf2 subfamily in plant genomes. It shows the occurrence of a complex series of evolutionary events. Its expansion, notably in tomato, suggests novel functionality in processes connected to chromatin remodeling. The results underpin and extend the Snf2 subfamily classification, which could help to determine the various functional roles of Snf2 ATPases and to target environmental stress tolerance and yield in future breeding with these genes.

In chapter 4, a new approach to improve the prediction of protein function in terms of biological processes is developed that is particularly attractive for sparsely annotated plant genomes. The combination of the network-based prediction method Bayesian Markov Random Field (BMRF) with the sequence-based prediction method Argot2 shows significantly improved performance compared to each of the methods separately, as well as compared to Blast2GO. The approach was applied to predict biological processes for the proteomes of rice, barrel clover, poplar, soybean and tomato. Analysis of the relationships between sequence similarity and predicted function similarity identifies numerous cases of divergence of biological processes in which proteins are involved, in spite of sequence similarity. Examples of potential divergence are identified for various biological processes, notably for processes related to cell development, regulation, and response to chemical stimulus. Such divergence in biological process annotation for proteins with similar sequences should be taken into account when analyzing plant gene and genome evolution. This way, the integration of network-based and sequence-based function prediction will strengthen the analysis of evolutionary relationships of plant genomes.

In chapter 5 the influence of network topology on network-based function prediction algorithms is investigated. The analysis of biological networks using algorithms such as Bayesian Markov Random Field (BMRF) is a valuable predictor of the biological processes that proteins are involved in. The topological properties and constraints that determine prediction performance in such networks are however largely unknown. This chapter presents analyses based on network centrality measures, such as node degree, to evaluate the performance of BMRF upon progressive removal of highly connected hub nodes (pruning). Three different protein-protein interaction networks with data from Arabidopsis, human and yeast were analyzed. All three show that the average prediction performance can improve significantly. The chapter paves the way for further improvement of network-based function prediction methods based on node pruning.

Chapter 6 discusses the results and methods developed in this thesis in the context of the vast amount of generated sequencing data. Sequencing or re-sequencing a (plant) genome has become fairly straightforward and affordable, but the interpretation for subsequent use of this sequence data is far from trivial. The topics addressed in this thesis, annotation of function, analysis of genome structure and identifying genomic variation, focus on this main bottleneck of biological research. Issues discussed in connection with this work and its future are data accuracy, error propagation, possible improvements and future implications for biological research in crop plants. In particular the shift of costs from sequencing to downstream analyses, with functional genome annotation as essential step, is covered. One of the biggest challenges biology and bioinformatics will face is the integration of results from such downstream analyses and other sources into a complete picture. Only this will allow understanding of complex biological systems.

Comparative genomics of Dothideomycete fungi
Burgt, A. van der - \ 2014
University. Promotor(en): Pierre de Wit, co-promotor(en): Jerome Collemare. - Wageningen : Wageningen University - ISBN 9789461739056 - 176
dothideomycetes - plantenziekteverwekkende schimmels - passalora fulva - dothistroma - genomen - vergelijkende genomica - plant pathogenic fungi - genomes - comparative genomics

Fungi are a diverse group of eukaryotic micro-organisms particularly suited for comparative genomics analyses. Fungi are important to industry, fundamental science and many of them are notorious pathogens of crops, thereby endangering global food supply. Dozens of fungi have been sequenced in the last decade and with the advances of the next generation sequencing, thousands of new genome sequences will become available in coming years. In this thesis I have used bioinformatics tools to study different biological and evolutionary processes in various genomes with a focus on the genomes of the Dothideomycetefungi Cladosporium fulvum, Dothistroma septosporumand Zymoseptoria tritici.

Chapter 1introduces the scientific disciplines of mycology and bioinformatics from a historical perspective. It exemplifies a typical whole-genome sequence analysis of a fungal genome, and focusses in particular on structural gene annotation and detection of transposable elements. In addition it shortly reviews the microRNA pathway as known in animal and plants in the context of the putative existence of similar yet subtle different small RNA pathways in other branches of the eukaryotic tree of life.

Chapter 2addresses the novel sequenced genomes of the closely related Dothideomyceteplant pathogenic fungi Cladosporium fulvumand Dothistroma septosporum. Remarkably, it revealed occurrence of a surprisingly high similarity at the protein level combined with striking differences at the DNA level, gene repertoire and gene expression. Most noticeably, the genome of C. fulvumappears to be at least twice as large, which is solely attributable to a much larger content in repetitive sequences.

Chapter 3describes a novel alignment-based fungal gene prediction method (ABFGP) that is particularly suitable for plastic genomes like those of fungi. It shows excellent performance benchmarked on a dataset of 7,000 unigene-supported gene models from ten different fungi. Applicability of the method was shown by revisiting the annotations of C. fulvumand D. septosporumand of various other fungal genomes from the first-generation sequencing era. Thousands of gene models were revised in each of the gene catalogues, indeed revealing a correlation to the quality of the genome assembly, and to sequencing strategies used in the sequencing centres, highlighting different types of errors in different annotation pipelines.

Chapter 4focusses on the unexpected high number of gene models that were identified by ABFGP that align nicely to informant genes, but only upon toleration of frame shifts and in-frame stop-codons. These discordances could represent sequence errors (SEs) and/or disruptive mutations (DMs) that caused these truncated and erroneous gene models. We revisited the same fungal gene catalogues as in chapter 3, confirmed SEs by resequencing and successively removed those, yielding a high-confidence and large dataset of nearly 1,000 pseudogenes caused by DMs. This dataset of fungal pseudogenes, containing genes listed as bona fide genes in current gene catalogues, does not correspond to various observations previously done on fungal pseudogenes. Moreover, the degree of pseudogenization showing up to a ten-fold variation for the lowest versus the highest affected species, is generally higher in species that reproduce asexually compared to those that in addition reproduce sexually.

Chapter 5describes explorative genomics and comparative genomics analyses revealing the presence of introner-like elements (ILEs) in various Dothideomycetefungi including Zymoseptoria triticiin which they had not identified yet, although its genome sequence is already publicly available for several years. ILEs combine hallmark intron properties with the apparent capability of multiplying themselves as repetitive sequence. ILEs strongly associate with events of intron gain, thereby delivering in silico proof of their mobility. Phylogenetic analyses at the intra- and inter-species level showed that most ILEs are related and likely share common ancestry.

Chapter 6provides additional evidence that ILE multiplication strongly dominates over other types of intron duplication in fungi. The observed high rate of ILE multiplication followed by rapid sequence degeneration led us to hypothesize that multiplication of ILEs has been the major cause and mechanism of intron gain in fungi, and we speculate that this could be generalized to all eukaryotes.

Chapter 7describes a new strategy for miRNA hairpin prediction using statistical distributions of observed biological variation of properties (descriptors) of known miRNA hairpins. We show that the method outperforms miRNA prediction by previous, conventional methods that usually apply threshold filtering. Using this method, several novel candidate miRNAs were assigned in the genomes of Caenorhabditis elegansand two human viruses. Although this chapter is not applied on fungi, the study does provide a flexible method to find evidence for existence of a putative miRNA-like pathway in fungi.

Chapter 8provides a general discussion on the advent of bioinformatics in mycological research and its implications. It highlights the necessity of a prioriplanning and integration of functional analysis and bioinformatics in order to achieve scientific excellence, and describes possible scenarios for the near future of fungal (comparative) genomics research. Moreover, it discusses the intrinsic error rate in large-scale, automatically inferred datasets and the implications of using and comparing those.

High-resolution mapping of the barley Ryd3 locus controlling tolerance to BYDV
Lüpken, T. ; Stein, N. ; Perovic, D. ; Habekuss, A. ; Serfling, A. ; Krämer, I. ; Hähnel, U. ; Steuernagel, B. ; Scholz, U. ; Ariyadasa, R. ; Martis, M. ; Mayer, K. ; Niks, R.E. ; Collins, N.C. ; Friedt, W. ; Ordon, F. - \ 2014
Molecular Breeding 33 (2014)2. - ISSN 1380-3743 - p. 477 - 488.
yellow-dwarf-virus - hordeum-vulgare l. - recessive bymovirus resistance - leaf rust resistance - comparative genomics - consensus map - winter barley - linkage map - yd2 gene - sequence
Barley yellow dwarf disease (BYD) is transmitted by aphids and is caused by different strains of Barley yellow dwarf virus (BYDV) and Cereal yellow dwarf virus (CYDV). Economically it is one of the most important diseases of cereals worldwide. Besides chemical control of the vector, growing of tolerant/resistant cultivars is an effective way of protecting crops against BYD. The Ryd3 gene in barley (Hordeum vulgare L.) confers tolerance to BYDV-PAV and BYDV-MAV and the locus was previously mapped on the short arm of barley chromosome 6H near the centromere. We applied a strategy for high-resolution mapping and marker saturation at the Ryd3 locus by exploiting recent genomic tools available in barley. In a population of 3,210 F2 plants, 14 tightly linked markers were identified, including 10 that co-segregated with Ryd3. The centromeric region where Ryd3 is located suffers suppressed recombination or reduced recombination rate, suggesting potential problems in achieving (1) map-based cloning of Ryd3 and (2) marker selection of the resistance in breeding programmes without the introduction of undesirable traits via linkage drag.
Birth, death and horizontal transfer of the fumonisin biosynthetic gene cluster during the evolutionary diversification of Fusarium
Proctor, R.H. ; Hove, F. van; Susca, A. ; Stea, G. ; Busman, M. ; Lee, T.A.J. van der; Waalwijk, C. ; Moretti, A. ; Ward, T.J. - \ 2013
Molecular Microbiology 90 (2013)2. - ISSN 0950-382X - p. 290 - 306.
fujikuroi species complex - maximum-likelihood - gibberella-moniliformis - aspergillus-parasiticus - secondary metabolism - comparative genomics - adaptive evolution - multigene family - section liseola - mixed models
Fumonisins are a family of carcinogenic secondary metabolites produced by members of the Fusarium fujikuroi species complex (FFSC) and rare strains of Fusarium oxysporum. In Fusarium, fumonisin biosynthetic genes (FUM) are clustered, and the cluster is uniform in gene organization. Here, sequence analyses indicated that the cluster exists in five different genomic contexts, defining five cluster types. In FUM gene genealogies, evolutionary relationships between fusaria with different cluster types were largely incongruent with species relationships inferred from primary-metabolism (PM) gene genealogies, and FUM cluster types are not trans-specific. In addition, synonymous site divergence analyses indicated that three FUM cluster types predate diversification of FFSC. The data are not consistent with balancing selection or interspecific hybridization, but they are consistent with two competing hypotheses: (i) multiple horizontal transfers of the cluster from unknown donors to FFSC recipients and (ii) cluster duplication and loss (birth and death). Furthermore, low levels of FUM gene divergence in F.¿bulbicola, an FFSC species, and F.¿oxysporum provide evidence for horizontal transfer of the cluster from the former, or a closely related species, to the latter. Thus, uniform gene organization within the FUM cluster belies a complex evolutionary history that has not always paralleled the evolution of Fusarium
Selective carbohydrate utilization by lactobacilli and bifidobacteria
Watson, D. ; O'Connell Motherway, M. ; Schoterman, M.H.C. ; Neerven, R.J.J. van; Nauta, A. ; Sinderen, D. van - \ 2013
Journal of Applied Microbiology 114 (2013)4. - ISSN 1364-5072 - p. 1132 - 1146.
galacto-oligosaccharides - comparative genomics - beta-galactosidase - gut microbiome - prebiotics - growth - inulin - milk - oligofructose - modulation
Aim - To evaluate the ability of specific carbohydrates, including commercially available products, to support the growth of representatives of two well-known groups of gut commensals, namely lactobacilli and bifidobacteria. Methods and Results - Sixty-eight bacterial strains, representing 29 human-derived lactobacilli and 39 bifidobacteria (both human- and animal-derived), were tested for their ability to metabolize 10 different carbohydrates. Analysis of growth and metabolic activity was performed using a combination of diagnostic parameters, such as final OD600, final pH, fermentation end products and growth rate. Conclusions - The data assembled in this study provide significant complementary and comparative information on the growth-promoting properties of a range of carbohydrates, while also investigating interspecies differences between lactobacilli and/or bifidobacteria with regard to their carbohydrate utilization abilities. Galacto-oligosaccharides (GOS) and lactulose were shown to support the most favourable growth characteristics, whereas relatively poor growth of lactobacilli and bifidobacteria was observed on inulin, maltodextrin and polydextrose. GOS/inulin (9 : 1) and fructo-oligosaccharides (FOS)/inulin mixtures supported mostly similar growth abilities to those obtained for GOS and FOS, respectively. Microbial consumption of GOS, as determined by high-performance anion-exchange chromatography with pulsed amperometric detection, was evident for both lactobacilli and bifidobacteria.
Transcriptome signatures of class I and III stress response deregulation in Lactobacillus plantarum reveal pleiotropic adaptation
Bokhorst-van de Veen, H. van; Bongers, R.S. ; Wels, M. ; Bron, P.A. ; Kleerebezem, M. - \ 2013
Microbial Cell Factories 12 (2013)1. - ISSN 1475-2859 - 15 p.
gram-positive bacteria - heat-shock response - lactic-acid bacteria - bacillus-subtilis - listeria-monocytogenes - gastrointestinal-tract - low gc - streptococcus-pneumoniae - comparative genomics - helicobacter-pylori
Background - To cope with environmental challenges bacteria possess sophisticated defense mechanisms that involve stress-induced adaptive responses. The canonical stress regulators CtsR and HrcA play a central role in the adaptations to a plethora of stresses in a variety of organisms. Here, we determined the CtsR and HrcA regulons of the lactic acid bacterium Lactobacillus plantarum WCFS1 grown under reference (28°C) and elevated (40°C) temperatures, using ctsR, hrcA, and ctsR-hrcA deletion mutants. Results - While the maximum specific growth rates of the mutants and the parental strain were similar at both temperatures (0.33¿±¿0.02 h-1 and 0.34¿±¿0.03 h-1, respectively), DNA microarray analyses revealed that the CtsR or HrcA deficient strains displayed altered transcription patterns of genes encoding functions involved in transport and binding of sugars and other compounds, primary metabolism, transcription regulation, capsular polysaccharide biosynthesis, as well as fatty acid metabolism. These transcriptional signatures enabled the refinement of the gene repertoire that is directly or indirectly controlled by CtsR and HrcA of L. plantarum. Deletion of both regulators, elicited transcriptional changes of a large variety of additional genes in a temperature-dependent manner, including genes encoding functions involved in cell-envelope remodeling. Moreover, phenotypic assays revealed that both transcription regulators contribute to regulation of resistance to hydrogen peroxide stress. The integration of these results allowed the reconstruction of CtsR and HrcA regulatory networks in L. plantarum, highlighting the significant intertwinement of class I and III stress regulons. Conclusions - Taken together, our results enabled the refinement of the CtsR and HrcA regulatory networks in L. plantarum, illustrating the complex nature of adaptive stress responses in this bacterium
Extensive chromosomal reshuffling drives evolution of virulence in an asexual pathogen
Jonge, R. de; Bolton, M.D. ; Kombrink, A. ; Berg, G.C.M. van den; Yadeta, K.A. ; Thomma, B.P.H.J. - \ 2013
Genome Research 23 (2013). - ISSN 1088-9051 - p. 1271 - 1282.
rice blast fungus - verticillium-albo-atrum - filamentous ascomycete fungi - chitin-triggered immunity - de-novo identification - magnaporthe-oryzae - mycosphaerella-graminicola - comparative genomics - resistance genes - rna-seq
Sexual recombination drives genetic diversity in eukaryotic genomes and fosters adaptation to novel environmental challenges. Although strictly asexual microorganisms are often considered as evolutionary dead ends, they comprise many devastating plant pathogens. Presently, it remains unknown how such asexual pathogens generate the genetic variation that is required for quick adaptation and evolution in the arms race with their hosts. Here we show that extensive chromosomal rearrangements in the strictly asexual plant pathogenic fungus Verticillium dahliae establish highly dynamic lineage-specific (LS) genomic regions that act as a source for genetic variation to mediate aggressiveness. We show that such LS regions are greatly enriched for in planta-expressed effector genes, encoding secreted proteins that enable host colonization. The LS regions occur at the flanks of chromosomal breakpoints and are enriched for retrotransposons and other repetitive sequence elements. Our results demonstrate that asexual pathogens may evolve by prompting chromosomal rearrangements, enabling rapid development of novel effector genes. Likely, chromosomal reshuffling can act as a general mechanism for adaptation in asexually propagating organisms.
Role of phosphate in the central metabolism of two lactic acid bacteria-a comparative systems biology approach
Levering, J. ; Musters, M.W.J.M. ; Bekker, M. ; Bellomo, D. ; Fiedler, T. ; Vos, W.M. de; Hugenholtz, F. ; Kreikemeyer, B. ; Kummer, U. ; Teusink, B. - \ 2012
FEBS Journal 279 (2012)7. - ISSN 1742-464X - p. 1274 - 1290.
pyruvate formate-lyase - group-a streptococci - lactococcus-lactis - phosphotransferase system - lactate-dehydrogenase - comparative genomics - in-vivo - glyceraldehyde-3-phosphate dehydrogenase - enterococcus-faecalis - inducer expulsion
Lactic acid-producing bacteria survive in distinct environments, but show common metabolic characteristics. Here we studied the dynamic interactions of the central metabolism in Lactococcus lactis, extensively used as a starter culture in the dairy industry, and Streptococcus pyogenes, a human pathogen. Glucose-pulse experiments and enzymatic measurements were performed to parameterize kinetic models of glycolysis. Significant improvements were made to existing kinetic models for L. lactis, which subsequently accelerated the development of the first kinetic model of S. pyogenes glycolysis. The models revealed an important role for extracellular phosphate in the regulation of central metabolism and the efficient use of glucose. Thus, phosphate, which is rarely taken into account as an independent species in models of central metabolism, should be considered more thoroughly in the analysis of metabolic systems in the future. Insufficient phosphate supply can lead to a strong inhibition of glycolysis at high glucose concentrations in both species, but this was more severe in S. pyogenes. S. pyogenes is more efficient at converting glucose to ATP, showing a higher tendency towards heterofermentative energy metabolism than L. lactis. Our comparative systems biology approach revealed that the glycolysis of L. lactis and S. pyogenes have similar characteristics, but are adapted to their individual natural habitats with respect to phosphate regulation
The Distribution of Mobile Genetic Elements (MGEs) in MRSA CC398 Is Associated with Both Host and Country
McCarthy, A.J. ; Witney, A.A. ; Gould, K.A. ; Moodley, A. ; Guardabassi, L. ; Voss, A. ; Denis, O. ; Broens, E.M. ; Hinds, J. ; Lindsay, J.A. - \ 2011
Genome Biology and Evolution 3 (2011). - ISSN 1759-6653 - p. 1164 - 1174.
resistant staphylococcus-aureus - factor-binding-protein - methicillin-resistant - comparative genomics - st398 - pigs - infection - humans - specificity - adaptation
Methicillin-resistant Staphylococcus aureus clonal complex (CC) 398 has emerged from pigs to cause human infections in Europe and North America. We used a new 62-strain S. aureus microarray (SAM-62) to compare genomes of isolates from three geographical areas ( Belgium, Denmark, and Netherlands) to understand how CC398 colonizes different mammalian hosts. The core genomes of 44 pig isolates and 32 isolates from humans did not vary. However, mobile genetic element (MGE) distribution was variable including SCCmec. phi 3 bacteriophage and human specificity genes (chp, sak, scn) were found in invasive human but not pig isolates. SaP15 and putative ruminant specificity gene variants (vwb and scn) were common but not pig specific. Virulence and resistance gene carriage was host associated but country specific. We conclude MGE exchange is frequent in CC398 and greatest among populations in close contact. This feature may help determine epidemiological associations among isolates of the same lineage.
Reconstruction of the regulatory network of Lactobacillus plantarum WCFS1 on basis of correlated gene expression and conserved regulatory motifs
Wels, M. ; Overmars, L. ; Francke, C. ; Kleerebezem, M. ; Siezen, R.J. - \ 2011
Microbial Biotechnology 4 (2011)3. - ISSN 1751-7907 - p. 333 - 344.
bacillus-subtilis - comparative genomics - transcriptional regulation - negative regulator - escherichia-coli - ctsr - regulon - identification - discovery - bacteria
P>Gene regulatory networks can be reconstructed by combining transcriptome data from many different experiments to elucidate relations between the activity of certain transcription factors and the genes they control. To obtain insight in the regulatory network of Lactobacillus plantarum, microarray transcriptome data from more than 70 different experimental conditions were combined and the expression profiles of the transcriptional units (TUs) were compared. The TUs that displayed correlated expression were used to identify putative cis-regulatory elements by searching the upstream regions of the TUs for conserved motifs. Predicted motifs were extended and refined by searching for motifs in the upstream regions of additional TUs with correlated expression. In this way, cis-acting elements were identified for 41 regulons consisting of at least four TUs (correlation > 0.7). This set of regulons included the known regulons of CtsR and LexA, but also several novel ones encompassing genes with coherent biological functions. Visualization of the regulons and their connections revealed a highly interconnected regulatory network. This network contains several subnetworks that encompass genes of correlated biological function, such as sugar and energy metabolism, nitrogen metabolism and stress response.
Comparative functional genomics of amino acid metabolism of lactic acid bacteria
Pastink, M.I. - \ 2009
University. Promotor(en): Willem de Vos; J. Hugenholtz. - [S.l. : S.n. - ISBN 9789085854616 - 184
aminozuurmetabolisme - melkzuurbacteriën - streptococcus thermophilus - vergelijkende genomica - functionele genomica - amino acid metabolism - lactic acid bacteria - comparative genomics - functional genomics
The amino acid metabolism of lactic acid bacteria used as starters in industrial fermentations has profound effects on the quality of the fermented foods. The work described in this PhD thesis was initiated to use genomics technologies and a comparative approach to link the gene content of some well-known lactic acid bacteria to flavor formation and to increase our general knowledge in the area of amino acid metabolism. The three well-known lactic acid bacteria that were used in these studies were Streptococcus thermophilus, Lactococcus lactis and Lactobacillus plantarum. The complete genomes of all these model bacteria have been sequenced and annotated in detail. Comparative experimental and in silico studies of Streptococcus thermophilus with the other two lactic acid bacteria, revealed the low degree of amino acid auxotrophies of this species; it only needs two amino acids for (minimal) growth and this strain is able to produce a varied amount of flavors. Lactococcus lactis and Lactobacillus plantarum require more amino acids and produce fewer flavors than S. thermophilus. Furthermore, S. thermophilus has a simple primary metabolism; homolactic growth is the only possible route under anaerobic conditions and, remarkably, it does not have a complete pentose phosphate pathway in contrast to the other two studied bacteria. This latter property has important consequences for the redox metabolism of S. thermophilus and particularly its ability to produce NADPH. A genome-scale metabolic model was developed and predicted that amino acid metabolism, and especially glutamate degradation, and citrate metabolism are the most obvious alternatives for NADPH generation. Several of these predictions were confirmed by constructing a glutamate dehydrogenase mutant of S. thermophilus. This mutant revealed theimportance of the citrate pathway (and other amino acid degradation pathways) in NADPH generation. A comparative and functional genomics study of the three lactic acid bacteria showed that amino acid depletion not only affects amino acid metabolism, but also flavor formation and overall growth. The comparative genomics approach presented in this thesis can be used to understand the amino acid metabolism of different lactic acid bacteria and their potential to produce flavors under different conditions. Finally, it can be applied for optimization of industrial fermentations
Prokaryotic homologs of Argonaute proteins are predicted to function as key components of a novel system of defense against mobile genetic elements
Makarova, K.S. ; Wolf, Y.I. ; Oost, J. van der; Koonin, E.V. - \ 2009
Biology Direct 4 (2009). - ISSN 1745-6150 - p. 29 - 29.
messenger-rna targets - silencing complex - crystal-structure - structural basis - archaeoglobus-fulgidus - comparative genomics - histone deacetylase - aeolicus argonaute - regulatory rnas - slicer activity
ABSTRACT: BACKGROUND: In eukaryotes, RNA interference (RNAi) is a major mechanism of defense against viruses and transposable elements as well of regulating translation of endogenous mRNAs. The RNAi systems recognize the target RNA molecules via small guide RNAs that are completely or partially complementary to a region of the target. Key components of the RNAi systems are proteins of the Argonaute-PIWI family some of which function as slicers, the nucleases that cleave the target RNA that is base-paired to a guide RNA. Numerous prokaryotes possess the CRISPR-associated system (CASS) of defense against phages and plasmids that is, in part, mechanistically analogous but not homologous to eukaryotic RNAi systems. Many prokaryotes also encode homologs of Argonaute-PIWI proteins but their functions remain unknown. RESULTS: We present a detailed analysis of Argonaute-PIWI protein sequences and the genomic neighborhoods of the respective genes in prokaryotes. Whereas eukaryotic Ago/PIWI proteins always contain PAZ (oligonucleotide binding) and PIWI (active or inactivated nuclease) domains, the prokaryotic Argonaute homologs (pAgos) fall into two major groups in which the PAZ domain is either present or absent. The monophyly of each group is supported by a phylogenetic analysis of the conserved PIWI-domains. Almost all pAgos that lack a PAZ domain appear to be inactivated, and the respective genes are associated with a variety of predicted nucleases in putative operons. An additional, uncharacterized domain that is fused to various nucleases appears to be a unique signature of operons encoding the short (lacking PAZ) pAgo form. By contrast, almost all PAZ-domain containing pAgos are predicted to be active nucleases. Some proteins of this group (e.g., that from Aquifex aeolicus) have been experimentally shown to possess nuclease activity, and are not typically associated with genes for other (putative) nucleases. Given these observations, the apparent extensive horizontal transfer of pAgo genes, and their common, statistically significant over-representation in genomic neighborhoods enriched in genes encoding proteins involved in the defense against phages and/or plasmids, we hypothesize that pAgos are key components of a novel class of defense systems. The PAZ-domain containing pAgos are predicted to directly destroy virus or plasmid nucleic acids via their nuclease activity, whereas the apparently inactivated, PAZ-lacking pAgos could be structural subunits of protein complexes that contain, as active moieties, the putative nucleases that we predict to be co-expressed with these pAgos. All these nucleases are predicted to be DNA endonucleases, so it seems most probable that the putative novel phage/plasmid-defense system targets phage DNA rather than mRNAs. Given that in eukaryotic RNAi systems, the PAZ domain binds a guide RNA and positions it on the complementary region of the target, we further speculate that pAgos function on a similar principle (the guide being either DNA or RNA), and that the uncharacterized domain found in putative operons with the short forms of pAgos is a functional substitute for the PAZ domain. CONCLUSIONS: The hypothesis that pAgos are key components of a novel prokaryotic immune system that employs guide RNA or DNA molecules to degrade nucleic acids of invading mobile elements implies a functional analogy with the prokaryotic CASS and a direct evolutionary connection with eukaryotic RNAi. The predictions of the hypothesis including both the activities of pAgos and those of the associated endonucleases are readily amenable to experimental tests. Reviewers This article was reviewed by Daniel Haft, Martijn Huynen, and Chris Ponting
An in silico analysis of T-box regulated genes and T-box evolution in prokaryotes, with emphasis on prediction of substrate specificity of transporters
Wels, M.W.W. ; Kormelink, T.G. ; Kleerebezem, M. ; Siezen, R.J. ; Francke, C. - \ 2008
BMC Genomics 9 (2008). - ISSN 1471-2164 - 16 p.
transfer-rna-synthetase - gram-positive bacteria - subtilis-tyrs gene - directed transcription antitermination - multiple sequence alignment - bacillus-subtilis - amino-acids - corynebacterium-glutamicum - comparative genomics - glyqs gene
BACKGROUND: T-box anti-termination is an elegant and sensitive mechanism by which many bacteria maintain constant levels of amino acid-charged tRNAs. The amino acid specificity of the regulatory element is related to a so-called specifier codon and can in principle be used to guide the functional annotation of the genes controlled via the T-box anti-termination mechanism. RESULTS: Hidden Markov Models were defined to search the T-box regulatory element and were applied to all completed prokaryotic genomes. The vast majority of the genes found downstream of the retrieved elements encoded functionalities related to transport and synthesis of amino acids and the charging of tRNA. This is completely in line with findings reported in literature and with the proposed biological role of the regulatory element. For several species, the functional annotation of a large number of genes encoding proteins involved in amino acid transport could be improved significantly on basis of the amino acid specificity of the identified T-boxes. In addition, these annotations could be extrapolated to a larger number of orthologous systems in other species. Analysis of T-box distribution confirmed that the element is restricted predominantly to species of the phylum Firmicutes. Furthermore, it appeared that the distribution was highly species specific and that in the case of amino acid transport some boxes seemed to "pop-up" only recently. CONCLUSION: We have demonstrated that the identification of the molecular specificity of a regulatory element can be of great help in solving notoriously difficult annotation issues, e.g. by defining the substrate specificity of genes encoding amino acid transporters on basis of the amino acid specificity of the regulatory T-box. Furthermore, our analysis of the species-dependency of the occurrence of specific T-boxes indicated that these regulatory elements propagate in a semi-independent way from the genes that they control
The growth limits of a large number of Listeria monocytogenes strains at combinations of stresses show serotype- and niche-specific traits
Veen, S. van der; Moezelaar, R. ; Abee, T. ; Wells-Bennik, M.H.J. - \ 2008
Journal of Applied Microbiology 105 (2008)5. - ISSN 1364-5072 - p. 1246 - 1258.
osmolyte transporters betl - food-borne pathogen - comparative genomics - dairy-products - water activity - inoculum size - lactic-acid - heat-shock - scott-a - low ph
Aims: The aim of this study was to associate the growth limits of Listeria monocytogenes during exposure to combined stresses with specific serotypes or origins of isolation, and identify potential genetic markers. Methods and Results: The growth of 138 strains was assessed at different temperatures using combinations of low pH, sodium lactate, and high salt concentrations in brain heart infusion broth. None of the strains was able to grow at pH ¿ 4·4, aw ¿ 0·92, or pH ¿ 5·0 combined with aw ¿ 0·94. In addition, none of the strains grew at pH ¿ 5·2 and NaLac ¿ 2%. At 30°C, the serotype 4b strains showed the highest tolerance to low pH and high NaCl concentrations at both pH neutral (pH 7·4) and mild acidic conditions (pH 5·5). At 7°C, the serotype 1/2b strains showed the highest tolerance to high NaCl concentrations at both pH 7·4 and 5·5. Serotype 1/2b meat isolates showed the highest tolerance to low pH in the presence of 2% sodium lactate at 7°C. ORF2110 and gadD1T1 were identified as potential biomarkers for phenotypic differences. Conclusions: Differences in growth limits were identified between specific L. monocytogenes strains and serotypes, which could in some cases be associated with specific genetic markers. Significance and Impact of the Study: Our data confirm the growth limits of L. monocytogenes as set out by the European Union for ready-to-eat foods and provides an additional criterion. The association of L. monocytogenes serotypes with certain stress responses might explain the abundance of certain serotypes in retail foods while others are common in clinical cases.
Mapping QTLs for mineral accumulation and shoot dry biomass under different Zn nutritional conditions in Chinese cabbage ( Brassica rapa L. ssp. pekinensis )
Wu, J. ; Yuan, Y.X. ; Zhang, X.W. ; Zhao, J. ; Song, X. ; Li, Y. ; Li, X. ; Sun, R. ; Koornneef, M. ; Aarts, M.G.M. ; Wang, X.W. - \ 2008
Plant and Soil 310 (2008)1-2. - ISSN 0032-079X - p. 25 - 40.
quantitative trait loci - genetic-linkage map - length-polymorphism loci - arabidopsis-thaliana - natural variation - amplified polymorphism - comparative genomics - syn campestris - thlaspi-caerulescens - zinc accumulation
Abstract Chinese cabbage (Brassica rapa L. ssp. pekinensis) is one of the most important vegetables in China. Genetic dissection of leaf mineral accumulation and tolerance to Zn stress is important for the improvement of the nutritional quality of Chinese cabbage by breeding. A mapping population with 183 doubled haploid (DH) lines was used to study the genetics of mineral accumulation and the growth response to Zn. The genetic map was constructed based on 203 AFLPs, 58 SSRs, 22 SRAPs and four ESTPs. The concentration of 11 minerals was determined in leaves for 142 DH lines grown in an open field. In addition shoot dry biomass (SDB) under normal, deficient and excessive Zn nutritional conditions were investigated in hydroponics experiments. Ten QTLs, each explaining 11.1¿17.1% of the Na, Mg, P, Al, Fe, Mn, Zn and Sr concentration variance, were identified by multiple-QTL model (MQM) mapping. One common QTL was found affecting SDB under normal, deficient and excessive Zn nutritional conditions. An additional QTL was detected for SDB under Zn excess stress only. These results offer insights into the genetic basis of leaf mineral accumulation and plant growth under Zn stress conditions in Chinese cabbage
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