| 1. |
- Ellegaard, Kirsten Maren, et al.
(författare)
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Comparative Genomics of Wolbachia and the Bacterial Species Concept
- 2013
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Ingår i: PLOS Genetics. - : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 9:4, s. e1003381-
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Tidskriftsartikel (refereegranskat)abstract
- The importance of host-specialization to speciation processes in obligate host-associated bacteria is well known, as is also the ability of recombination to generate cohesion in bacterial populations. However, whether divergent strains of highly recombining intracellular bacteria, such as Wolbachia, can maintain their genetic distinctness when infecting the same host is not known. We first developed a protocol for the genome sequencing of uncultivable endosymbionts. Using this method, we have sequenced the complete genomes of the Wolbachia strains wHa and wNo, which occur as natural double infections in Drosophila simulans populations on the Seychelles and in New Caledonia. Taxonomically, wHa belong to supergroup A and wNo to supergroup B. A comparative genomics study including additional strains supported the supergroup classification scheme and revealed 24 and 33 group-specific genes, putatively involved in host-adaptation processes. Recombination frequencies were high for strains of the same supergroup despite different host-preference patterns, leading to genomic cohesion. The inferred recombination fragments for strains of different supergroups were of short sizes, and the genomes of the co-infecting Wolbachia strains wHa and wNo were not more similar to each other and did not share more genes than other A- and B-group strains that infect different hosts. We conclude that Wolbachia strains of supergroup A and B represent genetically distinct clades, and that strains of different supergroups can co-exist in the same arthropod host without converging into the same species. This suggests that the supergroups are irreversibly separated and that barriers other than host-specialization are able to maintain distinct clades in recombining endosymbiont populations. Acquiring a good knowledge of the barriers to genetic exchange in Wolbachia will advance our understanding of how endosymbiont communities are constructed from vertically and horizontally transmitted genes.
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| 2. |
- Ellegaard, Kirsten Maren, et al.
(författare)
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Testing the Reproducibility of Multiple Displacement Amplification on Genomes of Clonal Endosymbiont Populations
- 2013
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:11, s. e82319-
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Tidskriftsartikel (refereegranskat)abstract
- The multiple displacement amplification method has revolutionized genomic studies of uncultured bacteria, where the extraction of pure DNA in sufficient quantity for next-generation sequencing is challenging. However, the method is problematic in that it amplifies the target DNA unevenly, induces the formation of chimeric reads and also amplifies contaminating DNA. Here, we have tested the reproducibility of the multiple displacement amplification method using serial dilutions of extracted genomic DNA and intact cells from the cultured endosymbiont Bartonella australis. The amplified DNA was sequenced with the Illumina sequencing technology, and the results were compared to sequence data obtained from unamplified DNA in this study as well as from a previously published genome project. We show that artifacts such as the extent of the amplification bias, the percentage of chimeric reads and the relative fraction of contaminating DNA increase dramatically for the smallest amounts of template DNA. The pattern of read coverage was reproducibly obtained for samples with higher amounts of template DNA, suggesting that the bias is non-random and genome-specific. A re-analysis of previously published sequence data obtained after amplification from clonal endosymbiont populations confirmed these predictions. We conclude that many of the artifacts associated with the use of the multiple displacement amplification method can be alleviated or much reduced by using multiple cells as the template for the amplification. These findings should be particularly useful for researchers studying the genomes of endosymbionts and other uncultured bacteria, for which a small clonal population of cells can be isolated.
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| 3. |
- Klasson, Lisa, et al.
(författare)
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Extensive duplication of the Wolbachia DNA in chromosome four of Drosophila ananassae
- 2014
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Ingår i: BMC Genomics. - : Springer Science and Business Media LLC. - 1471-2164. ; 15, s. 1097-
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Tidskriftsartikel (refereegranskat)abstract
- Background: Lateral gene transfer (LGT) from bacterial Wolbachia endosymbionts has been detected in similar to 20% of arthropod and nematode genome sequencing projects. Many of these transfers are large and contain a substantial part of the Wolbachia genome. Results: Here, we re-sequenced three D. ananassae genomes from Asia and the Pacific that contain large LGTs from Wolbachia. We find that multiple copies of the Wolbachia genome are transferred to the Drosophila nuclear genome in all three lines. In the D. ananassae line from Indonesia, the copies of Wolbachia DNA in the nuclear genome are nearly identical in size and sequence yielding an even coverage of mapped reads over the Wolbachia genome. In contrast, the D. ananassae lines from Hawaii and India show an uneven coverage of mapped reads over the Wolbachia genome suggesting that different parts of these LGTs are present in different copy numbers. In the Hawaii line, we find that this LGT is underrepresented in third instar larvae indicative of being heterochromatic. Fluorescence in situ hybridization of mitotic chromosomes confirms that the LGT in the Hawaii line is heterochromatic and represents similar to 20% of the sequence on chromosome 4 (dot chromosome, Muller element F). Conclusions: This collection of related lines contain large lateral gene transfers composed of multiple Wolbachia genomes that constitute >2% of the D. ananassae genome (similar to 5 Mbp) and partially explain the abnormally large size of chromosome 4 in D. ananassae.
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| 4. |
- Klasson, Lisa, et al.
(författare)
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Research on small genomes : Implications for synthetic biology
- 2010
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Ingår i: Bioessays. - : Wiley. - 0265-9247 .- 1521-1878. ; 32:4, s. 288-295
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Forskningsöversikt (refereegranskat)abstract
- Synthetic genomics is a new field of research in which small DNA pieces are assembled in a series of steps into whole genomes. The highly reduced genomes of host-associated bacteria are now being used as models for de novo synthesis of small genomes in the laboratory. Bacteria with the smallest genomes identified in nature provide nutrients to their hosts, such as amino acids, co-factors and vitamins. Comparative genomics of these bacteria enables predictions to be made about the gene sets required for core cellular functions and the associated metabolic network for the biosynthesis of host-selected compounds. Synthetic biology may ultimately enable researchers to make customized cell-specific organelles for the production and delivery of drugs to humans and domestic animals. Synthetic genomics may also become the method of choice for functional analyses of genes and genomes from bacteria that cannot be cultivated in the laboratory.
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| 5. |
- Nystedt, Björn, et al.
(författare)
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The Norway spruce genome sequence and conifer genome evolution
- 2013
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Ingår i: Nature. - : Nature Publishing Group. - 0028-0836 .- 1476-4687. ; 497:7451, s. 579-584
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Tidskriftsartikel (refereegranskat)abstract
- Conifers have dominated forests for more than 200 million years and are of huge ecological and economic importance. Here we present the draft assembly of the 20-gigabase genome of Norway spruce (Picea abies), the first available for any gymnosperm. The number of well-supported genes (28,354) is similar to the >100 times smaller genome of Arabidopsis thaliana, and there is no evidence of a recent whole-genome duplication in the gymnosperm lineage. Instead, the large genome size seems to result from the slow and steady accumulation of a diverse set of long-terminal repeat transposable elements, possibly owing to the lack of an efficient elimination mechanism. Comparative sequencing of Pinus sylvestris, Abies sibirica, Juniperus communis, Taxus baccata and Gnetum gnemon reveals that the transposable element diversity is shared among extant conifers. Expression of 24-nucleotide small RNAs, previously implicated in transposable element silencing, is tissue-specific and much lower than in other plants. We further identify numerous long (>10,000 base pairs) introns, gene-like fragments, uncharacterized long non-coding RNAs and short RNAs. This opens up new genomic avenues for conifer forestry and breeding.
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| 6. |
- Schneider, Daniela I., et al.
(författare)
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More than fishing in the dark : PCR of a dispersed sequence produces simple but ultrasensitive Wolbachia detection
- 2014
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Ingår i: BMC Microbiology. - : Springer Science and Business Media LLC. - 1471-2180. ; 14, s. 121-
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Tidskriftsartikel (refereegranskat)abstract
- Background: Detecting intracellular bacterial symbionts can be challenging when they persist at very low densities. Wolbachia, a widespread bacterial endosymbiont of invertebrates, is particularly challenging. Although it persists at high titers in many species, in others its densities are far below the detection limit of classic end-point Polymerase Chain Reaction (PCR). These low-titer infections can be reliably detected by combining PCR with DNA hybridization, but less elaborate strategies based on end-point PCR alone have proven less sensitive or less general. Results: We introduce a multicopy PCR target that allows fast and reliable detection of A-supergroup Wolbachia -even at low infection titers -with standard end-point PCR. The target is a multicopy motif (designated ARM: A-supergroup repeat motif) discovered in the genome of wMel (the Wolbachia in Drosophila melanogaster). ARM is found in at least seven other Wolbachia A-supergroup strains infecting various Drosophila, the wasp Muscidifurax and the tsetse fly Glossina. We demonstrate that end-point PCR targeting ARM can reliably detect both high-and low-titer Wolbachia infections in Drosophila, Glossina and interspecific hybrids. Conclusions: Simple end-point PCR of ARM facilitates detection of low-titer Wolbachia A-supergroup infections. Detecting these infections previously required more elaborate procedures. Our ARM target seems to be a general feature of Wolbachia A-supergroup genomes, unlike other multicopy markers such as insertion sequences (IS).
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| 7. |
- Siozios, Stefanos, et al.
(författare)
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The Diversity and Evolution of Wolbachia Ankyrin Repeat Domain Genes
- 2013
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:2, s. e55390-
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Tidskriftsartikel (refereegranskat)abstract
- Ankyrin repeat domain-encoding genes are common in the eukaryotic and viral domains of life, but they are rare in bacteria, the exception being a few obligate or facultative intracellular Proteobacteria species. Despite having a reduced genome, the arthropod strains of the alphaproteobacterium Wolbachia contain an unusually high number of ankyrin repeat domain-encoding genes ranging from 23 in wMel to 60 in wPip strain. This group of genes has attracted considerable attention for their astonishing large number as well as for the fact that ankyrin proteins are known to participate in protein-protein interactions, suggesting that they play a critical role in the molecular mechanism that determines host-Wolbachia symbiotic interactions. We present a comparative evolutionary analysis of the wMel-related ankyrin repeat domain-encoding genes present in different Drosophila-Wolbachia associations. Our results show that the ankyrin repeat domain-encoding genes change in size by expansion and contraction mediated by short directly repeated sequences. We provide examples of intragenic recombination events and show that these genes are likely to be horizontally transferred between strains with the aid of bacteriophages. These results confirm previous findings that the Wolbachia genomes are evolutionary mosaics and illustrate the potential that these bacteria have to generate diversity in proteins potentially involved in the symbiotic interactions.
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