| 1. |
- B. Moreno, Anaísa
(författare)
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Evolution and host-specific adaptations of Legionella pneumophila
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- How bacteria evolve pathogenic traits is shaped by their communities and environments. Legionella pneumophila is ubiquitous in aquatic habitats, where it persists by replicating within a broad range of protozoan hosts. Using the same mechanisms, L. pneumophila may also accidentally infect humans, causing a severe pneumonia known as Legionnaires’ disease. As hosts, humans are evolutionary dead-ends, resulting in the loss of human-specific adaptations after infection. This thesis aims to identify and characterise these host adaptations.In Paper I, we study the in-patient evolution of L. pneumophila. We collected a large set of strains from sporadic infections and outbreaks, pairing clinical isolates with their respective environmental sources. Using comparative genomic analyses, we identified two genes individually mutated in three independent infections. One gene encoded an outer membrane protein, a homolog from the OmpP1/FadL family, and the other an EAL domain-containing protein. These results suggest that convergent evolution may be at play and that these mutations are potential candidates for human-specific host adaptations.In Paper II, we investigate host adaptation and the selective pressures that drive it using a long-term experimental evolution approach. We passaged L. pneumophila in Acanthamoeba castellanii and U937 macrophages, separately and in alternation, for over 800 generations. We found 49 fixed mutations across the 18 evolved populations: two distinct mutations in RpsL, which confers streptomycin resistance, as well as two additional mutations, each consistently associated with one of the former, in the chaperonin GroES or in RpsD, a known compensatory mutation. Mutations in the lipopolysaccharide synthesis operon were observed only in lineages passaged in A. castellanii, whilst mutations in LerC were fixed in six lineages passaged in U937, making these candidate mutations for host-specific adaptations.In Paper III, we shift focus to A. castellanii, a natural host of L. pneumophila. We describe a novel method for high-efficiency transfection of this amoeba with a cationic polymer. Using a systematic approach to test different parameters, we found that widely available and inexpensive polyethylenimines can be used to transfect A. castellanii at a much greater efficiency than the currently used reagents.In conclusion, these studies suggest that although L. pneumophila can infect humans, it is sub-optimally adapted for it, and offer potential determinants of host-specificity in L. pneumophila.
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| 2. |
- Brandis, Gerrit, 1985-, et al.
(författare)
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Expression of the qepA1 gene is induced under antibiotic exposure
- 2021
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press. - 0305-7453 .- 1460-2091. ; 76:6, s. 1433-1440
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundThe qepA1 gene encodes an efflux pump that reduces susceptibility to ciprofloxacin. Little is known about the regulation of qepA1 expression.ObjectivesTo assess the potential role of ciprofloxacin and other antibiotics in the regulation of qepA1 gene expression. To identify the promoter that drives qepA1 expression and other factors involved in expression regulation. To assess whether the identified features are universal among qepA alleles.MethodsA translational qepA1-yfp fusion under the control of the qepA1 upstream region was cloned into the Escherichia coli chromosome. Expression of the fusion protein was measured in the presence of various antibiotics. Deletions within the upstream region were introduced to identify regions involved in gene expression and regulation. The qepA1 coding sequence and upstream region were compared with all available qepA sequences.ResultsCellular stress caused by the presence of various antibiotics can induce qepA1 expression. The qepA1 gene is fused to a class I integron and gene expression is driven by the Pc promoter within the integrase gene. A segment within the integron belonging to a truncated dfrB4 gene is essential for the regulation of qepA1 expression. This genetic context is universal among all sequenced qepA alleles.ConclusionsThe fusion of the qepA1 gene to a class I integron has created a novel regulatory unit that enables qepA1 expression to be under the control of antibiotic exposure. This setup mitigates potential negative effects of QepA1 production on bacterial fitness by restricting high-level expression to environmental conditions in which QepA1 is beneficial.
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| 3. |
- Fer, Evrim, et al.
(författare)
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Early divergence of translation initiation and elongation factors
- 2022
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Ingår i: Protein Science. - : John Wiley & Sons. - 0961-8368 .- 1469-896X. ; 31:9
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Tidskriftsartikel (refereegranskat)abstract
- Protein translation is a foundational attribute of all living cells. The translation function carried out by the ribosome critically depends on an assortment of protein interaction partners, collectively referred to as the translation machinery. Various studies suggest that the diversification of the translation machinery occurred prior to the last universal common ancestor, yet it is unclear whether the predecessors of the extant translation machinery factors were functionally distinct from their modern counterparts. Here we reconstructed the shared ancestral trajectory and subsequent evolution of essential translation factor GTPases, elongation factor EF-Tu (aEF-1A/eEF-1A), and initiation factor IF2 (aIF5B/eIF5B). Based upon their similar functions and structural homologies, it has been proposed that EF-Tu and IF2 emerged from an ancient common ancestor. We generated the phylogenetic tree of IF2 and EF-Tu proteins and reconstructed ancestral sequences corresponding to the deepest nodes in their shared evolutionary history, including the last common IF2 and EF-Tu ancestor. By identifying the residue and domain substitutions, as well as structural changes along the phylogenetic history, we developed an evolutionary scenario for the origins, divergence and functional refinement of EF-Tu and IF2 proteins. Our analyses suggest that the common ancestor of IF2 and EF-Tu was an IF2-like GTPase protein. Given the central importance of the translation machinery to all cellular life, its earliest evolutionary constraints and trajectories are key to characterizing the universal constraints and capabilities of cellular evolution.
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| 4. |
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| 5. |
- Garmendia, Eva, et al.
(författare)
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Chromosomal Location Determines the Rate of Intrachromosomal Homologous Recombination in Salmonella
- 2021
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Ingår i: mBio. - : American Society for Microbiology. - 2161-2129 .- 2150-7511. ; 12:3
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Tidskriftsartikel (refereegranskat)abstract
- Homologous recombination is an important mechanism directly involved in the repair, organization, and evolution of prokaryotic and eukaryotic chromosomes. We developed a system, based on two genetic cassettes, that allows the measurement of recombinational repair rates between different locations on the chromosome. Using this system, we analyzed 81 different positional combinations throughout the chromosome to answer the question of how the position and orientation of sequences affect intrachromosomal homologous recombination. Our results show that recombination was possible between any two locations tested in this study and that recombinational repair rates varied by just above an order of magnitude. The observed differences in rate do not correlate with distance between the recombination cassettes or with distance from the origin of replication but could be explained if each location contributes individually to the recombination event. The relative levels of accessibility for recombination vary 5-fold between the various cassette locations, and we found that the nucleoid structure of the chromosome may be the major factor influencing the recombinational accessibility of each chromosomal site. Furthermore, we found that the orientation of the recombination cassettes had a significant impact on recombination. Recombinational repair rates for the cassettes inserted as direct repeats are, on average, 2.2-fold higher than those for the same sets inserted as inverted repeats. These results suggest that the bacterial chromosome is not homogenous with regard to homologous recombination, with regions that are more or less accessible, and that the orientation of genes affects recombination rates. IMPORTANCE Bacterial chromosomes frequently carry multiple copies of genes at separate chromosomal locations. In Salmonella, these include the 7 rrn operons and the duplicate tuf genes. Genes within these families coevolve by homologous recombination, but it is not obvious whether their rates of recombination reflect general rates of intrachromosomal recombination or are an evolved property particularly associated with these conserved genes and locations. Using a novel experimental system, we show that recombination is possible between all tested pairs of locations at rates that vary by just above 1 order of magnitude. Differences in rate do not correlate with distance between the sites or distance to the origin of replication but may be explained if each location contributes individually to the recombination event. Our results suggest the existence of bacterial chromosomal domains that are differentially available for recombination and that gene orientation affects recombination rates.
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| 6. |
- Graells, Tiscar, et al.
(författare)
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Legionella pneumophila recurrently isolated in a Spanish hospital : Two years of antimicrobial resistance surveillance
- 2018
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Ingår i: Environmental Research. - : Elsevier. - 0013-9351 .- 1096-0953. ; 166, s. 638-646
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVES: The aim of this study was to monitor the spread, persistence and antibiotic resistance patterns of Legionella spp. strains found in a hospital water distribution system. These environmental studies are intended to help detect the presence of antibiotic resistant strains before they infect patients.METHODS: Antimicrobial surveillance tests were performed at 27 different sampling points of the water network of a large Spanish hospital over two years. Water samples were screened for Legionella according to ISO 11731:2007. Legionella spp. isolates were identified by serotyping and by mass spectrometry (MALDI-ToF). Epidemiological molecular typing was done by Pulse-Field Gel Electrophoresis (PFGE) and by Sequence-Based Typing (SBT). Antibiotic susceptibility tests were performed using disk diffusion and ETEST®.RESULTS: Legionella spp. were recurrently isolated for 2 years. All isolates belonged the same group, L. pneumophila serogroups 2-14. Isolates were all attributed by SBT to sequence type (ST) ST328, although PFGE revealed 5 different patterns. No significant change in antibiotic susceptibility could be observed for this study period, irrespectively of the method used.CONCLUSION: Colonization of water systems by Legionella spp. is still occurring, although all the prevention rules were strictly followed. Antibiotic resistance monitoring may help us to find resistance in bacteria with environmental reservoirs but difficult to isolate from patients. The knowledge of the antibiotic susceptibility in environmental strains may help us to predict changes in clinical strains. This study might also help reconsidering Legionnaires' disease (LD) diagnostic methods. L. pneumophila serogroups 2-14 present all along the time of the investigation in the water distribution system can cause LD. However, they may not be detected by routine urine tests run on patients, thereby missing an ongoing LD infection.
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| 7. |
- Graells, Tiscar, et al.
(författare)
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The all-intracellular order Legionellales is unexpectedly diverse, globally distributed and lowly abundant.
- 2018
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Ingår i: FEMS Microbiology Ecology. - : Oxford University Press. - 0168-6496 .- 1574-6941. ; 94:12
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Tidskriftsartikel (refereegranskat)abstract
- Legionellales is an order of the Gammaproteobacteria, only composed of host-adapted, intracellular bacteria, including the accidental human pathogens Legionella pneumophila and Coxiella burnetii. Although the diversity in terms of lifestyle is large across the order, only a few genera have been sequenced, owing to the difficulty to grow intracellular bacteria in pure culture. In particular, we know little about their global distribution and abundance.Here, we analyze 16/18S rDNA amplicons both from tens of thousands of published studies and from two separate sampling campaigns in and around ponds and in a silver mine. We demonstrate that the diversity of the order is much larger than previously thought, with over 450 uncultured genera. We show that Legionellales are found in about half of the samples from freshwater, soil and marine environments, and quasi-ubiquitous in man-made environments. Their abundance is low, typically 0.1%, with few samples up to 1%. Most Legionellales OTUs are globally distributed, while many do not belong to a previously identified species.This study sheds a new light on the ubiquity and diversity of one major group of host-adapted bacteria. It also emphasizes the need to use metagenomics to better understand the role of host-adapted bacteria in all environments.
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| 8. |
- Guy, Lionel, 1980-, et al.
(författare)
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genoPlotR : comparative gene and genome visualization in R
- 2010
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Ingår i: Bioinformatics. - : Oxford University Press (OUP). - 1367-4803 .- 1367-4811. ; 26:18, s. 2334-2335
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Tidskriftsartikel (refereegranskat)abstract
- The amount of gene and genome data obtained by next-generation sequencing technologies generates a need for comparative visualization tools. Complementing existing software for comparison and exploration of genomics data, genoPlotR automatically creates publication-grade linear maps of gene and genomes, in a highly automatic, flexible and reproducible way. Availability: genoPlotR is a platform-independent R package, available with full source code under a GPL2 license at R-Forge: http://genoplotr.r-forge.r-project.org/ Contact: lionel.guy@ebc.uu.se
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| 9. |
- Guy, Lionel, 1980-
(författare)
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phyloSkeleton : taxon selection, data retrieval and marker identification for phylogenomics
- 2017
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Ingår i: Bioinformatics. - : Oxford University Press (OUP). - 1367-4803 .- 1367-4811. ; 33:8, s. 1230-1232
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Tidskriftsartikel (refereegranskat)abstract
- With the wealth of available genome sequences, a difficult and tedious part of inferring phylogenomic trees is now to select genomes with an appropriate taxon density in the different parts of the tree. The package described here offers tools to easily select the most representative organisms, following a set of simple rules based on taxonomy and assembly quality, to retrieve the genomes from public databases (NCBI, JGI), to annotate them if necessary, to identify given markers in these, and to prepare files for multiple sequence alignment.AVAILABILITY AND IMPLEMENTATION: phyloSkeleton is a Perl module and is freely available under GPLv3 at https://bitbucket.org/lionelguy/phyloskeleton/ CONTACT: lionel.guy@imbim.uu.se.
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| 10. |
- Guy, Lionel, 1980-, et al.
(författare)
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The archaeal ‘TACK’ superphylum and the origin of eukaryotes
- 2011
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Ingår i: Trends in Microbiology. - : Elsevier BV. - 0966-842X .- 1878-4380. ; 19:12, s. 580-587
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Forskningsöversikt (refereegranskat)abstract
- Although most hypotheses to explain the emergence of the eukaryotic lineage are conflicting, some consensus exists concerning the requirement of a genomic fusion between archaeal and bacterial components. Recent phylogenomic studies have provided support for eocyte-like scenarios in which the alleged ‘archaeal parent’ of the eukaryotic cell emerged from the Crenarchaeota/Thaumarchaeota. Here, we provide evidence for a scenario in which this archaeal parent emerged from within the ‘TACK’ superphylum that comprises the Thaumarchaeota, Crenarchaeota and Korarchaeota, as well as the recently proposed phylum ‘Aigarchaeota’. In support of this view, functional and comparative genomics studies have unearthed an increasing number of features that are uniquely shared by the TACK superphylum and eukaryotes, including proteins involved in cytokinesis, membrane remodeling, cell shape determination and protein recycling.
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