| 1. |
- Wistrand-Yuen, Erik, 1980-, et al.
(författare)
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Evolution of high-level resistance during low-level antibiotic exposure
- 2018
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 9:1
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Tidskriftsartikel (refereegranskat)abstract
- It has become increasingly clear that low levels of antibiotics present in many environments can select for resistant bacteria, yet the evolutionary pathways for resistance development during exposure to low amounts of antibiotics remain poorly defined. Here we show that Salmonella enterica exposed to sub-MIC levels of streptomycin evolved high-level resistance via novel mechanisms that are different from those observed during lethal selections. During lethal selection only rpsL mutations are found, whereas at sub-MIC selection resistance is generated by several small-effect resistance mutations that combined confer high-level resistance via three different mechanisms: (i) alteration of the ribosomal RNA target (gidB mutations), (ii) reduction in aminoglycoside uptake (cyoB, nuoG, and trkH mutations), and (iii) induction of the aminoglycoside-modifying enzyme AadA (znuA mutations). These results demonstrate how the strength of the selective pressure influences evolutionary trajectories and that even weak selective pressures can cause evolution of high-level resistance.
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| 2. |
- Amlinger, Lina, et al.
(författare)
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Fluorescent CRISPR Adaptation Reporter for rapid quantification of spacer acquisition
- 2017
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Ingår i: Scientific Reports. - : NATURE PUBLISHING GROUP. - 2045-2322. ; 7
-
Tidskriftsartikel (refereegranskat)abstract
- CRISPR-Cas systems are adaptive prokaryotic immune systems protecting against horizontally transferred DNA or RNA such as viruses and other mobile genetic elements. Memory of past invaders is stored as spacers in CRISPR loci in a process called adaptation. Here we developed a novel assay where spacer integration results in fluorescence, enabling detection of memory formation in single cells and quantification of as few as 0.05% cells with expanded CRISPR arrays in a bacterial population. Using this fluorescent CRISPR Adaptation Reporter (f-CAR), we quantified adaptation of the two CRISPR arrays of the type I-E CRISPR-Cas system in Escherichia coli, and confirmed that more integration events are targeted to CRISPR-II than to CRISPR-I. The f-CAR conveniently analyzes and compares many samples, allowing new insights into adaptation. For instance, we show that in an E. coli culture the majority of acquisition events occur in late exponential phase.
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| 3. |
- Baldanzi, Gabriel, et al.
(författare)
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OSA Is Associated With the Human Gut Microbiota Composition and Functional Potential in the Population-Based Swedish CardioPulmonary bioImage Study
- 2023
-
Ingår i: Chest. - : Elsevier. - 0012-3692 .- 1931-3543. ; 164:2, s. 503-516
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Obstructive sleep apnea (OSA) is a common sleep-breathing disorder linked to increased risk of cardiovascular disease. Intermittent hypoxia and intermittent airway obstruction, hallmarks of OSA, have been shown in animal models to induce substantial changes to the gut microbiota composition and subsequent transplantation of fecal matter to other animals induced changes in blood pressure and glucose metabolism.RESEARCH QUESTION: Does obstructive sleep apnea in adults associate with the composition and metabolic potential of the human gut microbiota?STUDY DESIGN AND METHODS: We used respiratory polygraphy data from up to 3,570 individuals aged 50-64 from the population-based Swedish CardioPulmonary bioImage Study combined with deep shotgun metagenomics of fecal samples to identify cross-sectional associations between three OSA parameters covering apneas and hypopneas, cumulative sleep time in hypoxia and number of oxygen desaturation events with gut microbiota composition. Data collection about potential confounders was based on questionnaires, on-site anthropometric measurements, plasma metabolomics, and linkage with the Swedish Prescribed Drug Register.RESULTS: We found that all three OSA parameters were associated with lower diversity of species in the gut. Further, the OSA-related hypoxia parameters were in multivariable-adjusted analysis associated with the relative abundance of 128 gut bacterial species, including higher abundance of Blautia obeum and Collinsela aerofaciens. The latter species was also independently associated with increased systolic blood pressure. Further, the cumulative time in hypoxia during sleep was associated with the abundance of genes involved in nine gut microbiota metabolic pathways, including propionate production from lactate. Lastly, we observed two heterogeneous sets of plasma metabolites with opposite association with species positively and negatively associated with hypoxia parameters, respectively.INTERPRETATION: OSA-related hypoxia, but not the number of apneas/hypopneas, is associated with specific gut microbiota species and functions. Our findings lay the foundation for future research on the gut microbiota-mediated health effects of OSA.
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| 4. |
- Ghosh, Anirban, et al.
(författare)
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Contact-dependent growth inhibition induces high levels of antibiotic-tolerant persister cells in clonal bacterial populations
- 2018
-
Ingår i: EMBO Journal. - : WILEY. - 0261-4189 .- 1460-2075. ; 37:9
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial populations can use bet-hedging strategies to cope with rapidly changing environments. One example is non-growing cells in clonal bacterial populations that are able to persist antibiotic treatment. Previous studies suggest that persisters arise in bacterial populations either stochastically through variation in levels of global signalling molecules between individual cells, or in response to various stresses. Here, we show that toxins used in contact-dependent growth inhibition (CDI) create persisters upon direct contact with cells lacking sufficient levels of CdiI immunity protein, which would otherwise bind to and neutralize toxin activity. CDI-mediated persisters form through a feedforward cycle where the toxic activity of the CdiA toxin increases cellular (p)ppGpp levels, which results in Lon-mediated degradation of the immunity protein and more free toxin. Thus, CDI systems mediate a population density-dependent bet-hedging strategy, where the fraction of non-growing cells is increased only when there are many cells of the same genotype. This may be one of the mechanisms of how CDI systems increase the fitness of their hosts.
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| 5. |
- Jones, Allison M., et al.
(författare)
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Genetic Evidence for SecY Translocon-Mediated Import of Two Contact-Dependent Growth Inhibition (CDI) Toxins
- 2021
-
Ingår i: mBio. - : American Society for Microbiology. - 2161-2129 .- 2150-7511. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- The C-terminal (CT) toxin domains of contact-dependent growth inhibition (CDI) CdiA proteins target Gram-negative bacteria and must breach both the outer and inner membranes of target cells to exert growth inhibitory activity. Here, we examine two CdiA-CT toxins that exploit the bacterial general protein secretion machinery after delivery into the periplasm. A Ser281Phe amino acid substitution in transmembrane segment 7 of SecY, the universally conserved channel-forming subunit of the Sec translocon, decreases the cytotoxicity of the membrane depolarizing orphan10 toxin from enterohemorrhagic Escherichia coli EC869. Target cells expressing secY(S281F) and lacking either PpiD or YfgM, two SecY auxiliary factors, are fully protected from CDI-mediated inhibition either by CdiA-CTo10EC869 or by CdiA-CTGN05224, the latter being an EndoU RNase CdiA toxin from Klebsiella aerogenes GN05224 that has a related cytoplasm entry domain. RNase activity of CdiA-CTGN05224 was reduced in secY(S281F) target cells and absent in secY(S281F) Delta ppiD or secY(S281F) Delta yfgM target cells during competition co-cultures. Importantly, an allele-specific mutation in secY (secY(G313W)) renders DppiD or Delta yfgM target cells specifically resistant to CdiA-CTGN05224 but not to CdiA-CTo10EC869, further suggesting a direct interaction between SecY and the CDI toxins. Our results provide genetic evidence of a unique confluence between the primary cellular export route for unfolded polypeptides and the import pathways of two CDI toxins. IMPORTANCE Many bacterial species interact via direct cell-to-cell contact using CDI systems, which provide a mechanism to inject toxins that inhibit bacterial growth into one another. Here, we find that two CDI toxins, one that depolarizes membranes and another that degrades RNA, exploit the universally conserved SecY translocon machinery used to export proteins for target cell entry. Mutations in genes coding for members of the Sec translocon render cells resistant to these CDI toxins by blocking their movement into and through target cell membranes. This work lays the foundation for understanding how CDI toxins interact with the protein export machinery and has direct relevance to development of new antibiotics that can penetrate bacterial cell envelopes.
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| 6. |
- Kjellin, Jonas, et al.
(författare)
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Colicins and T6SS-based competition systems enhance enterotoxigenic E. coli (ETEC) competitiveness
- 2024
-
Ingår i: Gut microbes. - : Taylor & Francis. - 1949-0976 .- 1949-0984. ; 16:1
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Tidskriftsartikel (refereegranskat)abstract
- Diarrheal diseases are still a significant problem for humankind, causing approximately half a million deaths annually. To cause diarrhea, enteric bacterial pathogens must first colonize the gut, which is a niche occupied by the normal bacterial microbiota. Therefore, the ability of pathogenic bacteria to inhibit the growth of other bacteria can facilitate the colonization process. Although enterotoxigenic Escherichia coli (ETEC) is one of the major causative agents of diarrheal diseases, little is known about the competition systems found in and used by ETEC and how they contribute to the ability of ETEC to colonize a host. Here, we collected a set of 94 fully assembled ETEC genomes by performing whole-genome sequencing and mining the NCBI RefSeq database. Using this set, we performed a comprehensive search for delivered bacterial toxins and investi-gated how these toxins contribute to ETEC competitiveness in vitro. We found that type VI secretion systems (T6SS) were widespread among ETEC (n = 47). In addition, several closely related ETEC strains were found to encode Colicin Ia and T6SS (n = 8). These toxins provide ETEC compe-titive advantages during in vitro competition against other E. coli, suggesting that the role of T6SS as well as colicins in ETEC biology has until now been underappreciated.
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| 7. |
- Koskiniemi, Sanna, 1980-
(författare)
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Dynamics of the Bacterial Genome : Rates and Mechanisms of Mutation
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Bacterial chromosomes are highly dynamic, continuously changing with respect to gene content and size via a number of processes, including deletions that result in gene loss. How deletions form and at what rates has been the focus of this thesis. In paper II we investigated how chromosomal location affects chromosomal deletion rates in S. typhimurium. Deletion rates varied more than 100-fold between different chromosomal locations and some large deletions significantly increased the exponential growth rate of the cells. Our results suggest that the chromosome is heterogeneous with respect to deletion rates and that deletions may be genetically fixed as a consequence of natural selection rather than by drift or mutational biases. In paper I we examined in a laboratory setting how rapidly reductive evolution, i.e. gene loss, could occur. Using a serial passage approach, we showed that extensive genome reduction potentially could occur on a very short evolutionary time scale. For most deletions we observed little or no homology at the deletion endpoints, indicating that spontaneous deletions often form through a RecA independent process. In paper III we examined further how large spontaneous deletions form and, unexpectedly, showed that 90% of all spontaneous chromosomal deletions required error-prone translesion DNA polymerases for their formation. We propose that the translesion polymerases stimulate deletion formation by allowing extension of misaligned single-strand DNA ends. In paper IV we investigated how the translesion DNA polymerase Pol IV, RpoS and different types of stresses affect mutation rates in bacteria. Derepression of the LexA regulon caused a small to moderate increase in mutation rates that was fully dependent on functional endonucleases but only partly dependent on translesion DNA polymerases. RpoS levels and growth stresses had only minor effects on mutation rates. Thus, mutation rates appear very robust and are only weakly affected by growth conditions and induction of translesion polymerases and RpoS.
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| 8. |
- Koskiniemi, Sanna, 1980-, et al.
(författare)
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Effect of the translesion DNA polymerases, endonucleases and RpoS on mutation rates in Salmonella typhimurium
- 2010
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Ingår i: Genetics. - : Oxford University Press (OUP). - 0016-6731 .- 1943-2631. ; 185:3, s. 783-795
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Tidskriftsartikel (refereegranskat)abstract
- It has been suggested that bacteria have evolved mechanisms to increase their mutation rate in response to various stresses and that the translesion DNA polymerase Pol IV under control of the LexA regulon and the alternative sigma factor RpoS are involved in regulating this mutagenesis. Here we examined in Salmonella enterica serovar Typhimurium LT2 the rates for four different types of mutations (rifampicin-, nalidixic acid- and chlorate-resistance and Lac+ reversion) during various growth conditions and with different levels of four translesion DNA polymerases (Pol II, Pol IV, Pol V and SamAB) and RpoS. Constitutive de-repression of the LexA regulon by a lexA(def) mutation increased mutation rates 1.5- to 12-fold and the contribution of the translesion DNA polymerases to this mutagenesis varied with the type of mutation examined. In contrast, for all four types of mutations examined the increase in mutation rate in the lexA(def) mutant required the presence of the LexA-controlled endonucleases UvrB, UvrC and Cho. With regard to the potential involvement of RpoS in mutagenesis, neither an increase in RpoS levels conferred by artificial over-expression from a plasmid nor long-term stationary phase incubation or slow growth caused an increase in any of the four mutation rates measured, alone or in combination with over-expression of the translesion DNA polymerases. In conclusion, mutation rates are remarkably robust and no combination of growth conditions, induction of translesion polymerases by inactivation of LexA or increased RpoS expression could confer an increase in mutation rates higher than the moderate increase caused by de-repression of the LexA regulon alone.
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| 9. |
- Koskiniemi, Sanna, 1980-, et al.
(författare)
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Translesion DNA polymerases are required for spontaneous deletion formation in Salmonella typhimurium
- 2009
-
Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 106:25, s. 10248-10253
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Tidskriftsartikel (refereegranskat)abstract
- How spontaneous deletions form in bacteria is still a partly unresolved problem. Here we show that deletion formation in S. typhimurium requries the presence of functional translesion polymerases. First, in wild type bacteria, removal of the known translesion DNA polymerases: PolII (polB), PolIV (dinB), PolV (umuDC) and the PolV homologue SamAB (samAB) resulted in a 10-fold decrease in the deletion rate, indicating that 90% of all spontaneous deletions require these polymerases for their formation. Second, overexpression of these polymerases by de-repression of the DNA damage-inducible LexA regulon caused a 25-fold increase in deletion rate that depended on the presence of functional translesion polymerases. Third, overexpression of the polymerases PolII and PolIV from a plasmid increased the deletion rate 12- to 30-fold respectively. Last, in a recBC- mutant where dsDNA ends are stabilized due to the lack of the end-processing nuclease RecBC, the deletion rate was increased 20-fold. This increase depended on the translesion polymerases. In lexA(def) mutant cells with constitutive SOS-expression, a 10-fold increase in DNA breaks was observed. Inactivation of all 4 translesion polymerases in the lexA(def) mutant reduced the deletion rate 250-fold without any concomitant reduction in the amount of DNA breaks. Mutational inactivation of 3 endonucleases under LexA control, reduced the number of DNA breaks to the wild-type level in a lexA(def) mutant with a concomitant 50-fold reduction in deletion rate. These findings suggest that the translesion polymerases are not involved in forming the DNA breaks, but that they require them to stimulate deletion formation.
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| 10. |
- Koskiniemi, Sanna, 1980-, et al.
(författare)
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Variation in spontaneous deletion rates at different locations of the Salmonella typhimurium chromosome
-
Annan publikation (populärvet., debatt m.m.)abstract
- How and at what rates spontaneous deletions form is still a partly unresolved question. Here we have constructed a genetic tool that can be used to determine spontaneous chromosomal deletion rates at any chromosomal location. We measured deletion rates at 12 chromosomal locations and identified the deletable region as the largest deletion found at each location. Our data shows that spontaneous deletion rates can at least vary 100-fold between the different chromosomal locations when normalized to the size of the deletable region. The isolated deletions ranged in size from 1-200 kbp and the highest deletion rates were found around 2 Mbp of the S. typhiumurium chromosome, suggesting a potential hotspot for deletion formation. No long repeat sequences were found in this region that could explain the high deletion rate. Furthermore, no obvious correlation between fitness (measured as exponential growth rate) and deletion size could be seen. Surprisingly, since deletions are commonly considered deleterious certain deletions (ranging from 18- to 38 kbp in size) increased the growth rate of the cells with ~5% in both rich and poor growth media. These results suggest that the bacterial chromosome is heterogeneous with respect to the rate of deletion formation and that some deletions could become fixed as a consequence of natural selection rather than by drift and/or mutational biases.
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| 11. |
- Lee, Danna
(författare)
-
To defend or to offend? : Bacterial defence and competition systems
- 2022
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Bacteria are prokaryotic microorganisms that can be found in all niches of the biosphere. Bacteria must constantly evolve in order to survive and adapt to their environments. Defence systems and competition systems play a vital role in ensuring the survival of bacteria. The projects included in this thesis aim to further understand how these systems function, as well as adapting them in order to design probiotic strains with built-in biocontainment mechanisms. In paper I, we investigate how internal expression of Rearrangement hotspot (Rhs) toxins allows protection against Rhs toxin delivery in S. Typhimurium. In paper II, we examine bacterial competition systems found in enterotoxigenic E. coli (ETEC) strains and explore the evolution of toxin arsenals within ETEC family lineages. In paper III, we develop a fluorescence flow cytometry-based high-throughput screening method and apply it to isolate a highly competitive commensal E. coli strain that possesses a diverse arsenal of competition systems. In paper IV, we design and construct a synthetic CRISPR system that is able to protect E. coli from acquisition of antibiotic resistance genes through horizontal gene transfer. The work presented in this thesis contribute to our understanding of the functions of bacterial defence and competition systems, at the same time, laying the groundwork for how these systems can be studied in a high-throughput manner as well as how they can be adapted in the future to design synthetic microbial strains with clinical and environmental applications.
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| 12. |
- Michalska, Karolina, et al.
(författare)
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Functional plasticity of antibacterial EndoU toxins
- 2018
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Ingår i: Molecular Microbiology. - : WILEY. - 0950-382X .- 1365-2958. ; 109:4, s. 509-527
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Tidskriftsartikel (refereegranskat)abstract
- Bacteria use several different secretion systems to deliver toxic EndoU ribonucleases into neighboring cells. Here, we present the first structure of a prokaryotic EndoU toxin in complex with its cognate immunity protein. The contact‐dependent growth inhibition toxin CdiA‐CTSTECO31 from Escherichia coli STEC_O31 adopts the eukaryotic EndoU fold and shares greatest structural homology with the nuclease domain of coronavirus Nsp15. The toxin contains a canonical His‐His‐Lys catalytic triad in the same arrangement as eukaryotic EndoU domains, but lacks the uridylate‐specific ribonuclease activity that characterizes the superfamily. Comparative sequence analysis indicates that bacterial EndoU domains segregate into at least three major clades based on structural variations in the N‐terminal subdomain. Representative EndoU nucleases from clades I and II degrade tRNA molecules with little specificity. In contrast, CdiA‐CTSTECO31 and other clade III toxins are specific anticodon nucleases that cleave tRNAGlu between nucleotides C37 and m2A38. These findings suggest that the EndoU fold is a versatile scaffold for the evolution of novel substrate specificities. Such functional plasticity may account for the widespread use of EndoU effectors by diverse inter‐bacterial toxin delivery systems.
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| 13. |
- Ruhe, Zachary C., et al.
(författare)
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CdiA Effectors Use Modular Receptor-Binding Domains To Recognize Target Bacteria
- 2017
-
Ingår i: mBio. - : AMER SOC MICROBIOLOGY. - 2161-2129 .- 2150-7511. ; 8:2
-
Tidskriftsartikel (refereegranskat)abstract
- Contact-dependent growth inhibition (CDI) systems encode CdiA effectors, which bind to specific receptors on neighboring bacteria and deliver C-terminal toxin domains to suppress target cell growth. Two classes of CdiA effectors that bind distinct cell surface receptors have been identified, but the molecular basis of receptor specificity is not understood. Alignment of BamA-specific CdiAEC93 from Escherichia coli EC93 and OmpC-specific CdiA(EC536) from E. coli 536 suggests that the receptor-binding domain resides within a central region that varies between the two effectors. In support of this hypothesis, we find that CdiA(EC93) fragments containing residues Arg1358 to Phe1646 bind specifically to purified BamA. Moreover, chimeric CdiA(EC93) that carries the corresponding sequence from CdiA(EC536) is endowed with OmpC-binding activity, demonstrating that this region dictates receptor specificity. A survey of E. coli CdiA proteins reveals two additional effector classes, which presumably recognize distinct receptors. Using a genetic approach, we identify the outer membrane nucleoside transporter Tsx as the receptor for a third class of CdiA effectors. Thus, CDI systems exploit multiple outer membrane proteins to identify and engage target cells. These results underscore the modularity of CdiA proteins and suggest that novel effectors can be constructed through genetic recombination to interchange different receptor-binding domains and toxic payloads. IMPORTANCE CdiB/CdiA two-partner secretion proteins mediate interbacterial competition through the delivery of polymorphic toxin domains. This process, known as contact-dependent growth inhibition (CDI), requires stable interactions between the CdiA effector protein and specific receptors on the surface of target bacteria. Here, we localize the receptor-binding domain to the central region of E. coli CdiA. Receptor-binding domains vary between CdiA proteins, and E. coli strains collectively encode at least four distinct effector classes. Further, we show that receptor specificity can be altered by exchanging receptor-binding regions, demonstrating the modularity of this domain. We propose that novel CdiA effectors are naturally generated through genetic recombination to interchange different receptor-binding domains and toxin payloads.
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| 14. |
- Stenum, Thomas Søndergaard, et al.
(författare)
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RNA interactome capture in Escherichia coli globally identifies RNA-binding proteins
- 2023
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Ingår i: Nucleic Acids Research. - : Oxford University Press. - 0305-1048 .- 1362-4962. ; 51:9, s. 4572-4587
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Tidskriftsartikel (refereegranskat)abstract
- RNA-binding proteins (RPBs) are deeply involved in fundamental cellular processes in bacteria and are vital for their survival. Despite this, few studies have so far been dedicated to direct and global identification of bacterial RBPs. We have adapted the RNA interactome capture (RIC) technique, originally developed for eukaryotic systems, to globally identify RBPs in bacteria. RIC takes advantage of the base pairing potential of poly(A) tails to pull-down RNA-protein complexes. Overexpressing poly(A) polymerase I in Escherichia coli drastically increased transcriptome-wide RNA polyadenylation, enabling pull-down of crosslinked RNA-protein complexes using immobilized oligo(dT) as bait. With this approach, we identified 169 putative RBPs, roughly half of which are already annotated as RNA-binding. We experimentally verified the RNA-binding ability of a number of uncharacterized RBPs, including YhgF, which is exceptionally well conserved not only in bacteria, but also in archaea and eukaryotes. We identified YhgF RNA targets in vivo using CLIP-seq, verified specific binding in vitro, and reveal a putative role for YhgF in regulation of gene expression. Our findings present a simple and robust strategy for RBP identification in bacteria, provide a resource of new bacterial RBPs, and lay the foundation for further studies of the highly conserved RBP YhgF.
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| 15. |
- Stårsta, Magnus, et al.
(författare)
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RHS-elements function as type II toxin-antitoxin modules that regulate intra-macrophage replication of Salmonella Typhimurium
- 2020
-
Ingår i: PLOS Genetics. - : PUBLIC LIBRARY SCIENCE. - 1553-7390 .- 1553-7404. ; 16:2
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Tidskriftsartikel (refereegranskat)abstract
- RHS elements are components of conserved toxin-delivery systems, wide-spread within the bacterial kingdom and some of the most positively selected genes known. However, very little is known about how Rhs toxins affect bacterial biology. Salmonella Typhimurium contains a full-length rhs gene and an adjacent orphan rhs gene, which lacks the conserved delivery part of the Rhs protein. Here we show that, in addition to the conventional delivery, Rhs toxin-antitoxin pairs encode for functional type-II toxin-antitoxin (TA) loci that regulate S. Typhimurium proliferation within macrophages. Mutant S. Typhimurium cells lacking both Rhs toxins proliferate 2-times better within macrophages, mainly because of an increased growth rate. Thus, in addition to providing strong positive selection for the rhs loci under conditions when there is little or no toxin delivery, internal expression of the toxin-antitoxin system regulates growth in the stressful environment found inside macrophages.
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| 16. |
- Virtanen, Petra, et al.
(författare)
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Class II contact‐dependent growth inhibition (CDI) systems allow for broad‐range cross‐species toxin delivery within the Enterobacteriaceae family
- 2019
-
Ingår i: Molecular Microbiology. - : Wiley. - 0950-382X .- 1365-2958. ; 111:4, s. 1109-1125
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Tidskriftsartikel (refereegranskat)abstract
- Contact‐dependent growth inhibition (CDI) allows bacteria to recognize kin cells in mixed bacterial populations. In Escherichia coli, CDI mediated effector delivery has been shown to be species‐specific, with a preference for the own strain over others. This specificity is achieved through an interaction between a receptor‐binding domain in the CdiA protein and its cognate receptor protein on the target cell. But how conserved this specificity is has not previously been investigated in detail. Here, we show that class II CdiA receptor‐binding domains and their Enterobacter cloacae analog are highly promiscuous, and can allow for efficient effector delivery into several different Enterobacteriaceae species, including Escherichia, Enterobacter, Klebsiella and Salmonella spp. In addition, although we observe a preference for the own receptors over others for two of the receptor‐binding domains, this did not limit cross‐species effector delivery in all experimental conditions. These results suggest that class II CdiA proteins could allow for broad‐range and cross‐species growth inhibition in mixed bacterial populations.
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| 17. |
- Wäneskog, Marcus, et al.
(författare)
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Escherichia coli EC93 deploys two plasmid- encoded class I contact- dependent growth inhibition systems for antagonistic bacterial interactions
- 2021
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Ingår i: Microbial Genomics. - : MICROBIOLOGY SOC. - 2057-5858. ; 7:3
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Tidskriftsartikel (refereegranskat)abstract
- The phenomenon of contact- dependent growth inhibition (CDI) and the genes required for CDI (cdiBAI) were identified and isolated in 2005 from an Escherichia coli isolate (EC93) from rats. Although the cdiBAIEC93 locus has been the focus of extensive research during the past 15 years, little is known about the EC93 isolate from which it originates. Here we sequenced the EC93 genome and find two complete and functional cdiBAI loci (including the previously identified cdi locus), both carried on a large 127 kb plasmid. These cdiBAI systems are differentially expressed in laboratory media, enabling EC93 to outcompete E. coli cells lacking cognate cdiI immunity genes. The two CDI systems deliver distinct effector peptides that each dissipate the membrane potential of target cells, although the two toxins display different toxic potencies. Despite the differential expression and toxic potencies of these CDI systems, both yielded similar competitive advantages against E. coli cells lacking immunity. This can be explained by the fact that the less expressed cdiBAI system (cdiBAIEC93-2) delivers a more potent toxin than the highly expressed cdiBAIEC93-1 system. Moreover, our results indicate that unlike most sequenced CDI+ bacterial isolates, the two cdi loci of E. coli EC93 are located on a plasmid and are expressed in laboratory media.
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| 18. |
- Wäneskog, Marcus
(författare)
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Too close for comfort : The role of Contact-Dependent growth Inhibition (CDI) in interbacterial competition and cooperation
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Contact-Dependent growth inhibition (CDI) was discovered in 2005 in the E. coli isolate EC93. Since then our knowledge of CDI systems and their impact on bacterial communities have increased exponentially. Yet many biological aspects of CDI systems are still unknown and their impact on complex microbial communities have only just begun to be studied. CDI systems require the function of three proteins; CdiBAI. The outer-membrane transport protein, CdiB, allows for the transportation of the toxin delivery protein CdiA to the cell surface of an inhibitor cell. Through a contact- and receptor-dependent interaction with a target cell the toxic C-terminal domain of CdiA is cleaved off and delivered into the target cell were it mediates a growth arrest. Different CdiA-CT domains encodes for diverse toxic activities, such as nucleases and membrane ionophore toxins. Each unique CdiA-CT toxin has a cognate immunity protein (CdiI) that binds and neutralize against its toxic activity, thus preventing a possible self-inhibition.In this thesis I have studied the effect of CDI system(s) on both single cell and population level, within both intra- and interspecies bacterial communities. The findings presented here shows that multiple class I cdiBAI loci within a cell can function in a synergetic manner and act as versatile interbacterial warfare systems able to inhibit the growth of rival bacteria, even when CdiA expression is low. We also show that class II CdiA receptor-binding domains can mediate broad-range cross-species toxin delivery and growth inhibition, even when a non-optimal target cell receptor is expressed at a low level. Additionally, we show that the cdiA gene supports the expression of two separate proteins. The full-length CdiA protein, able to mediate an extracellular toxin delivery, but also the discrete CdiA-CT toxin domain. This stand-alone CdiA-CT expression was stress-dependent and together with its cognate CdiI immunity protein functioned as a selfish-genetic element. Moreover, we show that CDI systems can increase bacterial stress tolerance via an extracellular toxin delivery between kin-cells. This stress tolerance phenotype only occurred under conditions when we also observed a selective degradation of the CdiI immunity protein. Therefore, we suggest that a selective CdiI degradation allows for a sub-population of cells to self-intoxicate, thereby becoming transiently dormant, which confers an increase in stress tolerance. The findings presented in this thesis collectively suggest that CDI systems could function as a pseudo-quorum sensing system able to mediate behavioral changes and stress tolerance within a sub-population of cells in a bacterial community.
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| 19. |
- Xu, Xingxing, et al.
(författare)
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All-electrical antibiotic susceptibility testing within 30 min using silicon nano transistors
- 2022
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Ingår i: Sensors and actuators. B, Chemical. - : Elsevier. - 0925-4005 .- 1873-3077. ; 357
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Tidskriftsartikel (refereegranskat)abstract
- Rapid and reliable antibiotic susceptibility testing (AST) platform is highly desired to select the right antibiotics to treat infectious disease at early stage. Here, we demonstrate rapid ASTs using nanoscale silicon ion-selective field-effect transistor sensors. Our sensors profile bacterial metabolic kinetics by monitoring the metabolism induced acidification in the growth media with the absence and the presence of different antibiotics. Rapid AST results could be determined from the metabolic profiles with a total assay time less than 30 min for different bacterial strains. In addition, the sensors could also distinguish the bactericidal mechanisms for antibiotics with different modes of actions. Furthermore, the initial bacterial concentration in an unknown sample, a key parameter to determine its clinic relevance, could be estimated based on the metabolic profiles. Our demonstrated AST method is all-electrical, label-free and silicon technology compatible, and holds great promise for the development of a high-throughput and low-cost point-of-care device.
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