| 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
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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
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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
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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
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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
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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
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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
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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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