| 1. |
- Bartke, Katrin, et al.
(författare)
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Genetic Architecture and Fitness of Bacterial Interspecies Hybrids
- 2021
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Ingår i: Molecular biology and evolution. - : Oxford University Press. - 0737-4038 .- 1537-1719. ; 38:4, s. 1472-1481
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Tidskriftsartikel (refereegranskat)abstract
- Integration of a conjugative plasmid into a bacterial chromosome can promote the transfer of chromosomal DNA to other bacteria. Intraspecies chromosomal conjugation is believed responsible for creating the global pathogens Klebsiella pneumoniae ST258 and Escherichia coli ST1193. Interspecies conjugation is also possible but little is known about the genetic architecture or fitness of such hybrids. To study this, we generated by conjugation 14 hybrids of E. coli and Salmonella enterica. These species belong to different genera, diverged from a common ancestor >100 Ma, and share a conserved order of orthologous genes with similar to 15% nucleotide divergence. Genomic analysis revealed that all but one hybrid had acquired a contiguous segment of donor E. coli DNA, replacing a homologous region of recipient Salmonella chromosome, and ranging in size from similar to 100 to >4,000 kb. Recombination joints occurred in sequences with higher-than-average nucleotide identity. Most hybrid strains suffered a large reduction in growth rate, but the magnitude of this cost did not correlate with the length of foreign DNA. Compensatory evolution to ameliorate the cost of low-fitness hybrids pointed towards disruption of complex genetic networks as a cause. Most interestingly, 4 of the 14 hybrids, in which from 45% to 90% of the Salmonella chromosome was replaced with E. coli DNA, showed no significant reduction in growth fitness. These data suggest that the barriers to creating high-fitness interspecies hybrids may be significantly lower than generally appreciated with implications for the creation of novel species.
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| 2. |
- Becker, K., et al.
(författare)
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Antibacterial activity of apramycin at acidic pH warrants wide therapeutic window in the treatment of complicated urinary tract infections and acute pyelonephritis
- 2021
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Ingår i: EBioMedicine. - : Elsevier B.V.. - 2352-3964. ; 73
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Tidskriftsartikel (refereegranskat)abstract
- Background: The clinical-stage drug candidate EBL-1003 (apramycin) represents a distinct new subclass of aminoglycoside antibiotics for the treatment of drug-resistant infections. It has demonstrated best-in-class coverage of resistant isolates, and preclinical efficacy in lung infection models. However, preclinical evidence for its utility in other disease indications has yet to be provided. Here we studied the therapeutic potential of EBL-1003 in the treatment of complicated urinary tract infection and acute pyelonephritis (cUTI/AP). Methods: A combination of data-base mining, antimicrobial susceptibility testing, time-kill experiments, and four murine infection models was used in a comprehensive assessment of the microbiological coverage and efficacy of EBL-1003 against Gram-negative uropathogens. The pharmacokinetics and renal toxicology of EBL-1003 in rats was studied to assess the therapeutic window of EBL-1003 in the treatment of cUTI/AP. Findings: EBL-1003 demonstrated broad-spectrum activity and rapid multi-log CFU reduction against a phenotypic variety of bacterial uropathogens including aminoglycoside-resistant clinical isolates. The basicity of amines in the apramycin molecule suggested a higher increase in positive charge at urinary pH when compared to gentamicin or amikacin, resulting in sustained drug uptake and bactericidal activity, and consequently in potent efficacy in mouse infection models. Renal pharmacokinetics, biomarkers for toxicity, and kidney histopathology in adult rats all indicated a significantly lower nephrotoxicity of EBL-1003 than of gentamicin. Interpretation: This study provides preclinical proof-of-concept for the efficacy of EBL-1003 in cUTI/AP. Similar efficacy but lower nephrotoxicity of EBL-1003 in comparison to gentamicin may thus translate into a higher safety margin and a wider therapeutic window in the treatment of cUTI/API. Funding: A full list of funding bodies that contributed to this study can be found in the Acknowledgements section. © 2021 The Author(s)
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| 3. |
- Berryhill, Brandon A., et al.
(författare)
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Evaluating the potential efficacy and limitations of a phage for joint antibiotic and phage therapy of Staphylococcus aureus infections
- 2021
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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 (PNAS). - 0027-8424 .- 1091-6490. ; 118:10
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Tidskriftsartikel (refereegranskat)abstract
- In response to increasing frequencies of antibiotic-resistant pathogens, there has been a resurrection of interest in the use of bacteriophage to treat bacterial infections: phage therapy. Here we explore the potential of a seemingly ideal phage, PYOSa, for combination phage and antibiotic treatment of Staphylococcus aureus infections. This K-like phage has a broad host range; all 83 tested clinical isolates of S.aureus tested were susceptible to PYOSa. Because of the mode of action of PYOSa, S. aureus is unlikely to generate classical receptor-site mutants resistant to PYOSa; none were observed in the 13 clinical isolates tested. PYOSa kills S. aureus at high rates. On the downside, the results of our experiments and tests of the joint action of PYOSa and antibiotics raise issues that must be addressed before PYOSa is employed clinically. Despite the maintenance of the phage, PYOSa does not clear populations of S. aureus. Due to the ascent of a phenotyically diverse array of small-colony variants following an initial demise, the bacterial populations return to densities similar to that of phage-free controls. Using a combination of mathematical modeling and in vitro experiments, we postulate and present evidence for a mechanism to account for the demise-resurrection dynamics of PYOSa and S. aureus. Critically for phage therapy, our experimental results suggest that treatment with PYOSa followed by bactericidal antibiotics can clear populations of S. aureus more effectively than the antibiotics alone.
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| 4. |
- Brandis, Gerrit, 1985-, et al.
(författare)
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Mutant RNA polymerase can reduce susceptibility to antibiotics via ppGpp-independent induction of a stringent-like response
- 2021
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press. - 0305-7453 .- 1460-2091. ; 76:3, s. 606-615
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundMutations in RNA polymerase (RNAP) can reduce susceptibility to ciprofloxacin in Escherichia coli, but the mechanism of transcriptional reprogramming responsible is unknown. Strains carrying ciprofloxacin-resistant (CipR) rpoB mutations have reduced growth fitness and their impact on clinical resistance development is unclear.ObjectivesTo assess the potential for CipRrpoB mutations to contribute to resistance development by estimating the number of distinct alleles. To identify fitness-compensatory mutations that ameliorate the fitness costs of CipRrpoB mutations. To understand how CipRrpoB mutations reprogramme RNAP.MethodsE. coli strains carrying five different CipRrpoB alleles were evolved with selection for improved fitness and characterized for acquired mutations, relative fitness and MICCip. The effects of dksA mutations and a ppGpp0 background on growth and susceptibility phenotypes associated with CipRrpoB alleles were determined.ResultsThe number of distinct CipRrpoB mutations was estimated to be >100. Mutations in RNAP genes and in dksA can compensate for the fitness cost of CipRrpoB mutations. Deletion of dksA reduced the MICCip for strains carrying CipRrpoB alleles. A ppGpp0 phenotype had no effect on drug susceptibility.ConclusionsCipRrpoB mutations induce an ppGpp-independent stringent-like response. Approximately half of the reduction in ciprofloxacin susceptibility is caused by an increased affinity of RNAP to DksA while the other half is independent of DksA. Stringent-like response activating mutations might be the most diverse class of mutations reducing susceptibility to antibiotics.
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| 5. |
- Praski Alzrigat, Lisa, et al.
(författare)
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Resistance/fitness trade-off is a barrier to the evolution of MarR inactivation mutants in Escherichia coli
- 2021
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press. - 0305-7453 .- 1460-2091. ; 76:1, s. 77-83
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundMutations that inactivate MarR reduce susceptibility to ciprofloxacin and competitive growth fitness in Escherichia coli. Both phenotypes are caused by overexpression of the MarA regulon, which includes the AcrAB-TolC drug efflux pump.ObjectivesWe asked whether compensatory evolution could reduce the fitness cost of MarR-inactivating mutations without affecting resistance to ciprofloxacin.MethodsThe cost of overexpressing the AcrAB-TolC efflux pump was measured independently of MarA overexpression. Experimental evolution of MarR-inactive strains was used to select mutants with increased fitness. The acquired mutations were identified and their effects on drug susceptibility were measured.ResultsOverexpression of the AcrAB-TolC efflux pump was found not to contribute to the fitness cost of MarA regulon overexpression. Fitness-compensatory mutations were selected in marA and lon. The mutations reduced the level of MarA protein thus reducing expression of the MarA regulon. They restored growth fitness but also reduced resistance to ciprofloxacin.ConclusionsThe fitness cost caused by overexpression of the MarA regulon has multiple contributing factors. Experimental evolution did not identify any single pump-independent cost factor. Instead, efficient fitness compensation occurred only by mechanisms that reduce MarA concentration, which simultaneously reduce the drug resistance phenotype. This resistance/fitness trade-off is a barrier to the successful spread of MarR inactivation mutations in clinical isolates where growth fitness is essential.
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| 6. |
- Yadav, Kavita, et al.
(författare)
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Phenotypic and genetic barriers to establishment of horizontally transferred genes encoding ribosomal protection proteins
- 2021
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press. - 0305-7453 .- 1460-2091. ; 76:6, s. 1441-1447
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Tidskriftsartikel (refereegranskat)abstract
- Background: Ribosomal protection proteins (RPPs) interact with bacterial ribosomes to prevent inhibition of protein synthesis by tetracycline. RPP genes have evolved from a common ancestor into at least 12 distinct classes and spread by horizontal genetic transfer into a wide range of bacteria. Many bacterial genera host RPP genes from multiple classes but tet(M) is the predominant RPP gene found in Escherichia coli. Objectives: We asked whether phenotypic barriers (low-level resistance, high fitness cost) might constrain the fixation of other RPP genes in E. coli. Methods: We expressed a diverse set of six different RPP genes in E. coli, including tet(M), and quantified tetracycline susceptibility and growth phenotypes as a function of expression level, and evolvability to overcome identified phenotypic barriers. Results: The genes tet(M) and tet(Q) conferred high-level tetracycline resistance without reducing fitness; tet(O) and tet(W) conferred high-level resistance but significantly reduced growth fitness; tetB(P) conferred low-level resistance and while mutants conferring high-level resistance were selectable these had reduced growth fitness; otr(A) did not confer resistance and resistant mutants could not be selected. Evolution experiments suggested that codon usage patterns in tet(O) and tet(W), and transcriptional silencing associated with nucleotide composition in tetB(P), accounted for the observed phenotypic barriers. Conclusions: With the exception of tet(Q), the data reveal significant phenotypic and genetic barriers to the fixation of additional RPP genes in E. coli.
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