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- Hultling, C, et al.
(författare)
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Assisted ejaculation combined with in vitro fertilisation : an effective technique treating male infertility due to spinal cord injury.
- 1994
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Ingår i: Paraplegia. - : Springer Science and Business Media LLC. - 0031-1758. ; 32:7, s. 463-467
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Tidskriftsartikel (refereegranskat)abstract
- Infertility due to spinal cord injury (SCI) in males has been identified for decades as an area of major concern and techniques for assisted ejaculation are available. There has not been an overall consensus regarding which type of assisted procreation is the most appropriate for these couples. We describe here our experience from a programme based on assisted ejaculation combined with in vitro fertilization (IVF). Twelve couples have been treated so far and altogether 22 cycles with ovum pick-up have been completed. Fertilisation of the oocytes was obtained in 18 of these cycles. The overall oocyte fertilisation rate was 49%. Embryo transfer took place in 17 cycles, leading to seven clinical pregnancies. Four of the pregnancies are delivered or are ongoing, whereas three ended in first trimester spontaneous abortion. Thus our initial experience suggests that assisted ejaculation in combination with IVF is an effective option for these couples.
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| 5. |
- 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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- 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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- Cotman, Andrej Emanuel, et al.
(författare)
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Discovery and Hit-to-Lead Optimization of Benzothiazole Scaffold- Based DNA Gyrase Inhibitors with Potent Activity against Acinetobacter baumannii and Pseudomonas aeruginosa
- 2023
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Ingår i: Journal of Medicinal Chemistry. - : American Chemical Society (ACS). - 0022-2623 .- 1520-4804. ; 66:2, s. 1380-1425
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Tidskriftsartikel (refereegranskat)abstract
- We have developed compounds with a promising activity against Acinetobacter baumannii and Pseudomonas aerugi-nosa, which are both on the WHO priority list of antibiotic -resistant bacteria. Starting from DNA gyrase inhibitor 1, we identified compound 27, featuring a 10-fold improved aqueous solubility, a 10-fold improved inhibition of topoisomerase IV from A. baumannii and P. aeruginosa, a 10-fold decreased inhibition of human topoisomerase II alpha, and no cross-resistance to novobiocin. Cocrystal structures of 1 in complex with Escherichia coli GyrB24 and (S)-27 in complex with A. baumannii GyrB23 and P. aeruginosa GyrB24 revealed their binding to the ATP-binding pocket of the GyrB subunit. In further optimization steps, solubility, plasma free fraction, and other ADME properties of 27 were improved by fine-tuning of lipophilicity. In particular, analogs of 27 with retained anti-Gram-negative activity and improved plasma free fraction were identified. The series was found to be nongenotoxic, nonmutagenic, devoid of mitochondrial toxicity, and possessed no ion channel liabilities.
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| 11. |
- Garoff, Linnéa, et al.
(författare)
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Effect of aminoacyl-tRNA synthetase mutations on susceptibility to ciprofloxacin in Escherichia coli
- 2018
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press (OUP). - 0305-7453 .- 1460-2091. ; 73:12, s. 3285-3292
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Tidskriftsartikel (refereegranskat)abstract
- Background: Chromosomal mutations that reduce ciprofloxacin susceptibility in Escherichia coli characteristically map to drug target genes (gyrAB and parCE), and genes encoding regulators of the AcrAB-TolC efflux pump. Mutations in RNA polymerase can also reduce susceptibility, by up-regulating the MdtK efflux pump.Objectives: We asked whether mutations in additional chromosomal gene classes could reduce susceptibility to ciprofloxacin.Methods: Experimental evolution, complemented by WGS analysis, was used to select and identify mutations that reduce susceptibility to ciprofloxacin. Transcriptome analysis, genetic reconstructions, susceptibility measurements and competition assays were used to identify significant genes and explore the mechanism of resistance.Results: Mutations in three different aminoacyl-tRNA synthetase genes (leuS, aspS and thrS) were shown to re- duce susceptibility to ciprofloxacin. For two of the genes (leuS and aspS) the mechanism was partially dependent on RelA activity. Two independently selected mutations in leuS (Asp162Asn and Ser496Pro) were studied in most detail, revealing that they induce transcriptome changes similar to a stringent response, including up-regulation of three efflux-associated loci (mdtK, acrZ and ydhJK). Genetic analysis showed that reduced susceptibility depended on the activity of these loci. Broader antimicrobial susceptibility testing showed that the leuS mutations also reduce susceptibility to additional classes of antibiotics chloramphenicol, rifampicin, mecillinam, ampicillin and trimethoprim).Conclusions: The identification of mutations in multiple tRNA synthetase genes that reduce susceptibility to ciprofloxacin and other antibiotics reveals the existence of a large mutational target that could contribute to re- sistance development by up-regulation of an array of efflux pumps.
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| 12. |
- Garoff, Linnéa, et al.
(författare)
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Population Bottlenecks Strongly Influence the Evolutionary Trajectory to Fluoroquinolone Resistance in Escherichia coli
- 2020
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Ingår i: Molecular biology and evolution. - : Oxford University Press (OUP). - 0737-4038 .- 1537-1719. ; 37:6, s. 1637-1646
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Tidskriftsartikel (refereegranskat)abstract
- Experimental evolution is a powerful tool to study genetic trajectories to antibiotic resistance under selection. A confounding factor is that outcomes may be heavily influenced by the choice of experimental parameters. For practical purposes (minimizing culture volumes), most experimental evolution studies with bacteria use transmission bottleneck sizes of 5 x 10(6) cfu. We currently have a poor understanding of how the choice of transmission bottleneck size affects the accumulation of deleterious versus high-fitness mutations when resistance requires multiple mutations, and how this relates outcome to clinical resistance. We addressed this using experimental evolution of resistance to ciprofloxacin in Escherichia coli. Populations were passaged with three different transmission bottlenecks, including single cell (to maximize genetic drift) and bottlenecks spanning the reciprocal of the frequency of drug target mutations (10(8) and 10(10)). The 10(10) bottlenecks selected overwhelmingly mutations in drug target genes, and the resulting genotypes corresponded closely to those found in resistant clinical isolates. In contrast, both the 10(8) and single-cell bottlenecks selected mutations in three different gene classes: 1) drug targets, 2) efflux pump repressors, and 3) transcription-translation genes, including many mutations with low fitness. Accordingly, bottlenecks smaller than the average nucleotide substitution rate significantly altered the experimental outcome away from genotypes observed in resistant clinical isolates. These data could be applied in designing experimental evolution studies to increase their predictive power and to explore the interplay between different environmental conditions, where transmission bottlenecks might vary, and resulting evolutionary trajectories.
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| 14. |
- Huseby, Douglas L., et al.
(författare)
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Mutation supply and relative fitness shape the genotypes of ciprofloxacin-resistant Escherichia coli
- 2017
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Ingår i: Molecular biology and evolution. - : Oxford University Press (OUP). - 0737-4038 .- 1537-1719. ; 34:5, s. 1029-1039
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Tidskriftsartikel (refereegranskat)abstract
- Ciprofloxacin is an important antibacterial drug targeting Type II topoisomerases, highly active against Gram-negatives including Escherichia coli. The evolution of resistance to ciprofloxacin in E. coli always requires multiple genetic changes, usually including mutations affecting two different drug target genes, gyrA and parC. Resistant mutants selected in vitro or in vivo can have many different mutations in target genes and efflux regulator genes that contribute to resistance. Among resistant clinical isolates the genotype, gyrA S83L D87N, parC S80I is significantly overrepresented suggesting that it has a selective advantage. However, the evolutionary or functional significance of this high frequency resistance genotype is not fully understood. By combining experimental data and mathematical modeling, we addressed the reasons for the predominance of this specific genotype. The experimental data were used to model trajectories of mutational resistance evolution under different conditions of drug exposure and population bottlenecks. We identified the order in which specific mutations are selected in the clinical genotype, showed that the high frequency genotype could be selected over a range of drug selective pressures, and was strongly influenced by the relative fitness of alternative mutations and factors affecting mutation supply. Our data map for the first time the fitness landscape that constrains the evolutionary trajectories taken during the development of clinical resistance to ciprofloxacin and explain the predominance of the most frequently selected genotype. This study provides strong support for the use of in vitro competition assays as a tool to trace evolutionary trajectories, not only in the antibiotic resistance field.
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| 19. |
- 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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