| 1. |
- Nilsson, Annika I., et al.
(författare)
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Reducing the fitness cost of antibiotic resistance by amplification of initiator tRNA genes
- 2006
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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. ; 103:18, s. 6976-6981
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Tidskriftsartikel (refereegranskat)abstract
- Deformylase inhibitors belong to a novel antibiotic class that targets peptide deformylase, a bacterial enzyme that removes the formyl group from N-terminal methionine in nascent polypeptides. Using the bacterium Salmonella enterica, we isolated mutants with resistance toward the peptide deformylase inhibitor actinonin. Resistance mutations were identified in two genes that are required for the formylation of methionyl (Met) initiator tRNA (tRNAi)(fMet): the fmt gene encoding the enzyme methionyl-tRNA formyltransferase and the folD gene encoding the bifunctional enzyme methylenetetrahydrofolate-dehydrogenase and -cyclohydrolase. In the absence of antibiotic, these resistance mutations conferred a fitness cost that was manifested as a reduced growth rate in laboratory medium and in mice. By serially passaging the low-fitness mutants in growth medium without antibiotic, the fitness costs could be partly ameliorated either by intragenic mutations in the fmt/folD genes or by extragenic compensatory mutations. Of the extragenically compensated fmt mutants, approximately one-third carried amplifications of the identical, tandemly repeated metZ and metW genes, encoding tRNAi. The increase in metZW gene copy number varied from 5- to 40-fold and was accompanied by a similar increase in tRNAi levels. The rise in tRNAi level compensated for the lack of methionyl-tRNA formyltransferase activity and allowed translation initiation to proceed with nonformylated methionyl tRNAi. Amplified units varied in size from 1.9 to 94 kbp. Suppression of deleterious mutations by gene amplification may be involved in the evolution of new gene functions.
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| 2. |
- Cars, Otto, et al.
(författare)
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Resetting the agenda for antibiotic resistance through a health systems perspective
- 2021
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Ingår i: The Lancet Global Health. - : Elsevier. - 2214-109X. ; 9:7, s. E1022-E1027
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Forskningsöversikt (refereegranskat)abstract
- Although the individual and societal consequences of antibiotic resistance spiral upwards, coordinated action has not kept pace on a global scale. The COVID-19 pandemic has highlighted the need for resilient health systems and has resulted in an unprecedented rate of collaboration in scientific, medical, social, and political dimensions. The pandemic has also created a renewed awareness of the importance of infectious diseases and is a substantial entry point for reigniting the momentum towards containing the silent pandemic of antibiotic resistance. In this Viewpoint, we discuss the limitations in the current narrative on antibiotic resistance and how it could be improved, including concerted efforts to close essential data gaps. We discuss the need for capacity building and coordination at the national and global levels to strengthen the understanding of the importance of sustainable access to effective antibiotics for all health systems that could generate tangible links to current processes for global health and development.
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| 3. |
- Dittrich, Sabine, et al.
(författare)
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Target Product Profile for a Diagnostic Assay to Differentiate between Bacterial and Non-Bacterial Infections and Reduce Antimicrobial Overuse in Resource-Limited Settings : An Expert Consensus
- 2016
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 11:8
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Tidskriftsartikel (refereegranskat)abstract
- Acute fever is one of the most common presenting symptoms globally. In order to reduce the empiric use of antimicrobial drugs and improve outcomes, it is essential to improve diagnostic capabilities. In the absence of microbiology facilities in low-income settings, an assay to distinguish bacterial from non-bacterial causes would be a critical first step. To ensure that patient and market needs are met, the requirements of such a test should be specified in a target product profile (TPP). To identify minimal/optimal characteristics for a bacterial vs. non-bacterial fever test, experts from academia and international organizations with expertise in infectious diseases, diagnostic test development, laboratory medicine, global health, and health economics were convened. Proposed TPPs were reviewed by this working group, and consensus characteristics were defined. The working group defined non-severely ill, non-malaria infected children as the target population for the desired assay. To provide access to the most patients, the test should be deployable to community health centers and informal health settings, and staff should require <2 days of training to perform the assay. Further, given that the aim is to reduce inappropriate antimicrobial use as well as to deliver appropriate treatment for patients with bacterial infections, the group agreed on minimal diagnostic performance requirements of >90% and >80% for sensitivity and specificity, respectively. Other key characteristics, to account for the challenging environment at which the test is targeted, included: i) time-to-result <10 min (but maximally <2 hrs); ii) storage conditions at 0-40 degrees C, <= 90% non-condensing humidity with a minimal shelf life of 12 months; iii) operational conditions of 5-40 degrees C, <= 90% non-condensing humidity; and iv) minimal sample collection needs (50-100 mu L, capillary blood). This expert approach to define assay requirements for a bacterial vs. non-bacterial assay should guide product development, and enable targeted and timely efforts by industry partners and academic institutions.
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| 4. |
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| 5. |
- Högberg, Liselotte Diaz, et al.
(författare)
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Antibiotic use worldwide
- 2014
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Ingår i: The Lancet - Infectious diseases. - 1473-3099 .- 1474-4457. ; 14:12, s. 1179-1180
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Tidskriftsartikel (refereegranskat)
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| 6. |
- Jasovsky, Dusan, et al.
(författare)
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Antimicrobial resistance-a threat to the world's sustainable development
- 2016
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Ingår i: Upsala Journal of Medical Sciences. - : Uppsala Medical Society. - 0300-9734 .- 2000-1967. ; 121:3, s. 159-164
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Forskningsöversikt (refereegranskat)abstract
- This commentary examines how specific sustainable development goals (SDGs) are affected by antimicrobial resistance and suggests how the issue can be better integrated into international policy processes. Moving beyond the importance of effective antibiotics for the treatment of acute infections and health care generally, we discuss how antimicrobial resistance also impacts on environmental, social, and economic targets in the SDG framework. The paper stresses the need for greater international collaboration and accountability distribution, and suggests steps towards a broader engagement of countries and United Nations agencies to foster global intersectoral action on antimicrobial resistance.
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| 7. |
- Ke, Rongqin, et al.
(författare)
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Colorimetric Nucleic Acid Testing Assay for RNA Virus Detection Based on Circle-to-Circle Amplification of Padlock Probes
- 2011
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Ingår i: Journal of Clinical Microbiology. - 0095-1137 .- 1098-660X. ; 49:12, s. 4279-4285
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Tidskriftsartikel (refereegranskat)abstract
- We developed a molecular diagnostic method for detection of RNA virus based on padlock probes and colorimetric readout. The feasibility of our approach was demonstrated by using detection of Crimean-Congo hemorrhagic fever (CCHF) virus as a model. Compared with conventional PCR-based methods, our approach does not require advanced equipment, involves easier assay design, and has a sensitivity of 103 viral copies/ml. By using a cocktail of padlock probes, synthetic templates representing different viral strain variants could be detected. We analyzed 34 CCHF patient samples, and all patients were correctly diagnosed when the results were compared to those of the current real-time PCR method. This is the first time that highly specific padlock probes have been applied to detection of a highly variable target sequence typical of RNA viruses.
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| 8. |
- Mezger, Anja, et al.
(författare)
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A General Method for Rapid Determination of Antibiotic Susceptibility and Species in Bacterial Infections
- 2015
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Ingår i: Journal of Clinical Microbiology. - 0095-1137 .- 1098-660X. ; 53:2, s. 425-432
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Tidskriftsartikel (refereegranskat)abstract
- To ensure correct antibiotic treatment and reduce the unnecessary use of antibiotics, there is an urgent need for new rapid methods for species identification and determination of antibiotic susceptibility in infectious pathogenic bacteria. We have developed a general method for the rapid identification of the bacterial species causing an infection and the determination of their antibiotic susceptibility profiles. An initial short cultivation step in the absence and presence of different antibiotics was combined with sensitive species-specific padlock probe detection of the bacterial target DNA to allow a determination of growth (i.e., resistance) and no growth (i.e., susceptibility). A proof-of-concept was established for urinary tract infections in which we applied the method to determine the antibiotic susceptibility profiles of Escherichia coli for two drugs with 100% accuracy in 3.5 h. The short assay time from sample to readout enables fast appropriate treatment with effective drugs and minimizes the need to prescribe broad-spectrum antibiotics due to unknown resistance profiles of the treated infection.
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| 9. |
- Pavlov, Michael, et al.
(författare)
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Initiation factor 2 mutants promoting fast joining of ribosomal subunits in the absence of initiator tRNA or GTP
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- We have previously identified several mutations in initiation factor 2 (IF2) located outside the tRNA binding domain IV of IF2 that compensate for lack of initiator tRNA (Met-tRNAi) formylation in vivo. We have also shown that these IF2 mutants promote fast joining of ribosomal subunits even when non-formylated Met-tRNAi or deacylated tRNAi was present in the 30S pre-initiation complex (30S PIC) instead of formylated fMet-tRNAi. We demonstrate here in vitro that these IF2 mutants do not require any tRNA present on the 30S subunit to promote fast subunit joining provided that GTP is present. Moreover, in the presence of fMet-tRNAi the mutants promote fast subunit joining in the presence of only GDP. Thus, A-type IF2 mutants require either GTP or fMet-tRNAi for fast subunit joining. In contrast, fast subunit joining with wild type IF2 requires the presence of both GTP and fMet-tRNAi in the 30S PIC. These results imply that the presence of tRNA on the 30S subunit is per se not required for fast subunit joining but rather for switching the 30S:IF2 complex containing wild type IF2 into its 50S docking conformation. We show also that the rate of subunit joining with A-type IF2 is much less sensitive to the energy level in the reaction mixture than the rate of subunit joining with wild type IF2. We speculate that this insensitivity of initiation to energy levels may result in deteriorated adaptation of formylation-proficient strains harbouring A-type mutations in IF2 to growth under energy-limited conditions.
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| 10. |
- Pavlov, Michael Y., et al.
(författare)
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Activation of initiation factor 2 by ligands and mutations for rapid docking of ribosomal subunits
- 2011
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Ingår i: EMBO Journal. - : Wiley. - 0261-4189 .- 1460-2075. ; 30:2, s. 289-301
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Tidskriftsartikel (refereegranskat)abstract
- We previously identified mutations in the GTPase initiation factor 2 (IF2), located outside its tRNA-binding domain, compensating strongly (A-type) or weakly (B-type) for initiator tRNA formylation deficiency. We show here that rapid docking of 30S with 50S subunits in initiation of translation depends on switching 30S subunit-bound IF2 from its inactive to active form. Activation of wild-type IF2 requires GTP and formylated initiator tRNA (fMet-tRNA(i)). In contrast, extensive activation of A-type IF2 occurs with only GTP or with GDP and fMet-tRNA(i), implying a passive role for initiator tRNA as activator of IF2 in subunit docking. The theory of conditional switching of GTPases quantitatively accounts for all our experimental data. We find that GTP, GDP, fMet-tRNA(i) and A-type mutations multiplicatively increase the equilibrium ratio, K, between active and inactive forms of IF2 from a value of 4 × 10(-4) for wild-type apo-IF2 by factors of 300, 8, 80 and 20, respectively. Functional characterization of the A-type mutations provides keys to structural interpretation of conditional switching of IF2 and other multidomain GTPases.
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| 11. |
- Pietsch, Franziska, et al.
(författare)
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Ciprofloxacin selects for RNA polymerase mutations with pleiotropic antibiotic resistance effects
- 2017
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press. - 0305-7453 .- 1460-2091. ; 72:1, s. 75-84
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Tidskriftsartikel (refereegranskat)abstract
- ObjectivesResistance to the fluoroquinolone drug ciprofloxacin is commonly linked to mutations that alter the drug target or increase drug efflux via the major AcrAB-TolC transporter. Very little is known about other mutations that might also reduce susceptibility to ciprofloxacin. We discovered that an Escherichia coli strain experimentally evolved for resistance to ciprofloxacin had acquired a mutation in rpoB, the gene coding for the β-subunit of RNA polymerase. The aim of this work was to determine whether this mutation, and other mutations in rpoB, contribute to ciprofloxacin resistance and, if so, by which mechanism.MethodsIndependent lineages of E. coli were evolved in the presence of ciprofloxacin and clones from endpoint cultures were screened for mutations in rpoB. Ciprofloxacin-selected rpoB mutations were identified and characterized in terms of effects on susceptibility and mode of action.ResultsMutations in rpoB were selected at a high frequency in 3 out of 10 evolved lineages, in each case arising after the occurrence of mutations affecting topoisomerases and drug efflux. All ciprofloxacin-selected rpoB mutations had a high fitness cost in the absence of drug, but conferred a competitive advantage in the presence of ciprofloxacin. RNA sequencing and quantitative RT–PCR analysis showed that expression of mdtK, encoding a multidrug efflux transporter, was significantly increased by the ciprofloxacin-selected rpoB mutations. The susceptibility phenotype was shown to depend on the presence of an active mdtK and a mutant rpoB allele.ConclusionsThese data identify mutations in RNA polymerase as novel contributors to the evolution of resistance to ciprofloxacin and show that the phenotype is mediated by increased MdtK-dependent drug efflux.
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| 12. |
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| 13. |
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| 14. |
- Søgaard Jørgensen, Peter, et al.
(författare)
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Coevolutionary Governance of Antibiotic and Pesticide Resistance
- 2020
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Ingår i: Trends in Ecology & Evolution. - : Elsevier BV. - 0169-5347 .- 1872-8383. ; 35:6, s. 484-494
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Forskningsöversikt (refereegranskat)abstract
- Development of new biocides has dominated human responses to evolution of antibiotic and pesticide resistance. Increasing and uniform biocide use, the spread of resistance genes, and the lack of new classes of compounds indicate the importance of navigating toward more sustainable coevolutionary dynamics between human culture and species that evolve resistance. To inform this challenge, we introduce the concept of coevolutionary governance and propose three priorities for its implementation: (i) new norms and mental models for lowering use, (ii) diversifying practices to reduce directional selection, and (iii) investment in collective action institutions to govern connectivity. We highlight the availability of solutions that facilitate broader sustainable development, which for antibiotic resistance include improved sanitation and hygiene, strong health systems, and decreased meat consumption.
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| 15. |
- Zorzet, Anna, 1977-, et al.
(författare)
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Compensatory evolution restores fitness to actinonin-resistant Staphylococcus aureus
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- We have studied the emergence of actinonin-resistant mutants in S. aureus. In accordance with earlier studies we identified resistance mutations in the fmt gene that apart from conferring high-level resistance also reduced the growth rate. To study how fitness could be restored we performed compensatory evolution by serial passage of cells in liquid culture. Compensated mutants arose quickly, within 40 generations and sometimes in less than 10 generations. The mutants were fully resistant to actinonin and showed increased growth rates on plates. However, exponential growth rates in liquid media were not higher than for the parental resistant mutants. We sequenced the whole genomes of one slow-growing strain and three compensated and found alterations in three genes. These genes were SAOUHSC_01699 coding for shikimate 5-dehydrogenase, SAOUHSC_00945 coding for a magnesium transporter and SAOUHSC_02264, coding for accessory gene regulator protein C (AgrC). None of these mutants were obvious candidates for compensating lack of formylation. When sequencing these three loci in the remainder of the compensated strains, mutations in agrC were found in 11 of 16 strains. Further studies will show if these mutations are due to general environmental adaption, specific adaptation for slow growth or specific compensation for loss of formylation.
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| 16. |
- Zorzet, Anna, et al.
(författare)
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Compensatory mutations in agrC partly restore fitness in vitro to peptide deformylase inhibitor-resistant Staphylococcus aureus
- 2012
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Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press (OUP). - 0305-7453 .- 1460-2091. ; 67:8, s. 1835-1842
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVESTo determine how the fitness cost of deformylase inhibitor resistance conferred by fmt mutations can be genetically compensated.METHODSResistant mutants were isolated and characterized with regard to their growth rates in vitro and in neutropenic mice, MIC and DNA sequence. Faster-growing compensated mutants were isolated by serial passage in culture medium, and for a subset of the resistant and compensated mutants whole-genome sequencing was performed.RESULTSStaphylococcus aureus mutants resistant to the peptide deformylase inhibitor actinonin had mutations in the fmt gene that conferred high-level actinonin resistance and reduced bacterial growth rate. Compensated mutants that remained fully resistant to actinonin and showed increased growth rates appeared within 30-60 generations of growth. Whole-genome sequencing and localized DNA sequencing of mutated candidate genes showed that alterations in the gene agrC were present in the majority of compensated strains. Resistant and compensated mutants grew at similar rates as the wild-type in a mouse thigh infection model.CONCLUSIONSResistance to deformylase inhibitors due to fmt mutations reduces bacterial growth rates, but these costs can be reduced by mutations in the agrC gene. Mutants defective in fmt (with or without compensatory agrC mutations) grew well in an animal model, implying that they can also cause infection in a host.
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| 17. |
- Zorzet, Anna, 1977-, et al.
(författare)
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Error-prone initiation factor 2 mutations reduce the fitness cost of antibiotic resistance
- 2010
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Ingår i: Molecular Microbiology. - : Wiley-Blackwell. - 0950-382X .- 1365-2958. ; 75:5, s. 1299-1313
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Tidskriftsartikel (refereegranskat)abstract
- Mutations in the fmt gene (encoding formyl methionine transferase) that eliminate formylation of initiator tRNA (Met-tRNA(i)) confer resistance to the novel antibiotic class of peptide deformylase inhibitors (PDFIs) while concomitantly reducing bacterial fitness. Here we show in Salmonella typhimurium that novel mutations in initiation factor 2 (IF2) located outside the initiator tRNA binding domain can partly restore fitness of fmt mutants without loss of antibiotic resistance. Analysis of initiation of protein synthesis in vitro showed that with non-formylated Met-tRNA(i) IF2 mutants initiated much faster than wild-type IF2, whereas with formylated fMet-tRNA(i) the initiation rates were similar. Moreover, the increase in initiation rates with Met-tRNA(i) conferred by IF2 mutations in vitro correlated well with the increase in growth rate conferred by the same mutations in vivo, suggesting that the mutations in IF2 compensate formylation deficiency by increasing the rate of in vivo initiation with Met-tRNA(i). IF2 mutants had also a high propensity for erroneous initiation with elongator tRNAs in vitro, which could account for their reduced fitness in vivo in a formylation-proficient strain. More generally, our results suggest that bacterial protein synthesis is mRNA-limited and that compensatory mutations in IF2 could increase the persistence of PDFI-resistant bacteria in clinical settings.
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| 18. |
- Zorzet, Anna, 1977-
(författare)
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Mechanisms of Adaptation to Deformylase Inhibitors
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Antibiotic resistance is a growing problem on a global scale. Increasing numbers of bacteria resistant toward one or multiple antibiotics could return us to the high mortality rates for infectious diseases of the pre-antibiotic era. The need for development of new classes of antibiotics is great as is increased understanding of the mechanisms underlying the development of antibiotic resistance. We have investigated the emergence of resistance to peptide deformylase inhibitors, a new class of antibiotics that target bacterial protein synthesis. The fitness of resistant mutants as well as their propensity to acquire secondary compensatory mutations was assessed in order to gain some insight into the potential clinical risk of resistance development. Most of this work was done in the bacterium Salmonella typhimurium, due to the availability of excellent genetic tools to study these phenomena. In addition, we have studied the bacterium Staphylococcus aureus as peptide deformylase inhibitors have been shown to have the greatest effect on Gram-positive organisms. In the course of this work we also examined the mechanistic aspects of translation initiation. Using a cell-free in vitro translation system we studied the effects of various components on translation initiation. These results have been combined with results obtained from resistant and compensated bacterial strains in vivo to gain new insights into the mechanisms of translation initiation.
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| 19. |
- Zorzet, Anna
(författare)
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Overcoming scientific and structural bottlenecks in antibacterial discovery and development
- 2014
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Ingår i: Upsala Journal of Medical Sciences. - : Uppsala Medical Society. - 0300-9734 .- 2000-1967. ; 119:2, s. 170-175
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Forskningsöversikt (refereegranskat)abstract
- Antibiotic resistance is becoming an increasing threat, with too few novel antibiotics coming to market to replace those lost due to resistance development. Efforts by the pharmaceutical industry to screen for and design novel antibacterials have not been successful, with several companies minimizing or closing down their antibacterial research units, leading to a loss of skills and know-how. At the same time, antibiotic innovation in academia is not filling the void due to misaligned incentive structures and lack of vital knowledge of drug discovery. The scientific and structural difficulties in discovering new antibiotics have only begun to be appreciated in the latest years. Part of the problem has been a paradigm shift within both industry and academia to focus on 'rational' drug development with an emphasis on single targets and high-throughput screening of large chemical libraries, which may not be suited to target bacteria. The very particular aspects of 'targeting an organism inside another organism' have not been given enough attention. In this paper, researcher interviews have complemented literature studies to delve deeper into the specifics of the different scientific and structural barriers, and some potential solutions are offered.
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| 20. |
- Årdal, Christine, et al.
(författare)
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International cooperation to improve access to and sustain effectiveness of antimicrobials
- 2016
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Ingår i: The Lancet. - 0140-6736 .- 1474-547X. ; 387:10015, s. 296-307
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Tidskriftsartikel (refereegranskat)abstract
- Securing access to effective antimicrobials is one of the greatest challenges today. Until now, efforts to address this issue have been isolated and uncoordinated, with little focus on sustainable and international solutions. Global collective action is necessary to improve access to life-saving antimicrobials, conserving them, and ensuring continued innovation. Access, conservation, and innovation are beneficial when achieved independently, but much more effective and sustainable if implemented in concert within and across countries. WHO alone will not be able to drive these actions. It will require a multisector response (including the health, agriculture, and veterinary sectors), global coordination, and financing mechanisms with sufficient mandates, authority, resources, and power. Fortunately, securing access to effective antimicrobials has finally gained a place on the global political agenda, and we call on policy makers to develop, endorse, and finance new global institutional arrangements that can ensure robust implementation and bold collective action.
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