Dan I Andersson

Uppsala University, Uppsala, Uppsala, Sweden

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Publications (138)985.35 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Objectives: In silico pharmacokinetic/pharmacodynamic (PK/PD) models can be developed based on data from in vitro time-kill experiments and can provide valuable information to guide dosing of antibiotics. The aim was to develop a mechanism-based in silico model that can describe in vitro time-kill experiments of Escherichia coli MG1655 WT and six isogenic mutants exposed to ciprofloxacin and to identify relationships that may be used to simplify future characterizations in a similar setting. Methods: In this study, we developed a mechanism-based PK/PD model describing killing kinetics for E. coli following exposure to ciprofloxacin. WT and six well-characterized mutants, with one to four clinically relevant resistance mutations each, were exposed to a wide range of static ciprofloxacin concentrations. Results: The developed model includes susceptible growing bacteria, less susceptible (pre-existing resistant) growing bacteria, non-susceptible non-growing bacteria and non-colony-forming non-growing bacteria. The non-colony-forming state was likely due to formation of filaments and was needed to describe data close to the MIC. A common model structure with different potency for bacterial killing (EC50) for each strain successfully characterized the time-kill curves for both WT and the six E. coli mutants. Conclusions: The model-derived mutant-specific EC50 estimates were highly correlated (r(2) = 0.99) with the experimentally determined MICs, implying that the in vitro time-kill profile of a mutant strain is reasonably well predictable by the MIC alone based on the model.
    Journal of Antimicrobial Chemotherapy 09/2015; DOI:10.1093/jac/dkv233 · 5.31 Impact Factor
  • Diarmaid Hughes · Dan I Andersson
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    ABSTRACT: Drug therapy has a crucial role in the treatment of viral, bacterial, fungal and protozoan infections, as well as the control of human cancer. The success of therapy is being threatened by the increasing prevalence of resistance. We examine and compare mechanisms of drug resistance in these diverse biological systems (using HIV and Plasmodium falciparum as examples of viral and protozoan pathogens, respectively) and discuss how factors - such as mutation rates, fitness effects of resistance, epistasis and clonal interference - influence the evolutionary trajectories of drug-resistant clones. We describe commonalities and differences related to resistance development that could guide strategies to improve therapeutic effectiveness and the development of a new generation of drugs.
    Nature Reviews Genetics 07/2015; 16(8). DOI:10.1038/nrg3922 · 36.98 Impact Factor
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    Dan I. Andersson · Jon Jerlström-Hultqvist · Joakim Näsvall
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    ABSTRACT: How the enormous structural and functional diversity of new genes and proteins was generated (estimated to be 10(10)-10(12) different proteins in all organisms on earth [Choi I-G, Kim S-H. 2006. Evolution of protein structural classes and protein sequence families. Proc Natl Acad Sci 103: 14056-14061] is a central biological question that has a long and rich history. Extensive work during the last 80 years have shown that new genes that play important roles in lineage-specific phenotypes and adaptation can originate through a multitude of different mechanisms, including duplication, lateral gene transfer, gene fusion/fission, and de novo origination. In this review, we focus on two main processes as generators of new functions: evolution of new genes by duplication and divergence of pre-existing genes and de novo gene origination in which a whole protein-coding gene evolves from a noncoding sequence. Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.
    Cold Spring Harbor perspectives in biology 06/2015; 7(6). DOI:10.1101/cshperspect.a017996 · 8.68 Impact Factor
  • Dan I Andersson
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    ABSTRACT: The methods used today by academic researchers and the pharmaceutical industry to assess the risk of emergence of resistance, for example during development of new antibiotics or when assessing an old antibiotic, are sub-optimal. Even though easy to perform the presently used serial passage procedures, minimal prevention concentration measurements and determination of mutation rates in vitro are generally providing inadequate knowledge to use for risk assessment and decisions to continue/discontinue drug development. These methods need to be complemented and replaced with more relevant methods such as determination of whether resistance genes already pre-exist in various metagenomes, and the likelihood that these genes can transfer into the relevant pathogens and be stably maintained. Furthermore, to determine the risk of emergence of mutationally conferred resistance the fitness effect of the resistance mechanism is key, since this parameter will determine the ability of the resistant mutants to be maintained and enriched in the host after they have emerged. This information combined with knowledge of bacterial population sizes and growth and killing dynamics at relevant infection sites should allow for better forecasting of the risk of resistance emerging in clinical settings. Copyright © 2015. Published by Elsevier Ltd.
    Clinical Microbiology and Infection 05/2015; DOI:10.1016/j.cmi.2015.05.012 · 5.77 Impact Factor
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    Elisabeth Thulin · Martin Sundqvist · Dan I Andersson
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    ABSTRACT: Mecillinam is a β-lactam antibiotic that is mainly used for treatment of uncomplicated urinary tract infections. The objectives of this study were to identify mutations that confer mecillinam resistance in laboratory-isolated mutants and clinical isolates of Escherichia coli and to determine why mecillinam resistance remains rare clinically even though resistance is easily selected in the laboratory. In laboratory selections, mutation frequencies to mecillinam resistance varied between 8x10(-8) - 2x10(-5) per cell depending on the concentration of mecillinam used during selection. Several genes have been demonstrated to give mecillinam resistance, but here eight novel genes, previously unknown to be involved in mecillinam resistance were identified. These genes encode functions involved in the respiratory chain, the ribosome, cystein biosynthesis, tRNA synthesis and pyrophosphate metabolism. The clinical isolates exhibited significantly higher fitness and a different mutation spectrum than the laboratory-isolated mutants. The cysB gene was mutated (inactivated) in all the clinical isolates, in contrast to the laboratory-isolated mutants where mainly other types of more costly mutations were found. Our results suggest that the mutation frequency to mecillinam resistance is high because of the large mutational target (at least 38 genes). However, the majority of these resistant mutants have a low growth rate, reducing the probability that they are stably maintained in the bladder. Inactivation of the cysB gene, and a resulting loss of biosynthesis of cysteine, is the major mechanism of mecillinam resistance in clinical isolates of E. coli. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Antimicrobial Agents and Chemotherapy 01/2015; 59(3). DOI:10.1128/AAC.04819-14 · 4.48 Impact Factor
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    ABSTRACT: To assure correct antibiotic treatment and reduce 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 to rapidly identify the bacterial species causing an infection and determine their antibiotic susceptibility profiles. An initial short cultivation step in the absence and presence of different antibiotics was combined with a sensitive species-specific padlock probe detection of the bacterial target DNA to allow determination of growth (i.e. resistance) and no growth (i.e. susceptibility). A proof-of-concept was established for urinary tract infections where we applied the method to determine the antibiotic susceptibility profile of E. coli for two drugs with 100% accuracy in 3.5 hours. The short assay time from sample to readout enables fast appropriate treatment with effective drugs, and minimizes the need of prescribing broad-spectrum antibiotics due to unknown resistance profiles of the treated infection. Copyright © 2014, American Society for Microbiology. All Rights Reserved.
    Journal of Clinical Microbiology 11/2014; 53(2). DOI:10.1128/JCM.02434-14 · 3.99 Impact Factor
  • Hava Lofton · Naeem Anwar · Mikeal Rhen · Dan I. Andersson
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    ABSTRACT: Abstract OBJECTIVES: To examine the effects of mutations in the waaY, phoP and pmrB genes, which confer resistance to antimicrobial peptides (AMPs), on fitness of Salmonella Typhimurium. METHODS: Survival during low pH, oxidative stress, stationary-phase incubation, exposure to serum and bile and growth in mice and laboratory media were determined by time-kills, disc inhibition assays, competition experiments and optical density measurements. RESULTS: Individual mutations in the waaY gene (involved in LPS core biosynthesis) and in the phoP and pmrB genes (part of two different two-component regulatory systems, phoPQ and pmrAB) conferred no or minor effects on bacterial survival during stressful in vitro conditions or in mice. In contrast, a waaY-phoP-pmrB triple mutant was compromised under most assay conditions. CONCLUSIONS: Results from this study show that AMP resistance can be cost-free, as assessed by several assays that attempt to mimic the conditions a bacterium might encounter within a host. Our findings imply that future therapeutic use of AMPs could select for fit mutants with cross-resistance to human defence peptides and that potential resistance development in response to therapeutic use of AMPs needs to be carefully monitored.
    Journal of Antimicrobial Chemotherapy 09/2014; 70(2). DOI:10.1093/jac/dku423 · 5.31 Impact Factor
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    ABSTRACT: How sublethal levels of antibiotics and heavy metals select for clinically important multidrug resistance plasmids is largely unknown. Carriage of plasmids generally confers substantial fitness costs, implying that for the plasmid-carrying bacteria to be maintained in the population, the plasmid cost needs to be balanced by a selective pressure conferred by, for example, antibiotics or heavy metals. We studied the effects of low levels of antibiotics and heavy metals on the selective maintenance of a 220-kbp extended-spectrum β-lactamase (ESBL) plasmid identified in a hospital outbreak of Klebsiella pneumoniae and Escherichia coli. The concentrations of antibiotics and heavy metals required to maintain plasmid-carrying bacteria, the minimal selective concentrations (MSCs), were in all cases below (almost up to 140-fold) the MIC of the plasmid-free susceptible bacteria. This finding indicates that the very low antibiotic and heavy metal levels found in polluted environments and in treated humans and animals might be sufficiently high to maintain multiresistance plasmids. When resistance genes were moved from the plasmid to the chromosome, the MSC decreased, showing that MSC for a specific resistance conditionally depends on genetic context. This finding suggests that a cost-free resistance could be maintained in a population by an infinitesimally low concentration of antibiotic. By studying the effect of combinations of several compounds, it was observed that for certain combinations of drugs each new compound added lowered the minimal selective concentration of the others. This combination effect could be a significant factor in the selection of multidrug resistance plasmids/bacterial clones in complex multidrug environments.
    mBio 08/2014; 5(5). DOI:10.1128/mBio.01918-14 · 6.79 Impact Factor
  • Dan I Andersson · Diarmaid Hughes
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    ABSTRACT: The widespread use of antibiotics results in the generation of antibiotic concentration gradients in humans, livestock and the environment. Thus, bacteria are frequently exposed to non-lethal (that is, subinhibitory) concentrations of drugs, and recent evidence suggests that this is likely to have an important role in the evolution of antibiotic resistance. In this Review, we discuss the ecology of antibiotics and the ability of subinhibitory concentrations to select for bacterial resistance. We also consider the effects of low-level drug exposure on bacterial physiology, including the generation of genetic and phenotypic variability, as well as the ability of antibiotics to function as signalling molecules. Together, these effects accelerate the emergence and spread of antibiotic-resistant bacteria among humans and animals.
    Nature Reviews Microbiology 05/2014; 12(7). DOI:10.1038/nrmicro3270 · 23.57 Impact Factor
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    Song Sun · Maria Selmer · Dan I Andersson
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    ABSTRACT: Penicillin-binding proteins (PBPs) are enzymes responsible for the polymerization of the glycan strand and the cross-linking between glycan chains as well as the target proteins for β-lactam antibiotics. Mutational alterations in PBPs can confer resistance either by reducing binding of the antibiotic to the active site or by evolving a β-lactamase activity that degrades the antibiotic. As no systematic studies have been performed to examine the potential of all PBPs present in one bacterial species to evolve increased resistance against β-lactam antibiotics, we explored the ability of fifteen different defined or putative PBPs in Salmonella enterica to acquire increased resistance against penicillin G. We could after mutagenesis and selection in presence of penicillin G isolate mutants with amino-acid substitutions in the PBPs, FtsI, DacB and DacC (corresponding to PBP3, PBP4 and PBP6) with increased resistance against β-lactam antibiotics. Our results suggest that: (i) most evolved PBPs became 'generalists" with increased resistance against several different classes of β-lactam antibiotics, (ii) synergistic interactions between mutations conferring antibiotic resistance are common and (iii) the mechanism of resistance of these mutants could be to make the active site more accessible for water allowing hydrolysis or less binding to β-lactam antibiotics.
    PLoS ONE 05/2014; 9(5):e97202. DOI:10.1371/journal.pone.0097202 · 3.23 Impact Factor
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    ABSTRACT: An important mechanism for generation of new genes is by duplication-divergence of existing genes. Duplication-divergence includes several different sub-models, such as subfunctionalization where after accumulation of neutral mutations the original function is distributed between two partially functional and complementary genes, and neofunctionalization where a new function evolves in one of the duplicated copies while the old function is maintained in another copy. The likelihood of these mechanisms depends on the longevity of the duplicated state, which in turn depends on the fitness cost and genetic stability of the duplications. Here, we determined the fitness cost and stability of defined gene duplications/amplifications on a low copy number plasmid. Our experimental results show that the costs of carrying extra gene copies are substantial and that each additional kbp of DNA reduces fitness by approximately 0.15%. Furthermore, gene amplifications are highly unstable and rapidly segregate to lower copy numbers in absence of selection. Mathematical modelling shows that the fitness costs and instability strongly reduces the likelihood of both sub- and neofunctionalization, but that these effects can be off-set by positive selection for novel beneficial functions.
    Molecular Biology and Evolution 03/2014; 31(6). DOI:10.1093/molbev/msu111 · 9.11 Impact Factor
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    ABSTRACT: Clonally derived bacterial populations exhibit significant genotypic and phenotypic diversity that contribute to fitness in rapidly changing environments. Here, we show that serial passage of Salmonella enterica serovar Typhimurium LT2 (StLT2) in broth, or within a mouse host, results in selection of an evolved population that inhibits the growth of ancestral cells by direct contact. Cells within each evolved population gain the ability to express and deploy a cryptic "orphan" toxin encoded within the rearrangement hotspot (rhs) locus. The Rhs orphan toxin is encoded by a gene fragment located downstream of the "main" rhs gene in the ancestral strain StLT2. The Rhs orphan coding sequence is linked to an immunity gene, which encodes an immunity protein that specifically blocks Rhs orphan toxin activity. Expression of the Rhs orphan immunity protein protects ancestral cells from the evolved lineages, indicating that orphan toxin activity is responsible for the observed growth inhibition. Because the Rhs orphan toxin is encoded by a fragmented reading frame, it lacks translation initiation and protein export signals. We provide evidence that evolved cells undergo recombination between the main rhs gene and the rhs orphan toxin gene fragment, yielding a fusion that enables expression and delivery of the orphan toxin. In this manner, rhs locus rearrangement provides a selective advantage to a subpopulation of cells. These observations suggest that rhs genes play important roles in intra-species competition and bacterial evolution.
    PLoS Genetics 03/2014; 10(3):e1004255. DOI:10.1371/journal.pgen.1004255 · 7.53 Impact Factor
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    ABSTRACT: Genes introduced by horizontal gene transfer (HGT) from other species constitute a significant portion of many bacterial genomes and the evolutionary dynamics of HGTs are important for understanding the spread of antibiotic resistance and the emergence of new pathogenic strains of bacteria. The fitness effects of the transferred genes largely determine the fixation rates and the amount of neutral diversity of newly acquired genes in bacterial populations. Comparative analysis of bacterial genomes provides insight into what genes are commonly transferred, but direct experimental tests of the fitness constraints on HGT are scarce. Here, we address this paucity of experimental studies by introducing 98 random DNA fragments varying in size from 0.45 to 5 kb from Bacteroides, Proteus and human intestinal phage into a defined position in the Salmonella chromosome and measuring the effects on fitness. Using highly sensitive competition assays, we found that 8 inserts were deleterious with selection coefficients (s) ranging from ≈ -0.007 to -0.02 and ninety did not have significant fitness effects. When inducing transcription from a PBAD promoter located at one end of the inserts, 16 transfers were deleterious and 82 were not significantly different from the control. In conclusion, a major fraction of the inserts had minor effects on fitness implying that extra DNA transferred by HGT, even though it does not confer an immediate selective advantage, could be maintained at selection-transfer balance and serve as raw material for the evolution of novel beneficial functions.
    Molecular Biology and Evolution 02/2014; 31(5). DOI:10.1093/molbev/msu076 · 9.11 Impact Factor
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    Joakim Näsvall · Lei Sun · John R Roth · Dan I Andersson
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    Hervé Nicoloff · Dan I Andersson
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    ABSTRACT: Previous work demonstrated that selection for Escherichia coli mutants with low antibiotic resistance frequently resulted in co-selection of lon mutations and that lon(-) mutants evolved higher-level resistance faster than a lon(+) strain. Here we show that lon mutation causes a very low multidrug resistance by inducing the AcrAB-TolC pump via stabilization of the acrAB transcriptional activators MarA and SoxS, which are substrates of the Lon protease. Fast evolution of lon(-) mutants towards higher resistance involves selection of frequent next-step mutations consisting of large duplications including acrAB and the mutated lon gene. Resistance results from the combined effects of acrAB duplication and lon mutation increasing dosage of efflux pump. In contrast, when acrAB duplication occurs as the first step mutation, increased Lon activity caused by lon(+) co-duplication mitigates the effect of acrAB duplication on resistance, and faster evolution towards higher resistance is not observed. As predicted, when the functional lon gene is relocated far from acrAB to prevent their co-duplication, first-step acrAB duplication confers higher resistance, which then allows selection of frequent next-step mutations and results in faster evolution towards higher resistance. Our results demonstrate how order of appearance of mutations and gene location can influence the rate of resistance evolution.
    Molecular Microbiology 12/2013; 90(6):1233-48. DOI:10.1111/mmi.12429 · 4.42 Impact Factor
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    ABSTRACT: How pathogenic bacteria adapt and evolve in the complex and variable environment of the host remains a largely unresolved question. Here we have used whole genome sequencing of Salmonella enterica serovar Typhimurium LT2 populations serially passaged in mice to identify mutations that adapt bacteria to systemic growth in mice. We found unique pathoadaptive mutations in two global regulators, phoQ and stpA, which increase the competitive indexes of the bacteria 3- to 5-fold. Also, all mouse-adapted lineages had changed the orientation of the hin invertable element, resulting in production of a FliC type of flagellum. Competition experiments in mice with locked flagellum mutants showed that strains expressing the FliC type of flagellum had a 5-fold increase in competitive index as compared to those expressing FljB type flagellum. Combination of the flagellum cassette inversion with the stpA mutation increased competitive indexes up to 20-fold. These experiments show that Salmonella can rapidly adapt to a mouse environment by acquiring a few mutations of moderate individual effect that when combined confer substantial increases in growth.
    PLoS ONE 07/2013; 8(7):e70147. DOI:10.1371/journal.pone.0070147 · 3.23 Impact Factor
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    Hava Lofton · Maria Pränting · Elisabeth Thulin · Dan I Andersson
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    ABSTRACT: Antimicrobial peptides (AMPs) represent a potential new class of antimicrobial drugs with potent and broad-spectrum activities. However, knowledge about the mechanisms and rates of resistance development to AMPs and the resulting effects on fitness and cross-resistance is limited. We isolated antimicrobial peptide (AMP) resistant Salmonella typhimurium LT2 mutants by serially passaging several independent bacterial lineages in progressively increasing concentrations of LL-37, CNY100HL and Wheat Germ Histones. Significant AMP resistance developed in 15/18 independent bacterial lineages. Resistance mutations were identified by whole genome sequencing in two-component signal transduction systems (pmrB and phoP) as well as in the LPS core biosynthesis pathway (waaY, also designated rfaY). In most cases, resistance was associated with a reduced fitness, observed as a decreased growth rate, which was dependent on growth conditions and mutation type. Importantly, mutations in waaY decreased bacterial susceptibility to all tested AMPs and the mutant outcompeted the wild type parental strain at AMP concentrations below the MIC for the wild type. Our data suggests that resistance to antimicrobial peptides can develop rapidly through mechanisms that confer cross-resistance to several AMPs. Importantly, AMP-resistant mutants can have a competitive advantage over the wild type strain at AMP concentrations similar to those found near human epithelial cells. These results suggest that resistant mutants could both be selected de novo and maintained by exposure to our own natural repertoire of defence molecules.
    PLoS ONE 07/2013; 8(7):e68875. DOI:10.1371/journal.pone.0068875 · 3.23 Impact Factor
  • Marius Linkevicius · Linus Sandegren · Dan I Andersson
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    ABSTRACT: To stepwise select tigecycline-resistant Escherichia coli mutants in vitro, determine the mutation rates, identify the resistance mechanisms, determine the resistance level and cross-resistance to other antibiotic classes, evaluate the fitness costs of tigecycline resistance mechanisms and investigate if the same in vitro-identified target genes were mutated in clinical isolates. Spontaneous mutants with reduced susceptibility to tigecycline were selected on agar plates supplemented with tigecycline. Resistance levels and cross-resistance were evaluated by performing MIC assays and determining mutation rates using Luria-Delbruck fluctuation tests. Mutant fitness was estimated by measuring exponential growth rates, lag phase and total yield. Illumina whole-genome sequencing was used to identify mutations increasing MICs of tigecycline. Spontaneous mutants with reduced susceptibility to tigecycline were selected at a rate of ∼10(-8) to 10(-6) per cell per generation; however, the clinical MIC breakpoint was not reached. The resistance level of tigecycline was low and some of the mutants had elevated MICs of hydrophobic drugs (chloramphenicol, erythromycin and novobiocin) or decreased MICs of SOS response inducers (ciprofloxacin and nitrofurantoin). Mutations were identified in efflux regulatory network genes (lon, acrR and marR) or lipopolysaccharide core biosynthesis pathway genes (lpcA, rfaE, rfaD, rfaC and rfaF). Mutations in the same target genes were found in clinical isolates. Tigecycline selects for low-level resistance mutations with relatively high mutation rates and the majority of them come with a substantial fitness cost. Further in vivo experiments are needed to evaluate how these mutations affect bacterial virulence and ability to establish a successful infection.
    Journal of Antimicrobial Chemotherapy 07/2013; 68(12). DOI:10.1093/jac/dkt263 · 5.31 Impact Factor
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    Peter A Lind · Dan I Andersson
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    ABSTRACT: We previously reported that the distribution of fitness effects for non-synonymous and synonymous mutations in Salmonella typhimurium ribosomal proteins S20 and L1 are similar, suggesting that fitness constraints are present at the level of mRNA. Here we explore the hypothesis that synonymous mutations confer their fitness-reducing effect by alterating the secondary structure of the mRNA. To this end, we constructed a set of synonymous substitutions in the rpsT gene, encoding ribosomal protein S20, that are located in predicted paired regions in the mRNA and measured their effect on bacterial fitness. Our results show that for 3/9 cases tested, the reduced fitness conferred by a synonymous mutation could be fully or partly restored by introducing a second synonymous substitution that restore base pairing in a mRNA stem. In addition, random mutations in predicted paired regions had larger fitness effects than those in unpaired regions. Finally, we did not observe any correlation between fitness effects of the synonymous mutations and their rarity. These results suggest that for ribosomal protein S20, the deleterious effects of synonymous mutations are not generally due to codon usage effects, but that mRNA secondary structure is a major fitness constraint.
    PLoS ONE 05/2013; 8(5):e63373. DOI:10.1371/journal.pone.0063373 · 3.23 Impact Factor

Publication Stats

7k Citations
985.35 Total Impact Points


  • 1991–2015
    • Uppsala University
      • • Department of Medical Biochemistry and Microbiology
      • • Department of Cell and Molecular Biology
      Uppsala, Uppsala, Sweden
  • 2004–2009
    • Karolinska Institutet
      • Department of Microbiology, Tumor and Cell Biology (MTC)
      Solna, Stockholm, Sweden
    • Hospital Universitario Ramón y Cajal
      • Departamento de Microbiologia y Parasitología
      Madrid, Madrid, Spain
  • 1999–2006
    • Swedish Institute for Communicable Disease Control
      Tukholma, Stockholm, Sweden
  • 2005
    • KTH Royal Institute of Technology
      • School of Biotechnology (BIO)
      Tukholma, Stockholm, Sweden
  • 2003
    • Technische Universität München
      München, Bavaria, Germany
    • Uppsala University Hospital
      Uppsala, Uppsala, Sweden
  • 1990–2003
    • University of Utah
      • Department of Biology
      Salt Lake City, UT, United States