Evolution of the Pseudomonas aeruginosa Aminoglycoside Mutational Resistome
            <i>In Vitro</i>
            and in the Cystic Fibrosis Setting

López-Causapé, Carla; Rubio, Rosa; Cabot, Gabriel; Oliver, Antonio

doi:10.1128/aac.02583-17

Cited by 50 publications

(55 citation statements)

References 34 publications

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“…To examine if the precise mutations found in our in vitro study were also found in clinical isolates, we searched published genomes of P. aeruginosa clinical isolates from cystic fibrosis patients who had likely been treated with aminoglycosides like TOB (Bolard et al, 2017; Chung et al, 2012; López-Causapé et al, 2017, 2018; Markussen et al, 2014). We identified fusA1 and ptsP mutations in these genomes, suggesting that these mutations evolve during infections.…”

Section: Resultsmentioning

confidence: 99%

Parallel evolution of tobramycin resistance across species and environments

Scribner

Santos-López

Marshall

et al. 2019

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words)13 An important problem in evolution is identifying the genetic basis of how different species adapt 14 to similar environments. Understanding how various bacterial pathogens evolve in response to 15 antimicrobial treatment is a pressing example of this problem, where discovery of molecular 16 parallelism could lead to clinically useful predictions. Evolution experiments with pathogens in 17 environments containing antibiotics combined with periodic whole population genome 18 sequencing can be used to characterize the evolutionary dynamics of the pathways to 19 antimicrobial resistance. We separately propagated two clinically relevant Gram-negative 20 pathogens, Pseudomonas aeruginosa and Acinetobacter baumannii, in increasing 21 concentrations of tobramycin in two different environments each: planktonic and biofilm. 22Independent of the pathogen, populations adapted to tobramycin selection by parallel evolution 23 of mutations in fusA1, encoding elongation factor G, and ptsP, encoding phosphoenolpyruvate 24 phosphotransferase. As neither gene is a direct target of this aminoglycoside, both are relatively 25 novel and underreported causes of resistance. Additionally, both species acquired antibiotic-26 associated mutations that were more prevalent in the biofilm lifestyle than planktonic, in electron 27 transport chain components in A. baumannii and LPS biosynthesis enzymes in P. aeruginosa 28 populations. Using existing databases, we discovered both fusA1 and ptsP mutations to be 29 prevalent in antibiotic resistant clinical isolates. Additionally, we report site-specific parallelism of 30 fusA1 mutations that extend across several bacterial phyla. This study suggests that strong 31 selective pressures such as antibiotic treatment may result in high levels of predictability in 32 molecular targets of evolution despite differences between organisms' genetic background and 33 environment.The notion that evolution can be forecasted at the level of phenotype, gene, or even 36 amino acid is no longer a fantasy in the post-genomic era (Lässig et al., 2017). If we 37 acknowledge that most forecasting efforts rely on history to anticipate the future, the explosive 38 growth of whole-genome sequencing (WGS) sets the stage to resolve evolutionary phenomena 39 in action and suggest the next selected path. Among the best examples, bacterial populations 40 exposed to strong selection like antibiotics and analyzed by WGS are likely to identify gene 41 regions that produce resistance (Ahmed et al., 2018a; Cooper, 2018; Feng et al., 2016; Palmer 42 and Kishony, 2013). Repeated instances of the same antibiotic selection may enrich the same 43 types of mutations and ultimately enable some measure of predictability (Ibacache-Quiroga et 44 al., 2018; Wong et al., 2012). For instance, we can be confident that exposure of many bacteria 45 to high doses of fluoroquinolones like ciprofloxacin may select for substitutions in residues 83 or 46 87 of the drug target, DNA gyrase A (Fàbrega et al., 2009; Wong and Kassen, 2011).47 Furt...

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Section: Resultsmentioning

confidence: 99%

Parallel evolution of tobramycin resistance across species and environments

Scribner

Santos-López

Marshall

et al. 2019

Preprint

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“…We used PROVEAN to assess the frequency of likely change-of-function mutations in isolates of P. aeruginosa from patients. For comparison we analysed a panel of isolates from the general environment (Table 2) A high proportion of clinical isolates contained likely function-altering differences in genes that are established as contributing to clinical resistance to fluoroquinolones (gyrA and parE), carbapenems (mexR, nalD) and/ or aminoglycosides (mexY, amgS) (Table2) (27,28,(58)(59)(60)(61).…”

Section: Discussionmentioning

confidence: 99%

“…Experimental evolution of antibiotic-resistant mutants from sensitive parent strains, followed by whole genome sequencing to identify resistance-conferring mutations, is an approach that has been applied to a number of species (16)(17)(18)(19)(20)(21), including P. aeruginosa (22)(23)(24)(25)(26)(27)(28). Studies have confirmed the involvement of genes previously proposed to be associated with resistance in P. aeruginosa.…”

Section: Introductionmentioning

confidence: 87%

“…Studies have confirmed the involvement of genes previously proposed to be associated with resistance in P. aeruginosa. These include the fusA1 gene that encodes the EFG-1 protein and is associated with aminoglycoside resistance (27,29,30) and the ftsI gene that encodes a penicillin binding protein and is associated with carbapenem resistance (30)(31)(32)(33)(34). This approach has high potential for identifying previously unknown antibiotic resistanceassociated mutations and genes, but some key issues remain to be addressed.…”

Section: Introductionmentioning

confidence: 99%

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A large-scale comparison shows that genetic changes causing antibiotic resistance in experimentally evolvedPseudomonas aeruginosapredict those in naturally evolved bacteria

Wardell¹,

Rehman²,

Martin³

et al. 2019

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Pseudomonas aeruginosa is an opportunistic pathogen that causes a wide range of acute and chronic infections. An increasing number of isolates have acquired mutations that make them antibiotic resistant, making treatment more difficult. To identify resistance-associated mutations we experimentally evolved the antibiotic sensitive strain P. aeruginosa PAO1 to become resistant to three widely used anti-pseudomonal antibiotics, ciprofloxacin, meropenem and tobramycin. Mutants were able to tolerate up to 2048-fold higher concentrations of antibiotic than strain PAO1. Genome sequences were determined for thirteen mutants for each antibiotic. Each mutant had between 2 and 8 mutations. There were at least 8 genes mutated in more than one mutant per antibiotic, demonstrating the complexity of the genetic basis of resistance. Additionally, large deletions of up to 479kb arose in multiple meropenem resistant mutants. For all three antibiotics mutations arose in genes known to be associated with resistance, but also in genes not previously associated with resistance. To determine the clinical relevance of mutations uncovered in experimentallyevolved mutants we analysed the corresponding genes in 457 isolates of P. aeruginosa from patients with cystic fibrosis or bronchiectasis as well as 172 isolates from the general environment. Many of the genes identified through experimental evolution had changes predicted to be function-altering in clinical isolates but not in isolates from the general environment, showing that mutated genes in experimentally evolved bacteria can predict those that undergo mutation during infection. These findings expand understanding of the genetic basis of antibiotic resistance in P. aeruginosa as well as demonstrating the validity of experimental evolution in identifying clinically-relevant resistance-associated mutations.' ImportanceThe rise in antibiotic resistant bacteria represents an impending global health crisis. As such, understanding the genetic mechanisms underpinning this resistance can be a crucial piece of the puzzle to combatting it. The importance of this research is that by experimentally evolving P. aeruginosa to three clinically relevant antibiotics, we have generated a catalogue of genes that can contribute to resistance in vitro. We show that many (but not all) of these genes are clinically relevant, by identifying variants in clinical isolates of P. aeruginosa. This research furthers our understanding of the genetics leading to resistance in P. aeruginosa and provides tangible evidence that these genes can play a role clinically, potentially leading to new druggable targets or inform therapies.

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“…Particularly relevant is the activity of chromosomally-encoded multidrug-resistance (MDR) efflux pumps (10, 11). Further, the acquisition of mutation-driven resistance is common in this opportunistic pathogen, particularly along chronic infections (12–14), where the constitutive overexpression of MDR efflux pumps is one of the biggest problems to eradicate these infections (10, 15, 16). Efflux pumps exhibit different functions, with physiological and ecological significances that go beyond their activity as antibiotic resistance elements (10, 17–19).…”

Section: Introductionmentioning

confidence: 99%