Dataset Information

Dynamic Emergence of Mismatch Repair Deficiency Facilitates Rapid Evolution of Ceftazidime-Avibactam Resistance in Pseudomonas aeruginosa Acute Infection.

ABSTRACT: Strains of Pseudomonas aeruginosa with deficiencies in DNA mismatch repair have been studied in the context of chronic infection, where elevated mutational rates ("hypermutation") may facilitate the acquisition of antimicrobial resistance. Whether P. aeruginosa hypermutation can also play an adaptive role in the more dynamic context of acute infection remains unclear. In this work, we demonstrate that evolved mismatch repair deficiencies may be exploited by P. aeruginosa to facilitate rapid acquisition of antimicrobial resistance in acute infection, and we directly document rapid clonal succession by such a hypermutating lineage in a patient. Whole-genome sequencing (WGS) was performed on nine serially cultured blood and respiratory isolates from a patient in whom ceftazidime-avibactam (CZA) resistance emerged in vivo over the course of days. The CZA-resistant clone was differentiated by 14 mutations, including a gain-of-function G183D substitution in the PDC-5 chromosomal AmpC cephalosporinase conferring CZA resistance. This lineage also contained a substitution (R656H) at a conserved position in the ATPase domain of the MutS mismatch repair (MMR) protein, and elevated mutational rates were confirmed by mutational accumulation experiments with WGS of evolved lineages in conjunction with rifampin resistance assays. To test whether MMR-deficient hypermutation could facilitate rapid acquisition of CZA resistance, in vitro adaptive evolution experiments were performed with a mutS-deficient strain. These experiments demonstrated rapid hypermutation-facilitated acquisition of CZA resistance compared with the isogenic wild-type strain. Our results suggest a possibly underappreciated role for evolved MMR deficiency in facilitating rapid adaptive evolution of P. aeruginosa in the context of acute infection.IMPORTANCE Antimicrobial resistance in bacteria represents one of the most consequential problems in modern medicine, and its emergence and spread threaten to compromise central advances in the treatment of infectious diseases. Ceftazidime-avibactam (CZA) belongs to a new class of broad-spectrum beta-lactam/beta-lactamase inhibitor combinations designed to treat infections caused by multidrug-resistant bacteria. Understanding the emergence of resistance to this important new drug class is of critical importance. In this work, we demonstrate that evolved mismatch repair deficiency in P. aeruginosa, an important pathogen responsible for significant morbidity and mortality among hospitalized patients, may facilitate rapid acquisition of resistance to CZA in the context of acute infection. These findings are relevant for both diagnosis and treatment of antimicrobial resistance emerging in acute infection in the hypermutator background and additionally have implications for the emergence of more virulent phenotypes.

SUBMITTER: Khil PP

PROVIDER: S-EPMC6751058 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Dynamic Emergence of Mismatch Repair Deficiency Facilitates Rapid Evolution of Ceftazidime-Avibactam Resistance in Pseudomonas aeruginosa Acute Infection.

Khil Pavel P PP Dulanto Chiang Augusto A Ho Jonathan J Youn Jung-Ho JH Lemon Jamie K JK Gea-Banacloche Juan J Frank Karen M KM Parta Mark M Bonomo Robert A RA Dekker John P JP

mBio 20190917 5

Strains of Pseudomonas aeruginosa with deficiencies in DNA mismatch repair have been studied in the context of chronic infection, where elevated mutational rates ("hypermutation") may facilitate the acquisition of antimicrobial resistance. Whether P. aeruginosa hypermutation can also play an adaptive role in the more dynamic context of acute infection remains unclear. In this work, we demonstrate that evolved mismatch repair deficiencies may be exploited by P. aeruginosa to ...[more]

PMID: 31530672

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Project description:Klebsiella pneumoniae carbapenemase (KPC)-producing Pseudomonas aeruginosa (KPC-PA) has been reported sporadically. However, epidemiological and antimicrobial susceptibility data specific for KPC-PA are lacking. We collected 374 carbapenem-resistant P. aeruginosa (CRPA) isolates from seven hospitals in China from June 2016 to February 2019 and identified the blaKPC-2 gene in 40.4% (n = 151/374) of the isolates. Approximately one-half of all KPC-PA isolates (n = 76/151; 50.3%) were resistant to ceftazidime-avibactam (CAZ-AVI). Combining Kraken2 taxonomy identification and Nanopore sequencing, we identified eight plasmid types, five of which carried blaKPC-2, and 13 combination patterns of these plasmid types. In addition, we identified IS26-ΔTn6296 and Tn1403-like-ΔTn6296 as the two mobile genetic elements that mediated blaKPC-2 transmission. blaKPC-2 plasmid curing in 28 strains restored CAZ-AVI susceptibility, suggesting that blaKPC-2 was the mediator of CAZ-AVI resistance. Furthermore, the blaKPC-2 copy number was found to correlate with KPC expression and, therefore, CAZ-AVI resistance. Taken together, our results suggest that KPC-PA is becoming a clinical threat and that using CAZ-AVI to treat this specific pathogen should be done with caution. IMPORTANCE Previous research has reported several cases of KPC-PA strains and three KPC-encoding P. aeruginosa plasmid types in China. However, the prevalence and clinical significance of KPC-PA are not available. In addition, the susceptibility of the strains to CAZ-AVI remains unknown. Samples in this study were collected from seven tertiary hospitals prior to CAZ-AVI clinical approval in China. Therefore, our results represent a retrospective study establishing the baseline efficacy of the novel β-lactam/β-lactamase combination agent for treating KPC-PA infections. The observed correlation between the blaKPC copy number and CAZ-AVI resistance suggests that close monitoring of the susceptibility of the strain during treatment is required. It would also be beneficial to screen for the blaKPC gene in CRPA strains for antimicrobial surveillance purposes.

Project description:Carbapenem-resistant Pseudomonas aeruginosa (CRPA) has become a serious challenge in the clinic. Recently, the prevalence of CRPA isolates carrying the blaKPC-2 gene has been increasing in China. Ceftazidime-avibactam (CZA) has shown good efficacy against large portions of KPC-2-producing CRPA strains. However, with the increasing usage of this drug, CZA resistance in CRPA strains has been reported. Here, we reported for the first time that resistance of the ST463 CRPA strain to CZA was caused by a novel variant in the KPC gene that arose after CZA exposure. The CRPA strain PA2207 is a carbapenem- and CZA-resistant strain that harbors a mutated blaKPC gene, named blaKPC-90. Cloning and expression of blaKPC-90 in Escherichia coli DH5α revealed that KPC-90 led to a 64-fold increase in the MIC value of CZA. Conjugation experiments further confirmed that blaKPC-90 was located on a conjugative plasmid. Whole-genome sequencing analysis showed that this plasmid had high sequence similarity to a previously reported novel blaKPC-2-harboring plasmid in a clinical P. aeruginosa strain isolated in China. In addition, overexpression of an efflux pump (MexXY-OprM) might be associated with the CZA resistance phenotype, as determined by reverse transcription-quantitative PCR and efflux pump inhibition experiments. For the first time, we reported a KPC variant, KPC-90, in a clinical ST463 CRPA strain with CZA resistance that was mediated by a 2 amino acid insertion outside the KPC omega-loop region. Our study further highlights that diverse KPC variants that mediate CZA resistance have emerged in the CRPA strain. Furthermore, KPC-90 mutation combined with efflux pump overexpression resulted in a high level of resistance to CZA in the PA2207 isolate. Effective surveillance should be conducted to prevent CZA resistance from spreading in the CRPA strain. IMPORTANCE For the first time, we reported a KPC variant, KPC-90, in a clinical ST463 CRPA strain with CZA resistance. CZA resistance was mediated by a 2 amino acid insertion outside the KPC omega-loop region in CRPA. Our study further emphasized that CZA resistance caused by blaKPC gene mutation could be selected in CRPA after CZA therapy. Considering the widespread presence of the ST463 CRPA strain in China, clinicians should pay attention to the risk of the development of CZA resistance in CRPA strains under treatment pressure.

Project description:Broth microdilution antimicrobial susceptibility testing was performed for ceftazidime-avibactam and comparator agents against 7,062 clinical isolates of Pseudomonas aeruginosa collected from 2012 to 2014 in four geographic regions (Europe, Asia/South Pacific, Latin America, Middle East/Africa) as part of the International Network for Optimal Resistance Monitoring (INFORM) global surveillance program. The majority of isolates were susceptible to ceftazidime-avibactam, with the proportions susceptible differing marginally across the four regions (MIC90, 8 to 16 ?g/ml; 88.7 to 93.2% susceptible), in contrast to lower susceptibilities to the following comparator ?-lactam agents: ceftazidime (MIC90, 32 to 64 ?g/ml; 71.5 to 80.8% susceptible), meropenem (MIC90, >8 ?g/ml; 64.9 to 77.4% susceptible), and piperacillin-tazobactam (MIC90, >128 ?g/ml; 62.3 to 71.3% susceptible). Compared to the overall population, susceptibility to ceftazidime-avibactam of isolates that were nonsusceptible to ceftazidime (n = 1,627) was reduced to between 56.8% (Middle East/Africa; MIC90, 64 ?g/ml) and 68.9% (Asia/South Pacific; MIC90, 128 ?g/ml), but these percentages were higher than susceptibilities to other ?-lactam agents (0 to 44% susceptible, depending on region and agent; meropenem MIC90, >8 ?g/ml; 26.5 to 43.9% susceptible). For this subset of isolates, susceptibilities to amikacin (MIC90, >32 ?g/ml; 53.2 to 80.0% susceptible) and colistin (MIC90, 1 ?g/ml; 98.5 to 99.5% susceptible) were comparable to or higher than that of ceftazidime-avibactam. A similar observation was made with isolates that were nonsusceptible to meropenem (n = 1,926), with susceptibility to ceftazidime-avibactam between 67.8% (Middle East/Africa; MIC90, 64 ?g/ml) and 74.2% (Europe; MIC90, 32 ?g/ml) but again with reduced susceptibility to comparators except for amikacin (MIC90, >32 ?g/ml; 56.8 to 78.7% susceptible) and colistin (MIC90, 1 ?g/ml; 98.9 to 99.3% susceptible). Of the 8% of isolates not susceptible to ceftazidime-avibactam, the nonsusceptibility of half could be explained by their possession of genes encoding metallo-?-lactamases. The data reported here are consistent with results from other country-specific and regional surveillance studies and show that ceftazidime-avibactam demonstrates in vitro activity against globally collected clinical isolates of P. aeruginosa, including isolates that are resistant to ceftazidime and meropenem.

Dataset Information

Dynamic Emergence of Mismatch Repair Deficiency Facilitates Rapid Evolution of Ceftazidime-Avibactam Resistance in Pseudomonas aeruginosa Acute Infection.

Publications

Dynamic Emergence of Mismatch Repair Deficiency Facilitates Rapid Evolution of Ceftazidime-Avibactam Resistance in Pseudomonas aeruginosa Acute Infection.

Similar Datasets

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