β-lactamase-mediated resistance in MDR-Pseudomonas aeruginosa from Qatar

Background The distribution of β-lactam resistance genes in P. aeruginosa is often closely related to the distribution of certain high-risk international clones. We used whole-genome sequencing (WGS) to identify the predominant sequence types (ST) and β-lactamase genes in clinical isolates of multidrug-resistant (MDR)-P. aeruginosa from Qatar Methods Microbiological identification and susceptibility tests were performed by automated BD Phoenix™ system and manual Liofilchem MIC Test Strips. Results Among 75 MDR-P. aeruginosa isolates; the largest proportions of susceptibility were to ceftazidime-avibactam (n = 36, 48%), followed by ceftolozane-tazobactam (30, 40%), ceftazidime (n = 21, 28%) and aztreonam (n = 16, 21.3%). All isolates possessed Class C and/or Class D β-lactamases (n = 72, 96% each), while metallo-β-lactamases were detected in 20 (26.7%) isolates. Eight (40%) metallo-β-lactamase producers were susceptible to aztreonam and did not produce any concomitant extended-spectrum β-lactamases. High risk ST235 (n = 16, 21.3%), ST357 (n = 8, 10.7%), ST389 and ST1284 (6, 8% each) were most frequent. Nearly all ST235 isolates (15/16; 93.8%) were resistant to all tested β-lactams. Conclusion MDR-P. aeruginosa isolates from Qatar are highly resistant to antipseudomonal β-lactams. High-risk STs are predominant in Qatar and their associated MDR phenotypes are a cause for considerable concern.


Background
Due to their established efficacy and safety, anti-pseudomonal β-lactam antibiotics play a vital role in the clinical management of P. aeruginosa infections [1]. Key antimicrobial resistance mechanisms in P. aeruginosa include over-expression of efflux pumps, impermeability through porin modification or loss, target modification, and enzyme-mediated antimicrobial inactivation (e.g., β-lactamases). Multiple resistance mechanisms are frequently present in concert resulting in simultaneous resistance to multiple agents [1]. The epidemiology of β-lactamases is often closely related to the distribution of certain high-risk international clones [2]. In this study, we used whole-genome sequencing (WGS) to identify the predominant sequence types (STs) and β-lactamase genes in multi-drug resistant (MDR) P. aeruginosa clinical isolates from Qatar.

Methods
The study setting, bacterial identification, antimicrobial susceptibility testing, whole genome sequencing, and statistical analysis details are provided in Additional File 1. MDR status was defined as in-vitro resistance to at least one agent from three or more classes of anti-pseudomonal agents [3]. β-lactamases were classified according to their molecular groups [4]. Clinical data were retrieved from the electronic healthcare system.

Discussion
This study included data representative of the whole country, as it analyzed isolates from a national diagnostic laboratory. Notably, MDR-P. aeruginosa in Qatar are highly resistant to β-lactam agents. The most active β-lactam antibiotics in this study were those in combination with β-lactamase inhibitors, ceftazidime-avibactam and ceftolozane-tazobactam, were not available for clinical use at the time of the study. Yet, less than half of the isolates were susceptible. Given their recent availability for patients in Qatar, the results reported demonstrate the importance of their appropriate clinical use to minimize further loss of activity [5].
An interesting observation in this study was that 16 (21.33%) MDR-P. aeruginosa isolates were susceptible to aztreonam but resistant to several other antipseudomonal β-lactams tested (Additional file 1: Table S1). Aztreonam is a weak inducer of Class C enzymes and is not a substrate for Class B and narrowspectrum Class D β-lactamases [17]. The retained aztreonam activity in these isolates despite resistance to other antipseudomonal β-lactams may be explained by the absence of Class A ESBL in those isolates. Therefore, aztreonam should be included in routine antimicrobial susceptibility testing of clinical P. aeruginosa isolates.
Most MDR-P. aeruginosa isolates included in this study belonged to five STs and had consistent β-lactamase genetic profiles and β-lactam susceptibility patterns (Additional file 1: Table S2). ST235, ST233, and ST357 are already known as high-risk clones in Qatar, Saudi Arabia, Bahrain, and the United Arab Emirates [7]. These three STs are globally disseminated MDR-P. aeruginosa clones [2]. Often, these strains cause regional or nationwide outbreaks, express MDR phenotypes, and are associated with high mortality [12,18,19]. VIM-producing ST1284 P. aeruginosa have been described from Brazil, and ST389 from cystic fibrosis patients in Italy [20,21]. Both sequence types have otherwise limited geographic distribution.

Conclusion
MDR-P. aeruginosa isolates from Qatar are highly resistant to antipseudomonal β-lactams. Global high-risk STs predominate in Qatar and their associated multi-resistant phenotype is a cause for considerable concern.