To Örebro University

Ceftazidime-avibactam and ceftolozane-tazobactam are approved for the treatment of complicated Gram-negative bacterial infections including multidrug-resistant (MDR) Pseudomonas aeruginosa. Resistance to both agents has been reported, but the underlying mechanisms have not been fully explored. This study aimed to correlate β-lactamases with phenotypic resistance to ceftazidime-avibactam and/or ceftolozane-tazobactam in MDR-P. aeruginosa from Qatar. A total of 525 MDR-P. aeruginosa isolates were collected from clinical specimens between 2014 and 2017. Identification and antimicrobial susceptibility were performed by the BD Phoenix^TM system and gradient MIC test strips. Of the 75 sequenced MDR isolates, 35 (47%) were considered as having difficult-to-treat resistance, and 42 were resistant to ceftazidime-avibactam (37, 49.3%), and/or ceftolozane-tazobactam (40, 53.3%). They belonged to 12 sequence types, with ST235 being predominant (38%). Most isolates (97.6%) carried one or more β-lactamase genes, with bla_OXA-488 (19%) and bla_VEB-9 (45.2%) being predominant. A strong association was detected between class B β-lactamase genes and both ceftazidime-avibactam and ceftolozane-tazobactam resistance, while class A genes were associated with ceftolozane-tazobactam resistance. Co-resistance to ceftazidime-avibactam and ceftolozane-tazobactam correlated with the presence of bla_VEB-9, bla_PDC-35, bla_VIM-2, bla_OXA-10 and bla_OXA-488. MDR-P. aeruginosa isolates resistant to both combination drugs were associated with class B β-lactamases (bla_VIM-2) and class D β-lactamases (bla_OXA-10), while ceftolozane-tazobactam resistance was associated with class A (bla_VEB-9), class C (bla_VPDC-35), and class D β-lactamases (bla_OXA-488).

Background: Bloodstream infections (BSIs) caused by multidrug-resistant (MDR)-Pseudomonas aeruginosa are associated with poor clinical outcomes, at least partly due to delayed appropriate antimicrobial therapy. The characteristics of MDR-P. aeruginosa bloodstream isolates have not been evaluated in Qatar. Our study aimed to examine in vitro susceptibility, clinical and molecular characteristics, and mechanisms of resistance of MDR-P. aeruginosa bloodstream isolates from Qatar.

Materials and methods: We included all MDR-P. aeruginosa isolated from blood cultures taken between October 2014 and September 2017. Blood cultures were processed using BD BACTEC™ FX automated system. BD Phoenix™ was used for identification, Liofilchem® MIC Test Strips for MIC determination. Whole-genome sequencing was performed using the Illumina-HiSeq-2000.

Results: Out of 362 P. aeruginosa bloodstream isolates, 16 (4.4%) were MDR. The median patient age was 55 years (range 43-81) and all patients presented with septic shock. Most patients received meropenem (12/16) and/or colistin (10/16). Clinical response was achieved in eight patients, and five patients died within 30-days. MDR-P. aeruginosa isolates belonged to 13 different sequence types. All isolates were non-susceptible to cefepime and ciprofloxacin. The most active agents were colistin (16/16) and aztreonam (10/16). Seven isolates produced blaVIM, and four possessed genes encoding extended-spectrum β-lactamases. Aminoglycoside modifying enzymes were present in 15/16, transferable qnr-mediated quinolone resistance gene was detected in 3/16, and the novel ciprofloxacin modifying enzyme CrpP-encoding gene in one isolate.

Conclusion: MDR-P. aeruginosa BSIs are relatively uncommon in Qatar but are highly resistant, harbour multiple resistance genes, and are commonly associated with unfavourable clinical outcomes. Colistin was the only agent with consistent activity against the study isolates.Key messagesMDR-P. aeruginosa constituted <5% of P. aeruginosa blood isolates over three years.Typical risk factors for MDR infections were highly prevalent in the study population and overall clinical outcomes are consistent with those previously reported.Colistin was the only agent with consistent antibacterial activity against the study isolates.

Background: The emergence of multidrug-resistant Pseudomonas aeruginosa (MDR-PA) is a therapeutic challenge in patients with CF. β-lactamases are the most important mechanism of resistance to β-lactam antibiotics.

Aims: To identify the variability of β-lactamase resistance genes of MDR-PA in sputa of CF patients

Methods: Two hundred forty -six MDR-PA isolates were collected from lower respiratory samples of patients with pulmonary diseases over the 3-year period. Eighteen(7.32%) sputa samples were from 5 CF patients. The identification and antimicrobial susceptibility of the MDR-PA isolates were performed using BD PhoenixTM and E-test in compliance with CLSI guidelines. Whole-genome sequencing was used for gene identifications.

Results: Nine sputum samples were collected from CF patient 1;4 samples from patient 2;2 samples each of patients 3 and 4 and 1 sample from patient 5. The β-lactamase resistant P aeruginosa showed the highest resistance toward cefepime (100%), 55.56% toward each piperacillin-tazobactam and meropenem, and 50% to ceftazidime. β-lactamase genes in MDR-PA were variable within the same CF individual and from patient to patient. All classes of β-lactamase genes were identified (Class A; SHV-11, VEB-9, TEM-87, TEM-116, TEM-126. Class B;VIM-2. Class C;PDC-1, PDC-3, PDC-7, PDC-8. Class D;OXA-10, OXA-50, OXA-114a) with the highest percentage for OXA-50 harbored in 61.11% of isolates

Conclusions: Variability of β-lactamase genes in P aeruginosa isolates from CF patients may affect the progression of CF and personal specific antibiotics response to such infection

Background: The excessive and inappropriate use of antibiotics is universal across all healthcare facilities. In Qatar there has been a substantial increase in antimicrobial consumption coupled with a significant rise in antimicrobial resistance (AMR). Antimicrobial stewardship programmes (ASPs) have become a standard intervention for effective optimization of antimicrobial prescribing.

Methods: A before–after study was conducted in Hamad General Hospital (603 bed acute care hospital): 1 year before implementation of a comprehensive ASP compared with the following 2 years. The ASP included a hospital-wide pre-authorization requirement by infectious diseases physicians for all broad-spectrum antibiotics. Prevalence of MDR Pseudomonas aeruginosa was compared with antimicrobial consumption, calculated as DDD per 1000 patient-days (DDD/1000 PD). Susceptibility was determined using broth microdilution, as per CLSI guidelines. Antibiotic use was restricted through the ASP, as defined in the hospital’s antibiotic policy.

Results: A total of 6501 clinical isolates of P. aeruginosa were collected prospectively over 3 years (2014–17). Susceptibility to certain antimicrobials improved after the ASP was implemented in August 2015. The prevalence of MDR P. aeruginosa showed a sustained decrease from 2014 (9%) to 2017 (5.46%) (P"0.019). There was a significant 23.9% reduction in studied antimicrobial consumption following ASP implementation (P"0.008). They early consumption of meropenem significantly decreased from 47.32 to 31.90 DDD/1000 PD (P"0.012), piperacillin/tazobactam from 45.35 to 32.67 DDD/1000 PD (P,0.001) and ciprofloxacin from 9.71 to 5.63 DDD/1000 PD (P"0.015) (from 2014 to 2017).

Conclusions: The successful implementation of the ASP led to a significant reduction in rates of MDR P. aeruginosa, pointing towards the efficacy of the ASP in reducing AMR.

Inappropriate and excessive use of antibiotics promotes antimicrobial resistance (AMR), particularly in Gram-negative bacteria (GNB). There is a noticeable increase in nosocomial infections caused by multidrug-resistant (MDR) Pseudomonas aeruginosa, which is associated with significant morbidity, mortality, and an increase in cost management. Although this is a global problem, there is a lack of sufficient data on regional differences that can contribute towards effective AMR management. This thesis presents a study of MDR-P. aeruginosa at five different hospitals in Qatar conducted prospectively between October 2014 - September 2017. The aim was to study the epidemiology, microbiological and clinical characteristics of MDR-P. aeruginosa infections as well as investigate the activity of new antibiotic combinations against these bacteria. The prevalence of MDR-P. aeruginosa isolates in the first year was 8.1% (205/2533), isolated from different clinical specimens, but the majority were from respiratory infections (44.9%, n=92). Most cases were exposed to antibiotics during the 90 days prior to isolation (85.4%, n=177), and the resistance to cefepime, ciprofloxacin, piperacillin/tazobactam, meropenem was >90%. To compare pre- and post-Antimicrobial Stewardship Program, there was a significant reduction in antibiotic consumption by 30.4% of total inpatient antibiotic prescriptions (p=0.008) and the prevalence of MDR-P. aeruginosa significantly declined from 9% to 5.4% (p=0.019). The in vitro investigation of ceftazidime/avibactam (CZA) and ceftolozane/tazobactam (C/T) against MDR-P. aeruginosa isolates, showed promising results with susceptibility of 68.8% (n=141/205) and 62.9% (n=129/205), respectively, which was higher than other antipseudomonal agents except colistin. Seventy-five isolates that were sequenced belonged to 29 different sequence types, with ST235 being predominant at 21.3% (16/75). Among the 42 isolates that were resistant to CZA and/or C/T, the most prevalent genes were blaOXA-488 and blaVEB-9 detected in 45.2% (19/42) of isolates. Spearman’s analysis showed that resistance to CZA and C/T were positively correlated with the presence of blaOXA-10, blaPDC-2a, blaVIM-2, and blaVEB-9 , respectively. The study highlights potential key mechanisms that could explain the resistance of MDR-P. aeruginosa to the new antibiotic combinations.

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: A global rise in multidrug-resistant (MDR) nosocomial infections has led to a significant increase in morbidity and mortality. MDR Gram-negative bacteria (GNB) are recognised for rapidly developing drug resistance. Despite Pseudomonas aeruginosa being the second most common GNB isolated from healthcare associated infections, the magnitude of MDR P. aeruginosa (MDR-PA) has not been evaluated in Qatar.

Aim: To assess the prevalence and antimicrobial susceptibility patterns of MDR-PA from 5major hospitals in Qatar.

Methods: A total of 2533P. aeruginosaclinical isolates were collected over a one-year period. MDR-PA was defined as resistance to at least one agent of3 antibiotic classes. Clinical and demographic data were collected prospectively.

Findings: The overall prevalence of MDR-PA isolates was 8.1% (205/2533); the majority of isolates were from patients exposed to antibiotics during 90 days prior to isolation (85.4%,177/205), and the infections were mainly hospital-acquired (95.1%, 195/205) with only 4.9% from the community. The majority of MDR-PA isolates were resistant to cefepime (96.6%, 198/205), ciprofloxacin, piperacillin/tazobactam (91%, 186/205), and meropenem (90%, 184/205). Patient comorbidities with MDR-PA were diabetes mellitus (47.3%, n¼97), malignancy (17.1%, n¼35), end-stage renal disease (13.7%, n¼28) and heart failure (10.7%, n¼22).

Conclusion: There was a significant prevalence of MDR-PA in Qatar, primarily from healthcare facilities and associated with prior antibiotic treatment. There was an alarming level of antimicrobial resistance to carbapenems. Our results are part of a national surveillance of MDR to establish effective containment plans.

OBJECTIVES: To investigate the in vitro activity of ceftazidime/avibactam and ceftolozane/tazobactam against clinical isolates of MDR Pseudomonas aeruginosa from Qatar, as well as the mechanisms of resistance.

METHODS: MDR P. aeruginosa isolated between October 2014 and September 2015 from all public hospitals in Qatar were included. The BD PhoenixTM system was used for identification and initial antimicrobial susceptibility testing, while Liofilchem MIC Test Strips (Liofilchem, Roseto degli Abruzzi, Italy) were used for confirmation of ceftazidime/avibactam and ceftolozane/tazobactam susceptibility. Ten ceftazidime/avibactam- and/or ceftolozane/tazobactam-resistant isolates were randomly selected for WGS.

RESULTS: A total of 205 MDR P. aeruginosa isolates were included. Of these, 141 (68.8%) were susceptible to ceftazidime/avibactam, 129 (62.9%) were susceptible to ceftolozane/tazobactam, 121 (59.0%) were susceptible to both and 56 (27.3%) were susceptible to neither. Twenty (9.8%) isolates were susceptible to ceftazidime/avibactam but not to ceftolozane/tazobactam and only 8 (3.9%) were susceptible to ceftolozane/tazobactam but not to ceftazidime/avibactam. Less than 50% of XDR isolates were susceptible to ceftazidime/avibactam or ceftolozane/tazobactam. The 10 sequenced isolates belonged to six different STs and all produced AmpC and OXA enzymes; 5 (50%) produced ESBL and 4 (40%) produced VIM enzymes.

CONCLUSIONS: MDR P. aeruginosa susceptibility rates to ceftazidime/avibactam and ceftolozane/tazobactam were higher than those to all existing antipseudomonal agents, except colistin, but were less than 50% in extremely resistant isolates. Non-susceptibility to ceftazidime/avibactam and ceftolozane/tazobactam was largely due to the production of ESBL and VIM enzymes. Ceftazidime/avibactam and ceftolozane/tazobactam are possible options for some patients with MDR P. aeruginosa in Qatar.

Resistance to last-line polymyxins mediated by the plasmid-borne mobile colistin resistance gene (mcr-1) represents a new threat to global human health. Here we present the complete genome sequence of an mcr-1-positive multidrug-resistant Escherichia coli strain (MS8345). We show that MS8345 belongs to serotype O2:K1:H4, has a large 241,164-bp IncHI2 plasmid that carries 15 other antibiotic resistance genes (including the extended-spectrum β-lactamase bla_CTX-M-1) and 3 putative multidrug efflux systems, and contains 14 chromosomally encoded antibiotic resistance genes. MS8345 also carries a large ColV-like virulence plasmid that has been associated with E. coli bacteremia. Whole-genome phylogeny revealed that MS8345 clusters within a discrete clade in the sequence type 95 (ST95) lineage, and MS8345 is very closely related to the highly virulent O45:K1:H4 clone associated with neonatal meningitis. Overall, the acquisition of a plasmid carrying resistance to colistin and multiple other antibiotics in this virulent E. coli lineage is concerning and might herald an era where the empirical treatment of ST95 infections becomes increasingly more difficult.

Importance: Escherichia coli ST95 is a globally disseminated clone frequently associated with bloodstream infections and neonatal meningitis. However, the ST95 lineage is defined by low levels of drug resistance amongst clinical isolates, which normally provides for uncomplicated treatment options. Here, we provide the first detailed genomic analysis of an E. coli ST95 isolate that has both high virulence potential and resistance to multiple antibiotics. Using the genome, we predicted its virulence and antibiotic resistance mechanisms, which include resistance to last-line antibiotics mediated by the plasmid-borne mcr-1 gene. Finding an ST95 isolate resistant to nearly all antibiotics that also has a high virulence potential is of major clinical importance and underscores the need to monitor new and emerging trends in antibiotic resistance development in this important global lineage.

Introduction: Multidrug-resistant Pseudomonas aeruginosa (MDR-PA) is an important and growing issue in the care of patients with cystic fibrosis (CF), and a major cause of morbidity and mortality.

Objective: The objective of the study was to describe the frequency of MDR-PA recovered from the lower respiratory samples of pediatric and adult CF patients, and its antibiotic resistance pattern to commonly used antimicrobial agents including beta-lactams, aminoglycosides, and fluoroquinolones.

Materials and Methods: The lower respiratory isolates of P. aeruginosa were obtained from inpatients and outpatients CF clinics from a tertiary care teaching hospital for the period from October 2014 to September 2015. The identification and antimicrobial susceptibility for all the isolates were performed by using the BD Phoenix (TM) and E-test in compliance with Clinical and Laboratory Standards Institute (CLSI) guidelines.

Results: A total of 61 P. aeruginosa samples were isolated from thirty CF patients from twenty families. Twelve sputum samples were positive for MDR-PA (seven nonmucoid and five mucoid isolates) from five CF patients (five families) with moderate-to-very severe lung disease given MDR-PA frequency of 19.7%. The median age of the study group was 20 (range 10-30) years. Three CF patients were on chronic inhaled tobramycin and two on nebulized colistin. The antimicrobial patterns of isolates MDR-PA showed the highest rate of resistance toward each gentamycin, amikacin, and cefepime (100%), followed by 91.7% to ciprofloxacin, 75% to tobramycin, 58.3% to meropenem, and 50% to piperacillin-tazobactam. None of the isolates were resistant to colistin during the study period.

Conclusion: The study results emphasize that the emergence of a significant problem in the clinical isolates of P. aeruginosa in CF patients that dictate appropriate attention to the antibiotic management after proper surveillance.