Ceftazidime-avibactam (CAZ-AVI) is a promising novel treatment for infections caused by carbapenem-resistant (CRE). Despite improved treatment outcomes compared to those achieved with aminoglycoside- and colistin-based regimens, the rapid evolution of CAZ-AVI resistance during treatment has previously been reported in sequence type 258 (ST258) -harboring isolates. Here, we report the stepwise evolution and isolation of two phenotypically distinct CAZ-AVI-resistant isolates from a patient with pancreatitis. All susceptible ( = 3) and resistant ( = 5) isolates were of the ST307 clonal background, a rapidly emerging clone. Taking advantage of short-read Illumina and long-read Oxford Nanopore sequencing and full-length assembly of the core chromosome and plasmids, we demonstrate that CAZ-AVI resistance first occurred through a 532G → T point mutation in (D179Y protein substitution) following only 12 days of CAZ-AVI exposure. While subsequent isolates exhibited substantially decreased meropenem (MEM) MICs (≤2 μg/ml), later cultures demonstrated a second CAZ-AVI resistance phenotype with a lower CAZ-AVI MIC (12 μg/ml) but also MEM resistance (MIC > 128 μg/ml). These CAZ-AVI- and MEM-resistant isolates showed evidence of multiple genomic adaptations, mainly through insertions and deletions. This included amplification and transposition of wild-type into a novel plasmid, an IS insertion upstream of , and disruption of the gene locus in these isolates. Our findings illustrate the potential of CAZ-AVI resistance to emerge in non- ST258 clonal backgrounds and alternative variants. These results raise concerns about the strong selective pressures incurred by novel carbapenemase inhibitors, such as avibactam, on isolates previously considered invulnerable to CAZ-AVI resistance. There is an urgent need to further characterize non-KPC-mediated modes of carbapenem resistance and the intrinsic bacterial factors that facilitate the rapid emergence of resistance during treatment.

BackgroundCarbapenem-resistant Enterobacteriaceae (CRE) have emerged as an urgent public health threat. Intestinal colonization with CRE has been identified as a risk factor for the development of systemic CRE infection, but has not been compared to colonization with third and/or fourth generation cephalosporin-resistant (Ceph-R) Enterobacteriaceae. Moreover, the risk conferred by colonization on adverse outcomes is less clear, particularly in critically ill patients admitted to the intensive care unit (ICU).MethodsWe carried out a cohort study of consecutive adult patients screened for rectal colonization with CRE or Ceph-R upon ICU entry between April and July 2013. We identified clinical variables and assessed the relationship between CRE or Ceph-R colonization and subsequent systemic CRE infection within 30 days (primary outcome) and all-cause mortality within 90 days (secondary outcome).ResultsAmong 338 ICU patients, 94 (28%) were colonized with either Ceph-R or CRE. 26 patients developed CRE infection within 30 days of swab collection; 47% (N = 17/36) of CRE-colonized and 3% (N = 2/58) of Ceph-R colonized patients. 36% (N = 13/36) of CRE-colonized patients died within 90 days compared to 31% (N = 18/58) of Ceph-R-colonized and 15% (N = 37/244) of non-colonized patients. In a multivariable analysis, CRE colonization independently predicted development of a systemic CRE infection at 30 days (aOR 10.8, 95% CI2.8–41.9, p = 0.0006); Ceph-R colonization did not (aOR 0.5, 95% CI0.1–3.3, p = 0.5). CRE colonization was associated with increased 90-day mortality in a univariable analysis (p-value 0.001), in a multivariable model, previous hospitalization and medical ICU admission were independent predictors of 90-day mortality whereas CRE colonization approached significance (aOR 2.3, 95% CI1.0–5.3, p = 0.056).ConclusionsOur study highlights the increased risk of CRE infection and mortality in patients with CRE colonization at the time of ICU admission. Future studies are needed to assess how CRE colonization can guide empiric antibiotic choices and to develop novel decolonization strategies.

Background Patients hospitalized with COVID-19 are at increased risk of healthcare-associated infections especially with prolonged hospital stays. We sought to identify incidence, antimicrobial susceptibilities, and outcomes associated with bacterial/fungal secondary infections in a large cohort of patients with COVID-19. Methods We evaluated adult patients diagnosed with COVID-19 between March 2 to May 31, 2020 and hospitalized >24 hours. Data extracted from medical records included diagnoses, vital signs, laboratory results, microbiological data, and antibiotic use. Microbiologically-confirmed bacterial and fungal pathogens from clinical cultures were evaluated to characterize community- and healthcare-associated infections, including describing temporal changes in predominant organisms on presentation and throughout hospitalization. Univariable and multivariable logistic regression analyses were performed to investigate risk factors for healthcare-associated infections. Results A total of 3,028 patients were included and accounted for 899 positive clinical cultures. Overall, 516 (17%) with positive cultures met criteria for infection. Community-associated co-infections were identified in 183 (6%) patients, whereas healthcare-associated infections occurred in 350 (12%) patients. 57% of healthcare-associated infections were caused by Gram-negative bacteria and 19% by fungi. Antibiotic resistance increased with longer hospital stays, with incremental increases in the proportion of vancomycin-resistance among enterococci and ceftriaxone- and carbapenem-resistance among Enterobacterales. ICU stay, invasive mechanical ventilation, and steroids were associated with healthcare-associated infections. Conclusions Healthcare-associated infections occur in a small proportion of patients hospitalized with COVID-19 and are most often caused by Gram-negative and fungal pathogens. Antibiotic resistance is more prevalent with prolonged hospital stays. Antimicrobial stewardship is imperative in this population to minimize unnecessary broad-spectrum antibiotics.

Background Patients with COVID-19 may be at increased risk for secondary bacterial infections with MDR pathogens, including carbapenemase-producing Enterobacterales (CPE). Objectives We sought to rapidly investigate the clinical characteristics, population structure and mechanisms of resistance of CPE causing secondary infections in patients with COVID-19. Methods We retrospectively identified CPE clinical isolates collected from patients testing positive for SARS-CoV-2 between March and April 2020 at our medical centre in New York City. Available isolates underwent nanopore sequencing for rapid genotyping, antibiotic resistance gene detection and phylogenetic analysis. Results We identified 31 CPE isolates from 13 patients, including 27 Klebsiella pneumoniae and 4 Enterobacter cloacae complex isolates. Most patients (11/13) had a positive respiratory culture and 7/13 developed bacteraemia; treatment failure was common. Twenty isolates were available for WGS. Most K. pneumoniae (16/17) belonged to ST258 and encoded KPC (15 KPC-2; 1 KPC-3); one ST70 isolate encoded KPC-2. E. cloacae isolates belonged to ST270 and encoded NDM-1. Nanopore sequencing enabled identification of at least four distinct ST258 lineages in COVID-19 patients, which were validated by Illumina sequencing data. Conclusions While CPE prevalence has declined substantially in New York City in recent years, increased detection in patients with COVID-19 may signal a re-emergence of these highly resistant pathogens in the wake of the global pandemic. Increased surveillance and antimicrobial stewardship efforts, as well as identification of optimal treatment approaches for CPE, will be needed to mitigate their future impact.

Background Polymyxins are antimicrobials of last resort for the treatment of carbapenem-resistant Enterobacteriaceae, but resistance in 5% to >40% isolates has been reported. We conducted a genomic survey of clinical polymyxin-resistant (PR) Klebsiella pneumoniae to determine the molecular mechanisms of PR and the role of polymyxin exposure versus transmission in PR emergence. Methods We included 88 patients with PR K. pneumoniae from 2011–2018 and collected demographic, antimicrobial exposure, and infection data. Whole-genome sequencing was performed on 388 isolates, including 164 PR isolates. Variant calling and insertion sequence detection were performed, focusing on key genes associated with PR (mgrB, crrAB, phoPQ, and pmrAB). We conducted phylogenetic analyses of key K. pneumoniae multi-locus sequence types (ST258, ST17, ST307, and ST392). Results Polymyxin exposure was documented in 53/88 (60%) patients prior to PR detection. Through an analysis of key PR genes, we detected 129 individual variants and 72 unique variant combinations in PR isolates. This included multiple, distinct changes in 36% of patients with serial PR isolates. Insertion sequence disruption was limited to mgrB (P < .001). Polymyxin minimum inhibitory concentrations showed stepwise increases with the number of PR genes affected (P < .001). When clusters containing PR isolates in ≥2 patients were analyzed, 10/14 had multiple genetic events leading to PR. Conclusions Molecular mechanisms leading to PR in clinical K. pneumoniae isolates are remarkably heterogenous, even within clusters or individual patients. Polymyxin exposure with de novo PR emergence led to PR in the majority of patients, rather than transmission. Optimizing polymyxin use should be a key strategy in stopping the spread of PR.