Retrospective Tertiary Care-Based Cohort Study on Clinical Characteristics and Outcomes of Ceftazidime-Avibactam-Resistant Carbapenem-Resistant Klebsiella pneumoniae Infections

Introduction The advent of ceftazidime-avibactam (CAZ-AVI)-resistant carbapenem-resistant Klebsiella pneumoniae (CRKP) isolates has been steadily documented in recent years. We aimed to identify risk factors of CAZ-AVI-resistant CRKP infection and assess clinical outcomes of patients. Methods The study retrospectively examined the clinical and microbiological data of patients with ceftazidime avibactam susceptible and ceftazidime avibactam-resistant Klebsiella pneumonia carbapenem-resistant enterobacteriaceae infection to identify risk factors, clinical features, and outcomes using multivariate logistic regression analysis. Results A total of 152 patients with CRKP infection were enrolled in this study. Patients with CAZ-AVI-resistant CRKP isolates (20/34 = 58.8%) had prior exposure to carbapenems (p=0.003) and had more tracheostomies (16/34 = 47.1%) (p=0.001). Only 8/28 (28.6%) patients with CAZ-AVI susceptible CRKP isolates died amongst those administered ceftazidime-avibactam compared to 49/90 (54.4%) who did not receive the same (p=0.016). 1/9 (11.1%) patients with CAZ-AVI-resistant CRKP isolates who received colistin died compared to 13/25 (52%) who did not receive colistin (p=0.03). There was no association between presence of CAZ-AVI-resistant CRKP isolates and overall mortality (odds ratio: 0.7; 95% CI: 0.3, 1.6), and no independent predictors of risk factors to overall mortality in the group with CAZ-AVI-resistant CRKP isolates were noted. Conclusion Early advent of CAZ-AVI resistance in CRE isolates highlights the dynamic necessity of routine CAZ-AVI resistance laboratory testing and antimicrobial stewardship programmes focusing on the utilization of all antibiotics. Consolidating the hospital infection control of tracheostomies may help to prevent CAZ resistance in CRKP. Colistin may aid in decreasing of mortality rates among patients with CAZ-AVI CRKP isolates.

CAZ-AVI is recommended for the treatment of following infections in adults, i.e., complicated urinary tract infection (cUTI), complicated intra-abdominal infection (cIAI), including pyelonephritis, and hospital-acquired pneumonia (HAP) including ventilator-associated pneumonia (VAP) with susceptible Gram-negative microorganism and treatment of patients with bacteremia that occurs in association with or is suspected to be associated with any of the infections mentioned above [3].
Te in vitro susceptibility of carbapenemase-positive and MBL-negative isolates to CAZ-AVI was 99.8% between 2015 and 2017 in the International Network for Optimal Resistance Monitoring (INFORM) surveillance programme [1,9].Te in vitro susceptibility of CR K. pneumoniae against ceftazidime avibactam was reported to be 85.6% using the Clinical Laboratory and Standards Institute (CLSI) breakpoints according to the global ATLAS data collected between 2012 and 2016 [3,10].
CAZ-AVI-containing regimen notably increased 7-day microbiological clearance of CPKP infections and 28-day clinical success rate and diminished 28-day mortality rate compared with polymyxin B [11,12].
Unfortunately, CAZ-AVI-resistant CRKP infections have been reported escalating in adult patients with the inception of CAZ-AVI therapy in clinic [1,13,14].
Te paucity of sufcient source control alongside a prolonged antibiotic regimen (mean duration of 17 days) may advance evolution of resistance to CAZ-AVI [19].Patient-to-patient transmission may be a considerable factor in CAZ-AVI resistance in CRKP isolates as intestinal carriage of carbapenemase is an important source of transmission [13,20].Sparse rates of CAZ-AVIresistant isolates have been retrospectively documented in strains isolated before the advent of CAZ-AVI in clinical practice, even in neonates [13], constituting a reservoir of resistance, that could be potentially enhanced under inappropriate CAZ-AVI-based treatment [21].Resistance to CAZ-AVI can also emerge from selection pressure during treatment with meropenem emphasizing the need for dynamic antimicrobial stewardship programs for antibiotic utilization [22].Renal replacement therapy (RRT), pneumonia, and mechanical ventilation are shown to be risk factors for the development of CAZ-AVI resistance in a few studies [23,24].

Methodology
Tis study is a retrospective comparative analysis of clinical outcomes and possible risk factors of patients with ceftazidime avibactam susceptible and ceftazidime avibactamresistant Klebsiella pneumonia carbapenem-resistant enterobacteriaceae infection.Ethical clearance from research approval No: 8170122 was obtained from the Research Committee for Government Hospitals and need for consent was waived out as per institutional guidelines for retrospective studies.

Study Population. All clinically established ceftazidime avibactam susceptible and ceftazidime avibactam-resistant
Klebsiella pneumonia CRE infected hospitalized patients (Age >14 years) in Salmaniya Medical Complex, Bahrain, were included from January 2020 to December 2021 at the time of the most recently treated CRE infection.Patients with bacteremia will be analyzed as such regardless of the primary source.

Defnitions of Infection
(i) A patient with a culture positive for K. pneumoniae CRE was deemed to have an infection if K. pneumoniae CRE was isolated from blood or any other sterile source.(ii) Patients with bacteremia will be analyzed as such regardless of the primary source.(iii) For patients with positive respiratory cultures, the criteria outlined by the American Toracic Society and the Infectious Diseases Society of America will be used [25].(iv) For patients with positive cultures from urine or surgical wounds, the CDC National Healthcare Safety Network criteria will be applied [26].(v) Patients with cultures from nonsurgical wounds will be considered infected only if the treating physician documented infection in the medical record and evidence of systemic infammation on the day the positive culture was documented; this was defned as an abnormal systemic white cell count (either >10 × 10 3 or <4 × 10 3 cells/μl) and/or an abnormal body temperature (either >99.5 °F or <96 °F).(vi) All of the other culture episodes were designated colonizations.
2 Critical Care Research and Practice (vii) CRE was defned by the current centers for Disease Control and Prevention criteria as resistance to any carbapenem [27].

Methods
Te data were collected from the medical e-record database and medical records.Te data collected are enlisted in Table 1 and methods followed by van Duin et al. were utilized [28].A standard dosage of ceftazidime avibactam 2.5 g intravenously (IV) every 8 hours was used, with adjustments for renal impairment made according to manufacturer recommendation [29].All combination therapy antibiotic agents were started concomitantly with ceftazidime avibactam and administered for 72 hours or longer as per recommendations of antimicrobial stewardship of the infectious disease team.Previous antibiotic use was recorded as the class of antibiotic with documented exposure in 30 days leading up to the day of the frst positive culture.
Infections were defned as nosocomial if the frst K. pneumoniae CRE-positive culture was obtained >48 hours after hospital admission.Healthcare-associated infections were defned as infections in patients admitted from longterm care facilities or other hospitals who had a frst CR K. pneumoniae-positive culture within the frst 48 hours.Patients with a frst K. pneumoniae-CRE positive culture within 48 hours who were admitted from home were be deemed to have community-acquired infections.Renal insufciency is defned as any serum creatinine level of ≥2 mg/ dl.If patients were intubated and sedated and no further information on mental status was available, the mental status was coded as "stuporous" for the purpose of calculating this score only.Te Pitt bacteremia score (validated as a predictor of the outcome of K. pneumoniae CRE bloodstream infection) based on the day of the index culture was calculated, and patients with a score ≥4 will be considered critically ill.Te Charlson comorbidity index was determined at admission.Clearance was defned as survival and absence of recurrence at 14 days following the onset of infection, resolution of signs and symptoms of infection, and sterilization of site-specifc cultures within 14 days of treatment initiation.Antibiotic treatment initiated in ceftazidime avibactam-resistant group of patients will be recorded.Primary outcome of 30 day mortality was recorded.

Microbiology.
Guidelines from the Clinical and Laboratory Standards Institute will be used to defne CRE.Bacterial species-level identifcation was confrmed by using a mass spectrometry system (MALDI-TOF Bruker Daltonik GmbH, Bremen, Germany), and antibiotic susceptibility testing of isolates was performed with automated microbiological systems (VITEK-2 compact bioMerieux, Marcy L, Etoile, France).Mechanisms of resistance in CRE isolates using Xpert Carba-R were recorded.Minimum inhibitory concentrations (MICs) were determined using the reference Clinical and Laboratory Standards Institute broth microdilution methods; avibactam was tested at a fxed concentration of 4 μg/mL [30].

Statistical Analysis.
To detect signifcant diferences between groups, we use the chi-square test or the Fisher exact test for categorical variables and the two-tailed t-test or Mann-Whitney test for continuous variables, when appropriate.In a multivariate analysis of survival, the Cox regression model will be used to determine efects of diferent variables on 30-day survival.Wald confdence intervals and tests for odds ratios will be computed on the basis of the estimated standard errors.Possible confounding factors and interactions will be weighted during analyses.Statistical signifcance will be established at p ≤ 0.05.All reported p values are two tailed.Patients with CAZ-AVI-resistant CRKP isolates (20/ 34 � 58.8%) had prior exposure to carbapenems compared to patients with CAZ-AVI susceptible CRKP isolates which was statistically signifcant (p � 0.003).
In the group CAZ-AVI susceptible CRKP, (13/ 37 � 35.1%) with lower respiratory tract infection died compared to (44/81 � 54.3%) of other sites, and this difference of lower mortality was statistically signifcant (p � 0.05).In the group with CAZ-AVI-resistant CRKP isolates, no signifcant diferences were observed between any of the sites (blood stream/urine/DTA) and mortality.
Tere were no signifcant diferences noted in the Pitt bacteremia score, Charlson comorbidity index, SAPS 2 score, APACHE 2 score, days of ICU stay, clearance of infection, steroid use prior to index culture, time from admission to index culture, and mortality between both groups.
Tere is no association between presence of CAZ-AVIresistant CRKP isolates and overall mortality (odds ratio: 0.7; 95% CI: 0.3, 1.6), and no independent predictors of risk factors to overall mortality in the group with CAZ-AVIresistant CRKP isolates were noted.
Clearance rates were nearly identical among patients receiving ceftazidime-avibactam monotherapy or combination therapy.

Discussion
In this study, patients with CAZ-AVI-resistant CRKP isolates (6/34 = 17.6%) had prior Klebsiella pneumoniae CRE infections more compared to patients with CAZ-AVIsensitive CRKP isolates (6/118 = 5.1%) patients which was statistically signifcant (p � 0.03).Tis was similar to an earlier reported study where almost all the involved resistant bacteria were K. pneumoniae strains (64-87.8%)and twothirds of CAZ-AVI-resistant bacteria were isolated from patients with prior exposure to CAZ-AVI [31].
Only (13/37 � 35.1%) with lower respiratory tract infections (LRTI) infection died in patients with CAZ-AVIsusceptible CRKP isolates compared to (44/81 � 54.3%) of other sites, and this diference of lower mortality was statistically signifcant (p � 0.05) which was not noted in patients with CAZ-AVI-resistant isolates.Conficting evidences have been inferred regarding CAZ-AVI reaching adequate [32]/inadequate [33][34][35] concentrations in the airway epithelial lining fuid.Tis is in contrast to other cohorts where mortality was signifcantly higher among patients with LRTI [2] but had a higher severity of infections with INCREMENT scores of ≥8 compared to other subgroups [12].A few clinical studies revealed that the recommended therapeutic dose of CAZ-AVI therapy for KPC-K.pneumoniae was not as favorable for bacteremia [12,33,36].Pneumonia has shown to be an independent risk factor for CAZ-AVI treatment failure and resistance among patients with CRE infection [36].
In this study, there was no diference in comorbidities rate, Pitt bacteremia score, Charlson comorbidity index, SAPS 2 score, APACHE 2 score, days of ICU stay, clearance of infection, steroid use prior to index culture, time from admission to index culture, and mortality between both the population.Te extensive subjection to antimicrobials in patients with signifcant comorbidities increases the propensity of CAZ-AVI-resistant CRKP isolates [14,19].In previously reported studies, the overall mortality (37%) in patients with comorbidities (cancer, transplantation, and cardiomyopathy) is more than the mortality rate of patients with CAZ-AVI-resistant systemic infections (10%), thereby increasing the overall mortality rate accountable to CAZ-AVI-resistant isolates [16,21,40].
CAZ-AVI-resistant patients represented new cases of resistance originating from home at the time of hospital admission (23/34 � 67.6%), but all patients were noted to have prior history of hospitalizations more than 90 days ago.Low rates of CAZ-AVI-resistant CRKP isolates have been  Critical Care Research and Practice reported ex post facto in strains isolated prior to CAZ-AVI in clinical practice representing a reservoir of resistance [21] even in neonates [13].Patient-to-patient transmission may be a crucial factor in CAZ-AVI-resistant CRKP isolates due to intestinal carriage of carbapenemase [20] that could be potentially accentuated under inappropriate CAZ-AVIbased treatment [21].
Usually, CAZ-AVI-resistant CRKP isolates emerged in patients after 10-19 days of CAZ/AVI treatment.Suboptimal drug exposure to CAZ-AVI may lead to the emergence of CAZ-AVI-resistant KPC-K.pneumoniae strains in critically ill patients [41][42][43].85% of CAZ-AVI-resistant strains related to infections were commonly treated with combination therapy of CAZ-AVI with meropenem as the most common antibiotic used (65% of cases) [16,21].In our study, no emergence of resistance to CAZ-AVI was reported in patients who survived.
Te clearance rate and mortality rates were nearly identical among patients receiving ceftazidime-avibactam monotherapy or combination therapy in susceptible patients.It is suggested by in vitro studies that combination therapies may achieve higher rates of success, allow lower doses or shorter treatment periods, and impede the advent of resistance, but with a risk of possible side efects, inception of a higher level of resistance than already known with conficting evidence [44].
No independent predictors of risk factors to overall mortality in patients with CAZ-AVI-resistant CRKP isolates were noted.Data on ventilation days and continuous replacement therapy were not captured but previous studies have shown pneumonia and continuous renal replacement therapy as risk factors for CAZ-AVI treatment failure and resistance among patients with CRE infection due to potential suboptimal drug concentrations at the target infection site [40].
Tere are many limitations in this study.Due to a single centric, retrospective designed, small cohort observational study, our results may not be representative of experience at other institutions and may not provide a solid basis for recommendations for practice in clinical settings.Due to late introduction of carbapenemase molecular testing at our institute, suitability of CAZ-AVI in patients with CRKP-NDMs (or any other metallo-β-lactamase) producers is questionable considering lack of data [45].Due to small study size, we are unable to make defnitive conclusions about the role of combination regimens in improving outcomes and about the efectiveness of ceftazidimeavibactam.A pilot study at our institute revealed a very high resistant rate to most antibiotics including colistin and tigecycline and complex genetic environment of CRKP with the carriage of multiple resistance markers blaOXA-48 (91.6%) blaOXA-51 (45.8%) and blaOXA-23 (41.6%).Sequencing of the blaNDM amplicons revealed the presence of blaNDM-1 with 100% of isolates showing the presence of qnrS [45].Indication bias cannot be ruled out, as in all retrospective studies, as an explanation for the lack of beneft with ceftazidime-avibactam combination regimens.

Conclusion
Early advent of CAZ-AVI resistance in CRE isolates highlights the dynamic necessity of routine CAZ-AVI resistance laboratory testing and antimicrobial stewardship programmes focusing on the utilization of all antibiotics.Consolidating the hospital infection control of tracheostomies may help to prevent CAZ-AVI resistance in CRKP.Colistin may aid in decreasing mortality rates among patients with CAZ-AVI CRKP isolates.
Values are expressed in median (range) unless specifed otherwise.N � number; g � gram; L � liter.A

Table 3 :
Comparison of mortality in patients receiving colistin amongst the study participants.