The acquisition of Multi-drug resistant bacteria in patients admitted to COVID-19 intensive care units: a monocentric retrospective case control study

Background: Whether the risk of multidrug resistant bacteria (MDRB) acquisition in the intensive care unit (ICU) is increased during the COVID-19 crisis is unknown. Our aim was to measure the rate of MDRB acquisition in patients admitted in COVID-19 ICU and to compare it with pre-COVID-19 controls. Methods: This single center case control study included adult patients admitted to COVID-19 ICUs for more than 48h. Patients were screen twice a week for MDRB carriage during ICU stay. We compared the rate of MDRB acquisition of COVID-19 patients with a cohort of patients admitted for subarachnoid hemorrhage (SAH) and matched on length of ICU stay using a competing risk analysis. Results: Among 72 patients admitted to the COVID-19 ICUs, 24 (33%) patients acquired 31 MDRB during ICU stay. The rate of MDRB acquisition was 30/1000 patient-days. Patients that acquired MDRB had received more antimicrobial therapy [22 (92%) vs 34 (78%, p=0.05] and had a longer exposure time [12 days (8-18) vs 5 days (2-18), p=0.02]. The rate of MDRB acquisition in matched SAH patients was 18/1000 patient-days. When compared to SAH retrospective cohort, being admitted to a COVID-19 ICU was associated with a numerically higher proportion of MDRB acquisition. This association did not reach statistical signicance in the multivariable competing risk analysis (sHR 1.71 (CI 95% 0.93-3.21). Conclusion: Acquisition of MDRB was frequent during the COVID-19 rst wave in ICU patients. Despite physical isolation, it was similar to patients admitted to the same ICU in previous years.


Introduction
The respiratory infection COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread worldwide, being considered pandemic by the World Health Organization since March 11, 2020. [1] During this pandemic, health care systems worldwide became overloaded and experienced shortages of intensive care unit (ICU) beds [2] and protective personal equipment (PPE). [3] In the most severely affected countries, ICUs were lled with only COVID-19 patients, and named "COVID-19 ICUs"; new ICUs and hospital beds had to be opened, [4] health care professionals were reassigned and started working longer hours. These factors can contribute to a decreased adherence to infection prevention and control measures and, combined with a high antimicrobial selection pressure, [5] may facilitate the emergence of antimicrobial resistance (AMR). [6] AMR is a major health problem, [7] responsible for high morbi-mortality [8,9] and an elevated economic burden. [10] The ICU is one of the hospital locations where patients have the highest risk of acquiring multi drug resistant bacteria (MDRB). [11] However, until now, the risk of MDRB acquisition in the ICU during COVID-19 crisis remains unknown; whether this risk of MDRB acquisition is higher in COVID-19 units compared to normal functioning ICU has not been studied.
The aim of this study is to describe the rate of MDRB acquisition in COVID-19 ICUs compared with the MDRB acquisition in the same department before the COVID-19 crisis.

Study design and setting
We conducted a single center retrospective case-control study of COVID-19 patients admitted to the ICUs of Erasme Hospital, Brussels, Belgium from March to April 2020. The ethics committee approved this study (P2020/371) and waived the need for an informed consent.
Under normal circumstances, our ICU operates with 30 beds divided into ve ICUs with 6-single rooms each. We perform nasal and rectal surveillance cultures upon ICU admission and twice a week thereafter. Colonization by MRSA, ESBL, CPE and VRE is indicated at the room door and requires health care workers to wear gloves and gown before entry. We have a meeting with the antibiotic stewardship team twice a week.

COVID-19 crisis management
During COVID-19 crisis, we increased our ICU beds from 30 to 39, by opening 5 beds in the existing ICUs and building another 4-beds ICU in two operating rooms. Only one 6-beds ICU remained available for non-COVID-19 admissions. The remaining 33 beds were reserved for COVID-19 patients with a high risk of invasive mechanical ventilation. Patients with stable clinical condition on CPAP remained in the wards.

Patients' selection
Cases: All patients admitted to the COVID-19 ICUs of Erasme Hospital between 15/03 and 30/04, regardless of the nal etiological diagnosis, were eligible for inclusion; the sole exclusion criterion was an ICU stay < 48h. These patients were considered as cases and are named COVID-19 patients in this manuscript.

Controls:
The admission to the non-COVID 19 ICU was scarce during the study period. Therefore, it was not possible to use this population as control. Given the unusual duration of mechanical ventilation in COVID-19 patients, [12] we compared them to patients obtained from a preexisting subarachnoid hemorrhage (SAH) institutional database cohort. [13] These patients were admitted to our ICU from January 2016 -2019. Severe SAH patients are young with few comorbidities and can have prolonged ICU length of stay (LOS) [14] and duration of mechanical ventilation. [15] Patients were matched 1:1 according to ICU LOS and the use of mechanical ventilation. Patients with ICU LOS less than ten days were matched with controls with a difference of ± 2 days, and patients with ICU LOS > 10 days were matched with controls with a ± 20% difference in the LOS. When several controls could match one COVID-19 patient we selected the control with the closest ICU LOS.

Data collection and endpoints:
Demographic data and severity scores were collected at ICU admission. We de ned the length of exposure as the duration between ICU admission and the day of MDRB acquisition, or between ICU admission and ICU discharge in patients without MDRB acquisition. We collected the following data during the exposure time: antimicrobial use, presence of central venous catheter, urinary tract catheter, mechanical ventilation and the occurrence of surgery. The primary endpoint was the rate acquisition of MDRB in the COVID 19 units.

Microbiology data:
We considered patients MDRB + when a MDRB was found in any microbiological specimen. Patients that were MDRB+ within 48h after admission were considered index cases. Patients that acquired MDRB during ICU stay were considered new cases. Patients that didn't acquire MDRB during ICU stay were considered MDRB -. We collected the presence of possible cross-transmission. Cross-transmission was suspected if a patient acquired a MDR pathogen with the same antimicrobial susceptibility and resistance mechanism than another patient hospitalized at the same time in the same unit.
In our center, we perform routine surveillance cultures (rectal swab, tracheal aspirate and urinary cultures) on admission and then twice a week throughout the ICU stay.
For MDRB detection, rectal swabs are streaked onto selective plates as follow: chromID® CARBA SMART agar (bioMérieux, France) for the detection of carbapenemase-producing Enterobacteriaceae; MacConkey agar containing ceftazidime (bioTRADING, Netherlands) for the detection of third generation cephalosporin-resistant Pseudomonas aeruginosa, Klebsiella spp. and Enterobacter spp.; chromID® VRE agar for the detection of Vancomycin-resistant Enterococcus faecium. Identi cation of MDRB is performed using matrix-assisted-laser desorption ionization-time of ight analysis (MALDI-TOF).
Antimicrobial resistance was de ned according to breaking points recommended by the European Committee on Antimicrobial Susceptibility Testing (EUCAST) [16] using VITEK2 and disk diffusion method. Carbapenemases OXA-48, KPC, NDM, VIM and IMP were detected via Polymerase Chain Reaction (PCR) analysis or Coris Resist-5 O.O.K.N.V. antigenic detection (Coris BioConcept, Belgium). VanA and VanB genes were detected via PCR analysis. ESBL-producing Enterobacteriaceae and ampC derepression were identi ed using detection of synergy on disk diffusion test as recommended by EUCAST. For methicillin-resistant S.aureus detection, nasopharyngeal swabs were streaked onto ChromID® MRSA selective plates. MDR Pseudomonas and Acinetobacter spp. were de ned as recommended considering antimicrobial resistance phenotype. [17] Statistical analysis: Categorical variables are reported as count (%), continuous data that were normally distributed as mean ± standard deviation (SD) and skewed data as median [interquartile range].
Incidence of MDRB acquisition was calculated by dividing the number of new cases by the total number of patients admitted to the COVID-19 ICUs. The incidence rate was calculated by dividing the number of new cases by the total number of patient-days in the COVID-19 ICUs. We compared MDRB+ and MDRBpatients in the COVID 19 cohort using Student's T-test, Mann-Whitney test, χ² test or Fisher's exact test, as appropriate. In order to identify factors associated with the acquisition of MDRB, we compared patients admitted to COVID 19 ICUs with SAH patients admitted to our ICUs in previous years (2016-2019).
Cumulative incidence function of the acquisition of MDRB was used to describe the probability of MDRB acquisition at a given time. The Gray's test was used to test for the differences. Univariate and multivariable regression analyses were performed using the Fine and Gray competing risks proportional hazards regression model. Death was considered as a competing risk factor for the development of MDRB. In the multivariable model only variables that had p-value less than 0.2 by univariate analysis were considered.
All tests were two-tailed and a p value<0.05 was considered as statistically signi cant. Data were analyzed using IBM® SPSS® Statistics software, version 26 for Macintosh (IBM, Armonk, NY).

Characteristics of patients admitted in COVID-ICUs
We identi ed 75 patients admitted to COVID-19 units during the studied period. Three patients were excluded (ICU stay < 48h) letting 72 included patients representing 1104 patient-days of exposure. COVID-19 was diagnosed in 69 out of the 72 patients; the remaining 3 non-documented COVID-19 patients were kept in the analysis. Of note, ve patients (7%) were MDRB carriers at ICU-admission.

Characteristics of MDRB acquisition
Among the 72 patients, 24 (33%) acquired 31 MDRB (7 patients acquired more than one MDRB) during their ICU stay. The incidence rate of MDRB acquisition was 30/1000 patient-days. Figure 1 shows the isolated MDRB that were mostly Enterobacteriaceae. No MRSA were isolated. We identi ed 16/31 (52%) suspected cross-transmission events involving mainly ESBL K. pneumoniae, AmpC derepressed E. aerogenes and Vancomycin resistant E. faecium. MDRB positive and negative patients had similar demographic characteristics and severity at admission. Exposure time was longer in MDRB+ patients and antimicrobial therapy was more frequently used. In terms of clinical outcome, ICU and hospital mortality were similar between the two groups. These results are shown in table 1.

Risk of MDRB acquisition in COVID-19 patients compared to control cohort
To evaluate if the MDRB acquisition risk had changed during COVID-19 pandemic, we compared the COVID-19 patient cohort with a retrospective cohort of SAH patients hospitalized in the same ICU matched on the need for mechanical ventilation and the ICU LOS. Characteristics of both cohorts are shown in Table 2; SAH patients had a shorter duration of mechanical ventilation and received less antimicrobial therapy during exposure time.

Discussion
In this retrospective cohort of patients admitted to COVID-19 ICUs for suspected or con rmed SARS-CoV-2 infection, the rate of MDRB acquisition was 30/1000 patients-days. After adjustment and competitive risk analysis, the risk of MDRB acquisition tended to be higher in COVID-19 ICUs compared to control usual ICU.
The rate of acquisition of MDRB is highly dependent on local parameters, and thus vary greatly across centers and countries. [18] In another Belgium center 29% of patients acquired MDRB a proportion slightly higher but comparable to our control cohort. [19] Our study was not designed to explore whether the risk of MDRB acquisition is related to the COVID-19 per se or to the disruption in hospital functioning during the pandemic crisis. In the hospital setting, dissemination of MDRB happens through either cross-transmission or environmental sources, being favored at the individual level by antimicrobial therapy selection pressure [20]. In the ICU environment, cross transmission ranges from 23%-53% of patients' contacts [21,22] due to the frequent and complex cares, which facilitates the contamination of health care workers' hands and, consequently, the dissemination of MDRB [20]. Applying infection prevention and control measures and monitoring their observance are key factors to prevent MDRB spreading. [23] The possibility of dissemination of MDRB during a viral pandemic had been theoretically mentioned [24], but as many previous pandemic occurred before the antimicrobial resistance era, no data was available before the SARS-CoV-2 pandemic, including the H1N1 pandemic. Experts have raised concern about the dissemination of MDRB during the COVID-19 pandemic [6,25,26] and a report indicates an increase in blood stream infection [27]. In line with these concerns, we observed a high rate of MDRB acquisition in COVID-19 patients at least as high as in non-COVID-19 patients. The rate of MDRB acquisition in the COVID-19 ICUs was numerically higher, although not statistically different, than the rate in a control cohort of patients hospitalized in the previous years in the same ICUs. The lack of statistical signi cance may be due to the limited power of our study. Moreover, one can consider that 2 factors may have limited the MDRB acquisition rate: 1) COVID-19 patients were admitted to ICUs that had been entirely emptied and cleaned in contrast with normal ICU admission, which occurs in units where MDRB carriers are already present and 2) the physical isolation of COVID-19 patients should have provided an e cient barrier to MDRB cross-transmission.
The high rate of MDRB acquisition may be related to several di culties we experienced during this pandemic. We faced a gown shortage and needed to use the same gown for several patients. Considering the di culty of undressing PPE, it is possible that gloves were not systematically taken off at that moment. We could not document this phenomenon in our study. Beside shortage of PPE [28] other factors suspected of causing a MDRB epidemic during COVID-19 are work overload of the ICU staff, ICU overcrowding, reinforcement of less experimented staff leading to a decreased in adherence to infection prevention and control measures. [6,25,26,29] Our study was not designed to decipher the respective role of these different mechanisms but we can venture some hypothesis to explain our ndings: due to the surge of patients, we had to adapt our usual single room policy creating a new 4-beds ICU with 2 occupants per room. However, staying in our 4-beds ICU was not associated with MDRB acquisition. On the other hand, the number of concerned patients admitted to this ICU was low and no de nitive conclusion can be drawn on this point. Regarding prevention and control measures, we were not able to continue our hand hygiene and catheter-dressing audit. An increase in the use of antimicrobials is also a well-known factor associated with MDRB acquisition. [30] Patients with COVID-19, as for other viral infections, [31] were initially suspected to have a high risk of bacterial co-infection and secondary nosocomial infections. [32] In addition, initial COVID-19 symptoms may promote the initiation of antibiotic therapy even when there is no bacterial infection. [25] In line with early descriptions, [5] we observed a large antibiotic use in our study, which was signi cantly associated with a higher risk of MDRB acquisition in COVID-19 patients even if confounding variables might exist.
Our study has several limitations. First, generalization may be di cult since these results may depend on our center's characteristics. Second, we selected a control group without acute respiratory failure. However, ARDS patients are usually older and have more comorbidities than COVID-19 patients. [33] By contrast, SAH patients offered the advantage of being a cohort of patients with little comorbidity similarly to COVID 19 patients in our center as shown by the Charlson index. Moreover, taking into account predictable differences between the two cohorts, we matched the SAH patients to the COVID-19 patients according to the presence of mechanical ventilation and ICU LOS.

Conclusion
To conclude, the rate of MDRB acquisition was high in patients admitted in COVID-19. After adjustment, the risk of MDRB acquisition when being admitted to COVID 19 units was higher, but did not reach statistical signi cance, compared to control patients hospitalized for SAH before the pandemic and matched on ICU LOS and the use of mechanical ventilation. Larger multicentric studies will be necessary to assess how the viral pandemic impacts the MDRB one.

Declarations
Ethics approval and consent to participate: This study was approved by Erasme Hospital Ethics Committee under the protocol number P2020/371. Due to the retrospective nature of the study the need for informed written consent was waived.

Consent for publication: Not applicable
Availability of data and materials: The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request. All data generated or analyzed during this study are included in this published article [and its supplementary information les].
Competing interests: The authors declare that they have no competing interests Funding: This research did not receive any speci c grant from funding agencies in the public, commercial, or not-for-pro t sectors.
Authors' contributions:. DG, FT and EB were responsible for the study concept and design; EB, AI, AG, SH, DG, FT, JG, AB, NY, MH, BB, JC: acquisition of the data; DG, EB, HN, JLV: analysis and interpretation of the data; DG and EB: drafting of the manuscript; all authors: critical revision of the manuscript for important intellectual content. All authors read and approved the nal manuscript.

Supplementary Files
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