In July 2014, CPE was detected from a hospitalized patient at the cancer center for the very first time. In total, 18 hospitalized patients were found to have CPE during the study period confirmed by clinical culture (n = 13) or surveillance culture (n = 5) (Table 1, Table S1). Seventeen and one isolates were identified as E. claocae and Cronobacter sakazakii, respectively, with MicroScan WalkAway. All isolates were positive for production of MBLs by the SMA disk testing. Date of first isolation of CPE from all but one (Patient-18) patients was clustered in a 16-months period (between July 2014 and Oct 2015). The mean age of 18 patients was 68.8 years. Sixteen patients (88.9%) were male. Seventeen patients (94.4%) had gastrointestinal malignancy. Six (33.3%), fourteen (77.8%), and sixteen (88.9%) patients had received chemotherapy within 90 days, surgery within 90 days, and antimicrobial therapy within 30 days of the first isolation of CPE, respectively. Although the patients were under the care of 7 different departments, Surgery-A (n = 7), Surgery-B (n = 4), and Medical Oncology-E (n = 3) departments were involved in the care of multiple patients. At the time of CPE isolation, the patients were located at one of five different wards or one intensive care unit (ICU). Six, five, and three patients were at ICU, Ward-V, and Ward-X, respectively. Fifteen patients (83.3%) received endoscopic procedure within 90 days of isolation of CPE. All six patients from whom CPE was isolated within 3 days after surgery (Patient-2, -5, -10, -14, -15, and -16) received routine airway nebulization and frequent airway suctioning at ICU and bronchoscopy was also performed at operating rooms in two patients (Patient 5 and -15). Thirteen (76%) of seventeen case patients between July 2014 and Oct 2015 had history of hospitalization at multiple wards before the isolation of CPE (Figure 1).
A ward-wide active surveillance was performed twice at Ward-V (November 2014 and February 2015), three times at Ward-X (May, June, and July 2015), and once at Ward Y (November 2014). A total of 191 patients were screened and two patients were positive for carriage of CPE (Patient-7 and -12). Additionally, a patient with a history of admission in Ward-V was screened upon a later admission at Ward-X and was positive for the growth of CPE (Patient-9). Surveillance cultures performed at the discretion of primary physicians immediately after surgery without using selective media turned positive for carriage of CPE in two patients (Patient-2 and -6).
Six patients had infections due to CPE (Table 2). Although one patient died of an intraabdominal infection following intestinal perforation on the 11th day of the onset of the infection, infections in other patients were cured without relapse. Two other patients died within 90 days due to reasons unrelated to the carriage of CPE (Table S1).
The patients with a history of isolation of CPE were cared for in a private room under contact precautions according to the infection prevention protocol of the hospital. After July 2014 when the isolation of CPE from patients hospitalized in different wards was documented, occurrence of an institutional outbreak of CPE was notified to the all hospital staffs and strict compliance to the infection prevention protocol was enforced. In addition, direct observation of hand hygiene compliance was initiated by infection preventionists and the data were fed back to each hospital department. Compliance to the infection prevention protocol was thoroughly checked especially at the wards where the patients with CPE was hospitalized. Sampling from hospital environment was not performed. Compliance to the cleaning and disinfection protocol of endoscopes was confirmed and bacterial cultures of the relevant endoscopes, including bronchoscopes for the operating rooms and ICU and duodenoscope, were negative for the growth of CPE.
Microbiological and molecular analysis of CPE
All CPE isolates were non-susceptible to cefepime and all but one isolates were non-susceptible to piperacillin-tazobactam with BD Phoenix NMIC/ID-208 panel. Three isolates were susceptible to aztreonam. Although most of the isolates were non-susceptible to carbapenems, MIC of >4 mg/mL for imipenem and meropenem was observed only in two isolates and one isolate, respectively. Most of the isolates was susceptible to non-b-lactam antibiotics tested (Table 3).
Fourteen of eighteen CPE isolates were identified as E. hormaechei by ANI and other isolates were E. hormaechei subsp. steigerwaltii (n = 2), E. asburiae (n = 1), and E. xiangfangensis (n = 1) (Table 3). All E. hormaechei isolates were ST78 and isolated between July 2014 and August 2015. While one isolate (TUM17942) carried blaIMP-11, all other isolates carried blaIMP-1. As expected, all isolates carried chromosomal ampC genes, but acquired genes for extended-spectrum b-lactamases (ESBL), AmpC, and carbapenemases other than blaIMP were not identified in any isolates. Plasmid replicon for IncHI2 was documented in 16 of 18 isolates and all 6 isolates in which full nucleotide sequences required for pMLST of IncHI2 plasmids were obtained were pMLST-ST1 (smr0119:1-smr0018:1). Remaining ten isolates also had allele profile of ‘1’ for smr00119 (smr00118 of these isolates had 0.3% difference from allele 1 (n = 4), were not fully sequenced (n = 2), or were non-typable (n = 4)).
All isolates carrying blaIMP-1 had In316 (intI1-blaIMP-1-aac(6’)-IIc-sul1)-like structure. In 7 isolates (TUM17941, TUM17943, TUM17945, TUM17946, TUM17947, TUM17948, and TUM17949), nucleotide sequences of In316 were completely preserved and other isolates had single nucleotide difference (n = 5) or had fragmentation of the structure into multiple contigs (n = 5).
ST78 isolates were divided into three clades by core-genome (4,257,370bp, 82.8% of the genomic sequence of the reference strain) based SNP analysis (Figure 2). All patients from whom isolates of clade A were identified had a history of admission at ICU, Ward-V, or Ward-Y. On the other hand, all patients from whom isolates of clade C were identified had a history of multiple admission at Ward-X (Figure 1).
In 13 isolates, blaIMP was successfully transferred into recipient E. coli cell by conjugation experiment (Table 3). All transconjugant were positive for PCR using primers for blaIMP-1-group. TUM17942 yielded transconjugants positive for IncL/M by PCR-based replicon typing. Transconjugants of the remaining isolates were positive for IncHI2 by PCR-based replicon typing, which suggested the location of blaIMP-1 on IncHI2 plasmids.