Bacterial isolation and identification
Over the 5-month period, 250 bronchoscopes were sampled and 500 samples were collected, including 250 samples after clinical procedures and 250 samples after usual decontamination procedures. All bronchoscope samples were tested for bacterial screens. A total of 358 isolates and 13 isolates were recovered from samples after clinical procedures and samples after decontamination procedures, respectively (Table S2 and Table S3). This might explain by the low-level contamination with environmental and skin bacteria, since bronchoscope samples were collected after the clinical procedures without any disinfection or cleaning processes. Of note, most of the detected microorganisms were Gram-positive bacteria, such as Staphylococcus epidermidis (n = 69), Streptococcus salivarius (n = 42), and Streptococcus oralis (n = 23). Among the Gram-negative bacilli isolates, most of them were that belong to the Enterobacteriaceae group (Table S2 and Table S3). Outbreaks and pseudo-outbreaks associated with bronchoscopes have been well documented in the literature [32]. These nosocomial infections commonly associated with Mycobacterium spp. and Enterobacteriaceae isolates [33-37]. Moreover, K. aerogenes was found in both groups (before and after the cleaning procedures). All this prompted us to further investigated the phenotypic and genomic characteristics of 7 K. aerogenes identified in this work.
AST and MLST of K. aerogenes isolates
AST results of 7 K. aerogenes isolates were detailed in Table 1. The full resistance rate was observed for amoxicillin-clavulanic acid and cefoxitin (100%). All the isolates were susceptible to piperacillin-tazobactam, cefuroxime, ceftazidime, ceftriaxone, cefoperazone-sulbactam, cefepime, ertapenem, amikacin, levofloxacin, tigecycline, trimethoprim-sulfamethoxazole. Interestingly, 5 isolates were intermediate to imipenem. Among 7 K. aerogenes isolates, we found 5 sequence types (STs), which were ST135 (n = 2) and ST1358 (n = 2), followed by ST1357 (n = 1), ST1359 (n = 1), and ST1363 (n = 1). K. aerogenes is reported to associated with nosocomial infections and displaying multidrug resistance [16], and the most prevalent STs were ST93 and ST4 [38]. However, we didn’t detect any multidrug resistant K. aerogenes in this work. Furthermore, STs of K. aerogenes found in this work have not been described in the literature. These data suggested that these colonized isolates may have an environmental origin.
Whole genome sequencing and comparison analysis
Roary matrix-based gene sequence analysis generated a large pan-genome of 18,105 gene clusters of 58 full genomes. The whole-genome phylogeny (Figure 1) revealed a population structure that was generally concordant with MLST (data not shown). Genetic diversity was observed in our bacterial collection, which clustered into three main clades. The 2 ST1358 strains identified in this study were aggregated in 1 clade with 1 ST1364 human isolate from Spain, which suggested that ST1358 and ST1364 might originate from the same ancestor. The results also indicate that 11 ST93 isolates and 9 ST56 were clustered into one separate sub-cluster, respectively, which exhibited a slight difference in the core genome sequence. Recent studies found that ST93 was the most prevalent clone in the global K. aerogenes genome database, which indicated that ST93 might be dominant sequence type global clones in clinical settings [13, 39]. Furthermore, the emergence of fecal carriage and human infection MDR K. aerogenes isolates resistant to multiple antibiotics, especially resistant to carbapenems is considered a substantial threat to public health [17, 40]. Although phenotypic and genomic evidence from the current study revealed that isolates recovered in this work are not multi-resistant, active surveillance of bronchoscope-associated K. aerogenes isolates would improve our understanding on the population structure of this species. Of note, all isolates recovered from this study have a close relation to environmental or human isolates. A recent study investigated the population structure, virulence, and antimicrobial resistance in K. aerogenes [38]. Their findings showed that K. aerogenes has an open pangenome and a large effective population size, which is in line with our results.
Resistant determinants of K. aerogenes isolates
We identified a total of 43 antimicrobial resistance genes in the K. aerogenes core genomes (Figure 2). The resistome of K. aerogenes comprise a high number of antibiotic efflux pumps as well as narrow and extended spectrum β-lactamases. As expected, human isolates encoded more antimicrobial resistance genes than those environmental isolates. Of note, 3 isolate from this work possessed only one resistance gene, fosA, which is consistent whit phenotypic characteristics.