Abstract
Enterobacter cloacae complex (ECC) is composed of multiple species and the taxonomic status is consecutively updated. In last decades ECC is frequently associated with multidrug resistance and become an important nosocomial pathogen. Currently, rapid and accurate identification of ECC to the species level remains a technical challenge, thus impedes our understanding of the population at the species level. Here, we aimed to develop a simple, reliable, and economical method to distinguish four epidemiologically prevalent species of ECC with clinical significance, i.e., E. cloacae, E. hormaechei, E. roggenkampii, and E. kobei. A total of 977 ECC genomes were retrieved from the GenBank, and unique gene for each species was obtained by core-genome comparisons. Four pairs of species-specific primers were designed based on the unique genes. A total of 231 ECC clinical strains were typed both by hsp60 typing and by species-specific PCRs. The specificity and sensitivity of the four species-specific PCRs ranged between 96.56% and 100% and between 76.47% and 100%, respectively. The PCR for E. cloacae showed the highest specificity and sensitivity. A one-step multiplex PCR was subsequently established by combining the species-specific primers. Additional 53 hsp60-typed ECC and 20 non-ECC isolates belonging to six species obtained from samples of patients, sewage water and feces of feeding animals were tested by the multiplex PCR. The identification results of both techniques were concordant. The multiplex PCR established in this study provides an accurate, expeditious, and cost-effective way for routine diagnosis and molecular surveillance of ECC strains at species level.
Highlights
Enterobacter spp. are natural commensals of the animal and human gut microbiota (Sanders and Sanders, 1997; Mesa et al, 2006)
The primary goal of our study is to setup and optimize a one-step multiplex PCR assay capable of differentiating the major E. cloacae complex (ECC) species (i.e. E. cloacae, E. hormaechei, E. roggenkampii, and E. kobei) with clinical significance
According to the ANI results, 808 genomes were assigned to E. hormaechei (n=714), E. cloacae (n=50), E. kobei (n=26), and E. roggenkampii (n=18)
Summary
Enterobacter spp. are natural commensals of the animal and human gut microbiota (Sanders and Sanders, 1997; Mesa et al, 2006). Among Enterobacter spp., the E. cloacae complex (ECC) is of major importance, accounting for 65%–75% of infections in clinical settings (Sanders and Sanders, 1997). The ECC is frequently associated with a multidrug resistance (MDR) phenotype, mainly due to the ability to acquire numerous genetic mobile elements carrying resistance genes. The chromosome of ECC intrinsically encodes ampC genes, resulting in a constitutive production of the AmpC b-lactamase and is intrinsically resistant to ampicillin, amoxicillin, amoxicillin/ clavulanate, first-generation cephalosporins, and cefoxitin (Mezzatesta et al, 2012). Available epidemiological data show that the ECC has become the third major drug-resistant Enterobacteriaceae species involved in nosocomial infections after Escherichia coli and K. pneumoniae (Zhou et al, 2018). It is important to control the dissemination of MDR-ECC for public health
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