Comparison of Six Phenotypic Assays with Reference Methods for Assessing Colistin Resistance in Clinical Isolates of Carbapenemase-Producing Enterobacterales: Challenges and Opportunities

Abstract

The global escalation of severe infections due to carbapenemase-producing Enterobacterales (CPE) isolates has prompted increased usage of parenteral colistin. Considering the reported difficulties in assessing their susceptibility to colistin, the purpose of the study was to perform a comparative evaluation of six phenotypic assays—the colistin broth disc elution (CBDE), Vitek 2 Compact (bioMérieux SA, Marcy l’Etoile, France), the Micronaut MIC-Strip Colistin (Merlin Diagnostika GMBH, Bornheim-Hensel, Germany), the gradient diffusion strip Etest (bioMérieux SA, Marcy l’Etoile, France), ChromID Colistin R Agar (COLR) (bioMérieux SA, Marcy l’Etoile, France), and the Rapid Polymyxin NP Test (ELITechGroup, Signes, France)—versus the reference method of broth microdilution (BMD). All false resistance results were further assessed using population analysis profiling (PAP). Ninety-two nonrepetitive clinical CPE strains collected from two hospitals were evaluated. The BMD confirmed 36 (39.13%) isolates susceptible to colistin. According to the BMD, the Micronaut MIC-Strip Colistin, the CBDE, and the COLR medium exhibited category agreement (CA) of 100%. In comparison with the BMD, the highest very major discrepancy (VMD) was noted for Etest (n = 15), and the only false resistance results were recorded for the Rapid Polymyxin NP Test (n = 3). Only the PAP method and the Rapid Polymyxin NP Test were able to detect heteroresistant isolates (n = 2). Thus, there is an urgent need to further optimize the diagnosis strategies for colistin resistance.

Characterization of Carbapenemase-Producing Klebsiella pneumoniae Isolates from Two Romanian Hospitals Co-Presenting Resistance and Heteroresistance to Colistin

“…Consequently, in the first case, a combination of mutations in mgrB (M27K), phoP (L4F), and phoQ (Q426L) genes was noted, while in the second case, an L26Q substitution in the phoQ gene was associated with R256G mutation in the pmrB gene. However, several reports have mentioned that Vitek 2 Compact is inappropriate for colistin susceptibility testing [ 63 , 64 , 65 ], especially for strains showing an MIC > 1 mg/L by BMD [ 63 ]. The manufacturer of Vitek 2 Compact has also recommended using an alternative method prior to reporting colistin results obtained with several types of testing cards in this automated system [ 66 ], at least until this issue can be addressed through technology upgrades to be implemented in the future.…”

“…Remarkably, another significant aspect of our study is the molecular basis of the two pairs of ChR (parental and the corresponding resistant mutants) isolates, which are reported for the first time in Romania [ 65 ]. These two pairs of strains co-harbored an mgrB alteration caused by an ISKpn25 element insertion at the same position (with the deletion of nucleotides 1–5) and an R256G substitution in pmrB .…”

“…The retrospective nature of the study did not allow us to provide a reliable picture of all significant clinical and therapeutic aspects or previous hospitalizations, which was beyond the scope of this manuscript. However, the data on previous colistin therapy in patients diagnosed with ChR strains may be found elsewhere [ 65 ]. BMD was performed only for colistin as the research was focused on detecting the molecular basis of resistance and heteroresistance to this antimicrobial agent.…”

“…Additionally, 2 ChR K. pneumoniae strains obtained from Târgu Mureș in March 2019 and their corresponding colistin-resistant mutants were included in the study. The collection comprising the 10 CR and 2 ChR isolates has been phenotypically characterized in an earlier study [ 65 ].…”

“…All K. pneumoniae CPE strains were additionally assessed for colistin resistance by BMD and using the following 5 phenotypic methods, as described elsewhere [ 65 ]: Micronaut MIC-Strip (Merlin Diagnostika GmbH, Bornheim-Hersel, Germany), Etest gradient diffusion strip on Mueller Hinton E agar (MHE) (BioMérieux SA, Marcy l’Etoile, France), ChromID Colistin R agar (COLR) assay (BioMérieux SA, Marcy l’Etoile, France), Rapid Polymyxin NP test (ELITechGroup, Signes, France), and colistin broth disc elution. Moreover, the 2 ChR strains were further confirmed using the population analysis profiling (PAP) assay, as also described in [ 65 ].…”