Colistin Susceptibility Testing of Enterobacteriaceae by Agar Dilution (AD), Broth Microdilution (BMD) and Polymyxin NP

BACKGROUND: Polymyxin resistance among Enterobacteriaceae is increasingly reported worldwide, with plasmid-mediated colistin resistance, conferred by mcr-1, recently reported. In 2017, CLSI set colistin Epidemiological Cutoff Values (ECVs) for Enterobacteriaceae. There are limited accurate methods for colistin susceptibility testing. The new rapid polymyxin NP (PBNP) test detects bacterial growth in the presence of colistin. We evaluated AD and BMD in comparison to PBNP using clinical isolates of Enterobacteriaceae, which we also tested for mcr-1. We additionally gathered colistin MIC data among Enterobacteriaceae isolates over a period of 6 years. METHODS: Colistin MICs were determined by BMD and AD for 100 clinical isolates of Enterobacteriaceae submitted to our laboratory from August 2016 to February 2017. mcr-1 testing was performed via a laboratory developed real-time PCR assay on a LightCycler 480 platform. PBNP was also performed. Colistin MIC distributions, determined using AD, were reviewed for all isolates of Enterobacteriaceae submitted to our laboratory from 2011 to 2017 after excluding species with intrinsic resistance to colistin. RESULTS: With BMD as the reference method, the essential and categorical agreement of AD was 86.3 and 97.7%, respectively. The very major and major error rates for AD were 2.5% (1/40) and 2.9% (1/34), respectively. Sensitivity and specificity of PBNP were 90.7 and 94.1%, respectively. One isolate tested positive for mcr-1 (Escherichia coli, MIC 4 µg/mL by AD and BMD and positive PBNP). Excluding species with intrinsic resistance to colistin, 1153/48,441 isolates (2.4%) had colistin MICs ≥ 4 µg/mL by AD. Enterobacter cloacae complex, Klebsiella pneumoniae and E. coli were the most common species with colistin MICs ≥ 4 µg/mL (by AD). 2.7% (31/1153) of isolates with colistin MICs ≥ 4 µg/mL (by AD) were also resistant to a carbapenem; K. pneumoniae was the most common species with concomitant colistin MICs ≥ 4 µg/mL by AD and carbapenem resistance. CONCLUSION: A low percentage of isolates surveyed over the past 6 years demonstrated elevated MICs to colistin by AD. AD did not meet essential agreement criteria for colistin susceptibility testing. PBNP was found to have good sensitivity and specificity when compared with BMD. DISCLOSURES: R. Patel, ASM: Board Member, None CD Diagnostics, BioFire, Curetis, Merck, Hutchison Biofilm Medical Solutions, Accelerate Diagnostics, Allergan, and The Medicines Company: Grant Investigator, Grant recipient Curetis: Consultant, Monies paid to my employer A patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication with royalties paid by Samsung to Mayo Clinic, and a patent on an anti-biofilm substance issued: Patents, Patents, any money is paid to my employer Actelion: DSMB, Money paid to my employer ASM and IDSA: Editor’s stipends, Editor’s stipends NBME, Up-to-Date and the Infectious Diseases Board Review Course: NBME, Up-to-Date and the Infectious Diseases Board Review Course, Honoraria