Feasibility of Heteroresistance Screening on Escherichia coli to Predict the Presence of Inner Colonies (IC) During Fosfomycin Disk Diffusion Testing
Feasibility of Heteroresistance Screening on Escherichia coli to Predict the Presence of Inner Colonies (IC) During Fosfomycin Disk Diffusion Testing Jenna M. Salay1, Morgan L. Bixby1, Elizabeth B. Hirsch1 1University of Minnesota College of Pharmacy, Minneapolis, MN Background: The presence of IC within the zone of inhibition during disk diffusion (DD) testing in E.coli have been noted, but CLSI and EUCAST interpretation criteria for IC are contradictory. Because of the presence of resistant IC, there is a need to determine the most appropriate method of interpretation to prevent a potential increase of resistant infections, thus evaluating the susceptibility of IC E.coli isolates in comparison to the corresponding parent isolates will help determine a more optimal method for DD testing. A heteroresistance (subpopulations with increased resistance) screening test has potential to identify isolates with IC. We sought to determine the feasibility of heteroresistance screening to predict the presence of IC in clinical E.coli isolates. Materials/Methods: A convenience collection (n=46) of E. coli parent isolates underwent DD testing to isolate those with IC. Broth microdilution (BMD) testing was performed on all isolates and their IC, when present (n=40), to determine minimal inhibitory concentrations (MICs) and to establish inclusion criteria for the heteroresistance screening. All parent isolates with MICs of ≤ 64 ug/mL and IC isolates with MICs of ≥ 128 ug/mL were chosen for heteroresistance screening. A difference of at least 3 dilutions between parent and IC isolate MIC established the inclusion criteria. A disk elution test for heteroresistance screening was performed in duplicate on separate days. In tubes of Mueller-Hinton broth, 6 commercial fosfomycin disks (each 200 ug fosfomycin and 50 ug glucose-6-phosphate) were eluted for 90 minutes. One hundred uL of each bacterial isolate from an overnight culture was suspended in the tubes. A positive test was a turbid tube after 72 hours of incubation. Results: The parent isolates (n=46) had an MIC range between 1 and >256 ug/mL with a median of 4 ug/mL. The subset of IC isolates (n=34/40) tested to date had an MIC range between ≤8 and >1024 with a median of 128 ug/mL. Nineteen IC isolates met the inclusion criteria for the heteroresistance screening. Of the 8 isolates with heteroresistance screening completed to date, 5 isolates produced IC (MIC range of 1-8 ug/mL) and 3 isolates did not produce IC (MIC range of 1- 32 ug/mL). Of these 8 isolates, only 2 (25%) were heteroresistant using the disk elution test. Of those with IC, 2 (40%) were heteroresistant while 0 (0%) isolates without IC were heteroresistant. Conclusion: A heteroresistance screening test did not provide consistent data to predict the presence of IC among this E.coli collection. A larger isolate set and further studies are needed to understand the feasibility of a heteroresistance screening test and increased resistance in IC in E.coli.