Differential DNA accessibility to polymerase enables 30-minute phenotypic β-lactam antibiotic susceptibility testing of carbapenem-resistant Enterobacteriaceae
Fig 5
Validation of the pol-aAST method using lysed control and antibiotic-treated aliquots.
(a) Example calculation of TTPD between the lysed control and antibiotic-treated aliquots (TTPDLT). The TTP (in minutes) in the lysed control and antibiotic-treated aliquots are used to calculate TTPDLT. (b–d) The pol-aAST results using E. coli (b), K. pneumoniae (c), and Enterobacter spp. (d) isolates exposed to CRO, ETP, and MEM for 15 min. Red points represent isolates with either no detectable carbapenemase genes (Ec and Kp isolates) according to a published genotypic assay [67] and commercial assay [68], or no predictive genotype (Ebs isolates) according to the CDC [66]. S/R thresholds (dashed lines) were set halfway between the lowest susceptible and the highest resistant TTPDLT values except in the case of Enterobacter spp. treated with CRO (see text). Raw data are provided in S3 Table. +ABX, antibiotic-treated; AST, antibiotic susceptibility testing; CDC, Centers for Disease Control and Prevention; ctrl, control; CRO, ceftriaxone; Ebs, E. aerogenes and E. cloacae collectively; Ec, E. coli; ETP, ertapenem; Kp, K. pneumoniae; lc, lysed control; MEM, meropenem; pol-aAST, polymerase-accessibility AST; R, resistant; RFU, relative fluorescent units; S, susceptible; TTP, time-to-positive; TTPD, time-to-positive difference; TTPDLT, TTPD lysed-control to treated.