Direct-on-Target Microdroplet Growth Assay for Detection of Bacterial Resistance in Positive Blood Cultures

Introduction: The recently developed DOT-MGA (direct-on-target microdroplet growth assay) has shown the desirability of direct application of this approach in positive blood cultures and its good performance in detection. This study selected 44 Enterobacteriaceae strains and implemented a DOT-MGA assay on blood cultures to detect their resistance to seven antibiotics. The results of DOT-MGA were compared with the other two antimicrobial susceptibility testing (AST) methods to analyze the detection performance of DOT-MGA.

Methods: We adopted the differential centrifugation to process positive blood-culture (BC). Processed BC broth was directly used for rapid AST using DOT-MGA. Droplets of 6 µL with and without antibiotics at the EUCAST breakpoint concentration were spotted in triplicates onto the surface of a MALDI target. The plates were incubated in a wet box for 4 h before the broth was removed with filter paper. Bruker Biotyper software was used to analyze the test results compared with standard database, and the scores were used to quantify and determine the results.

Results: DOT-MGA results were compared with the direct-from-BC disk-diffusion method and results were reported by broth microdilution method, respectively. The comparison demonstrated a 100% growth efficiency in DOT-MGA, a 100% classification consistency for ampicillin, ceftriaxone, and gentamicin, and >93% classification consistency for tobramycin, aztreonam, trimethoprim-sulfamethoxazole (TMP-SMX), and ceftazidime.

Discussion: These study results have shown that DOT-MGA is suitable for directly identifying bacterial resistance to positive blood cultures in clinical microbiology laboratories. Furthermore, it is conducive for early diagnosis and treatment of patients with bloodstream infection due to its convenience, time efficiency, and good performance in identifying multiple antibiotic-insensitive bacteria.