Because of the advancements in medicine and science, the numbers of patients surviving complicated diseases are continuously increasing, which in turn leads to elevated chances of anaerobic infections by endogenous bacteria. Traditional growth yield-based antibiotic susceptibility tests (ASTs) against anaerobic bacteria are very time-consuming (≥48 h) and labor intensive, which delays the timely guidance of antibiotic prescription and increases the mortality of patients. Inspired by a fluorescent d-amino acid (FDAA) labeling-based AST (FaAST) that we recently developed for quick determination of aerobic bacteria's susceptibilities, here we report an accurate and fast AST method for anaerobic pathogens. Based on flow cytometry analysis of anaerobes that have been treated with various doses of antibiotics and metabolically labeled with FDAA, the intensities of which can reflect their affected metabolic status by the drugs, the MICs of each drug can then be determined. The whole process can be completed in 5 h. After testing 40 combinations of the representative anaerobic bacteria and antibiotics, our method demonstrates a high susceptibility category accuracy of 95.0%. This FaAST-based protocol is helpful in accurately and quickly guiding antibiotic decisions when treating critical infections caused by anaerobic bacteria.
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