Efficient Absorbance-Based Assay for Rapid Antibiotic Susceptibility Testing of Enterobacterales

It is increasingly important to rapidly receive information on the antimicrobial susceptibility of bacteria due to the rise in antimicrobial resistance worldwide. However, traditional phenotypic methods are time-consuming. Thus, the objective of this study was to develop a rapid assay that can detect antibiotic-resistant bacterial isolates phenotypically in less than 2 h. The microplate assay used in this study is based on absorbance measurements of pure bacterial isolates grown in liquid media with and without antibiotics. Absorbance was measured at the beginning of the assay and after 90 min of incubation at 37 °C. Susceptibility was calculated for bacterial isolates that, in the absence of antibiotics, exhibited more than a 50% growth increase by comparing the absorbance value of the culture in the presence of an antibiotic at 90 min with its initial value. Here, we show that it is possible to phenotypically screen the antibiotic susceptibility of Enterobacterales and spp. isolates to three different antibiotics in 90 min. The test demonstrated an accuracy of 98.8%, sensitivity of 97.6%, and specificity of 99.6%. The false susceptibility rates were 0.2% and false resistance rates were 1.0%. This rapid and simple absorbance test has proven suitable for the screening of antibiotic susceptibility for a large number of strains with high accuracy and sensitivity. This method can be performed without specialized and costly materials and/or equipment, which makes it highly suitable for laboratories with limited resources.