# Verification and application of an automated real-time antimicrobial susceptibility testing system: for accelerated antibiotic susceptibility testing and high-throughput screening of antibiotic synergistic effects

## TL;DR

A new automated system for testing antibiotic susceptibility is developed, significantly speeding up resistance detection and identifying effective drug combinations.

## Contribution

The AR-AST system introduces a novel approach combining early growth kinetics with threshold and slope verification for rapid and accurate antimicrobial testing.

## Key findings

- The AR-AST system reduced resistance identification time by 75.54% for Acinetobacter baumannii and Escherichia coli in a hospital setting.
- The system identified a synergistic effect between kanamycin and polymyxin B against resistant E. coli with 100% inhibition and 68.03% time savings.
- The system demonstrated high accuracy and stability with a detection range of six orders of magnitude for E. coli.

## Abstract

Bacterial infections remain a serious global health threat, exacerbated by the widespread use of antibiotics leading to bacterial resistance. Faced with the continuous evolution and accelerated spread of drug-resistant bacteria, there is an urgent need to establish an efficient and automated system to accelerate bacterial resistance identification and combination therapy screening in clinical practice. Here, we present an Automated Real-time Antimicrobial Susceptibility Testing (AR-AST) system, which represents an innovation in the AST field by enabling continuous, high-resolution monitoring of microbial growth. This system demonstrated exceptional adaptability, stability, and robustness in monitoring the growth kinetics of six common pathogenic bacteria. Through the establishment of a combined interpretation system of “early bacterial growth kinetics + PPT threshold + slope verification”, it effectively reduces the risk of misclassification. This approach achieved a reduction in the time required for resistance identification of clinical Acinetobacter baumannii 186 and Escherichia coli 15,017 by approximately 75.54% (95% CI: 64.54% − 86.55%) in a hospital setting. Furthermore, when applied to screen pairwise combinations of polymyxin B, polymyxin E, and kanamycin against resistant E. coli15017, a significant synergistic effect was observed for the kanamycin and polymyxin B combination (FIC index = 0.42 < 0.5). The complete inhibition rate in the binary combination was 100% (95% CI: 47.86% − 100%), with a time saving of approximately 68.03% (95% CI: 53.20% − 82.86%). Additionally, the AR-AST System successfully facilitated the quantitative analysis of E. coli15017, exhibiting a broad detection range of six orders of magnitude (10¹ to 10⁶ CFU/mL), high accuracy (|Δ Log₁₀| < ± 0.5), and good stability (P > 0.05), thereby providing effective auxiliary validation for evaluating combination therapy efficacy.

The online version contains supplementary material available at 10.1186/s13036-026-00630-3.