Development of Microfluidic Chip-Based Loop-Mediated Isothermal Amplification (LAMP) Method for Detection of Carbapenemase Producing Bacteria

Abstract

ABSTRACT The rapid and accurate diagnostic methods to identify carbapenemase-producing organisms (CPO) is of great importance for controlling the CPO infection. Herein, we have developed a microfluidic chip-based technique to detect CPO and assessed its clinical value in detecting CPO directly from blood cultures (BCs). The detection performance of the microfluidic chip-based LAMP amplification method was analyzed retrospectively on a collection of 192 isolates including molecularly characterized 108 CPO and 84 non-CPO and prospectively on a collection of 133 positive BCs with or without CPO suspicion, respectively. In the retrospective study, the microfluidic chip-based LAMP amplification method exhibited 87.5% accuracy (95% CI [82.0–91.5]), 97.7% sensitivity (95% CI [91.2–99.6]), 78.8% specificity (95% CI [69.5–86.0]), 79.6% positive predictive value (PPV) (95% CI [70.6–86.5]) and 97.6% negative predictive value (NPV) (95% CI [90.9–99.6]). Among the 192 isolates, 22 (11.5%) false-positives (FP) and 2 (1.0%) false negatives (FN) were observed. In the prospective study, the 133 routine isolates of positive BCs including 18 meropenem-resistant CPO and 115 non-CPO were assessed, and 4 FP were observed in non-CPO and CPO, respectively. The current method showed a total detection performance of 94.0% accuracy (95% CI [88.4–97.1]), 100.0% sensitivity (95% CI [73.2–100.0]), 93.2% specificity (95% CI [86.7–96.8]), 63.6% PPV (95% CI [40.8–82.0]) and 100.0% NPV (95% CI [95.8–100.0]). In summary, the microfluidic chip-based LAMP amplification method is reliable for the rapid screening and detection of CPO with high accuracy, sensitivity, and specificity, and could easily be implemented in clinical microbiology laboratories. IMPORTANCE Rapid and accurate identification of CPO may reduce the genetic exchanges among bacteria and prevent further dissemination of carbapenemases to non-CPO. The current method had designed microfluidic chip-based LAMP amplification method for multiplex detection of carbapenemase genes and evaluated the detection performance of the newly method. The current method can rapidly screen and detect CPO with high accuracy, sensitivity, and specificity, and could easily be implemented in clinical microbiology laboratories, as this will reduce the carbapenem resistance issues worldwide.

Keywords

• microfluidic chips
• carbapenemase-producing organisms
• loop-mediated isothermal amplification
• blood cultures
• clinical bacteria isolates
• detection performance