IoT-integrated and portable real-time LAMP platform with hydroxy naphthol blue (HNB) for universal, low-cost pathogen...

The timely and cost-effective detection of infectious pathogens is critical for their prevention and control. In this study, a low-cost, portable, and user-friendly WiFi-enabled Internet of Things (IoT) monitoring system was developed for the detection of infectious pathogens. The detection mechanism relies on real-time monitoring of hydroxy naphthol blue (HNB) colorimetric signals, which are triggered by the depletion of Mg²⁺ ions during loop-mediated isothermal amplification (LAMP). A 3D-printed, portable reader was developed for both LAMP incubation and real-time HNB signal monitoring. Analysis at an incubation temperature of 65 °C demonstrated the excellent thermal stability of HNB and its linear correlation (R² = 0.9973) with Mg²⁺ ion consumption, highlighting its suitability as a real-time indicator. The detection capability was validated using the Nosema bombycis SSU rRNA gene and the Bombyx mori nucleopolyhedrovirus gp41 gene as model targets. The results showed linear relationships (R² > 0.99) between threshold time and log target concentration, with the limit of detection (LOD) reaching 1 copy µL^-1 for both targets. Analysis of Nosema bombycis in real samples showed comparable performance to the conventional female moth microscopy method. Furthermore, the IoT-enabled reader allowed for fully automated result reporting via email, facilitating timely implementation of containment measures. This study reported on the development of a portable and user-friendly IoT-based test platform, presenting attractive potential for point-of-care diagnosis in infectious diseases, particularly in resource-constrained settings.