Mycobacteriophage Functionalized Magnetic Nanocrystal Clusters for Highly Sensitive and Rapid Detection of <i>Mycobacterium tuberculosis</i>

Tuberculosis caused by Mycobacterium tuberculosis (Mtb) is one of the most dangerous diseases globally. Mtb poses a heavy death toll, especially in low-resource settings, where inadequate diagnostic capabilities greatly hinder treatment and prevention. Here, we present a rapid and cost-effective bacilli-capturing method that uses magnetic nanoclusters conjugated with mycobacteriophages. The mycobacteriophages provide Mtb recognition functionality, and the binding of the nanoparticles with attenuated Mtb H37Rv and Mycobacterium bovis Bacillus Calmette-Guérin (BCG) was visualized by electron microscopy. The magnetic nanocrystal clusters have an excellent separation efficiency. A nearly 100% capturing efficiency and high specificity toward mycobacteria species were obtained. Magnetically separated mycobacteria were disrupted by ultrasound to facilitate the rapid release of cellular adenosine triphosphate (ATP) for bioluminescent detection. Using portable and inexpensive devices, we achieved rapid detection of Mtb at as low as 1000 bacilli per sample in artificial sputum, urine, and whole porcine blood within 35 min. This method demonstrates excellent potential for point-of-care tuberculosis diagnosis in resource-limited settings.