Palm Multidiagnostic of Mycoplasma pneumoniae, Chlamydia pneumoniae, Haemophilus influenzae, and Streptococcus pneumoniae Using One-Tube CRISPR/Cas12a

Abstract

The recent high incidence of Mycoplasma pneumoniae (Mp) infections has raised widespread public health concerns. Therefore, rapid and accurate diagnosis of respiratory pathogenic microbial infections is of paramount importance to provide clinicians with accurate diagnostic insights and guide clinical medication. In response to this urgent need, we developed a one-tube Palm CRISPR/Cas12a Diagnostic (PaCD) method. This method facilitates the rapid detection of Mp infections, as well as three other prevalent respiratory pathogens, Chlamydia pneumoniae (Cp), Haemophilus influenzae (Hi), and Streptococcus pneumoniae (SP). In addition, 3D printing was employed to fabricate a compact detection device that includes a temperature control module set at 39°C and a blue light irradiation module, significantly enhancing the feasibility of point-of-care testing. The PaCD diagnostic process takes only 30 min with a detection limit of 50 copies/test, making it suitable for analysis of sputum and throat swab samples. PaCD demonstrated 100% concordance (72/72) with next-generation sequencing and exhibited high concordance with computed tomography test results. These findings demonstrate the clinical feasibility of PaCD for the rapid and accurate diagnosis of infections caused by four prevalent respiratory pathogens, offering theoretical insights into the versatile application of point-of-care tests for the detection of other respiratory pathogens in various clinical scenarios.