Comparative diagnostic performance of metagenomic and two targeted sequencing methods in lower respiratory infection

Lower respiratory tract infections are characterized by high morbidity and mortality, the latter associated with the low sensitivity and long turnaround time (TAT) of traditional diagnostic methods. Advances in next-generation sequencing (NGS) offers a promising solution, but in the face of so many different NGS products, how to use them appropriately remains a great challenge for clinicians. This study included 205 patients with suspected lower respiratory tract infections from the department of respiratory and critical care medicine, and collected their lower respiratory tract samples for metagenomic NGS (mNGS) and two different targeted NGS (tNGS), amplification-based tNGS and capture-based tNGS. We analyzed their microorganisms reported, and evaluated their detection performance based on the comprehensive clinical diagnosis. Compared to the two tNGS, mNGS showed significant higher cost ($840) and longer TAT (20 h). Conversely, it identified the highest number of species, totaling 80, compared to 71 species identified by capture-based tNGS and 65 species by amplification-based tNGS. When benchmarked against the comprehensive clinical diagnosis, the capture-based tNGS demonstrated significantly higher diagnostic performance than the other two NGS, with an accuracy of 93.17% and a sensitivity of 99.43%. However, it showed lower specificity compared to the amplification-based tNGS in identifying DNA virus (74.78% vs. 98.25%). The amplification-based tNGS exhibited a poor sensitivity for both gram-positive (40.23%) and gram-negative bacteria (71.74%). Moreover, tNGS was able to identify genotypes, antimicrobial resistance genes and virulence factors. In conclusion, mNGS is suited for the detection of rare pathogens; the capture-based tNGS is preferable for routine diagnostic testing; the amplification-based tNGS can be an alternative in situations requiring rapid results and constrained by limited resources.