Application of Metagenomic and Targeted Next-Generation Sequencing in Diagnosis of Pulmonary Tuberculosis in Bronchoalveolar Lavage Fluid

Zhen Yang,* Qian Liu,* Yi Hu, Shuang Geng, Ji-Xiang Ni Department of Respiratory and Critical Care Medicine, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, People’s Republic of China*These authors contributed equally to this workCorrespondence: Shuang Geng, Department of Respiratory and Critical Care Medicine, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, No. 26 Shengli Street, Jiangan District, Wuhan, Hubei, 430014, People’s Republic of China, Tel +86-13476192468, Email [email protected] Ji-Xiang Ni, Department of Respiratory and Critical Care Medicine, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, No. 26 Shengli Street, Jiangan District, Wuhan, Hubei, 430014, People’s Republic of China, Email [email protected]: To explore the application value of metagenomic next-generation sequencing (mNGS) and targeted next-generation sequencing (tNGS) in the diagnosis of pulmonary tuberculosis (PTB) in bronchoalveolar lavage fluid (BALF).Patients and Methods: Data from 202 patients with suspected PTB at Wuhan Central Hospital (Jan 2022 - Jan 2024) were retrospectively analyzed. BALF samples were collected and examined using mNGS and tNGS, comparing their sensitivity to traditional methods like acid-fast staining, TB culture, and TB-DNA. Mixed microbial species were identified from the BALF using mNGS and tNGS, and the pros and cons of tNGS were evaluated against mNGS.Results: Of the 202 patients evaluated, 94 were diagnosed with PTB. The BALF mNGS and tNGS exhibited a sensitivity of 77.66% and a specificity of 100%, with positive and negative predictive values of 100% and 83.72%, respectively, outperforming conventional diagnostic methods. It was possible to compare the AUC values of the ROC curves of the BALF mNGS and tNGS with the corresponding values of the other three assay methods (0.89 vs 0.56, p < 0.05), MTB culture (0.89 vs 0.71, p < 0.05), and TB-DNA (0.89 vs 0.68, p < 0.05). Additionally, these techniques identified mixed microbial species in 52.13% of the BALF samples. Although both mNGS and tNGS demonstrated similar diagnostic rates, tNGS proved to be faster, more cost-effective, and incorporated a tuberculosis-specific wall-breaking technology, thereby suggesting greater clinical utility.Conclusion: BALF mNGS and tNGS technologies quickly and accurately detect PTB patients with greater sensitivity and specificity than traditional MTB methods. While both mNGS and tNGS demonstrate enhanced capacity for polymicrobial detection, the clinical significance of co-detected microorganisms requires integration with clinical context to differentiate colonization from active infection. Compared to mNGS, tNGS provides distinct advantages in clinical utility.Keywords: metagenomic next-generation sequencing, targeted next-generation sequencing, pulmonary tuberculosis, bronchoalveolar lavage fluid, diagnosis