There is an urgent need for precise diagnosis to distinguish nontuberculous mycobacterial (NTM) diseases from pulmonary tuberculosis (PTB) and other respiratory diseases. The aim of this study is to evaluate the diagnostic performance of Interferon-gamma (IFN-γ) release assays (IGRAs), including antigen-specific peripheral blood-based quantitative T cell assay (T-SPOT.TB) and QuantiFERON-TB-Gold-Test (QFT-G), in differentiating NTM infections (N = 1,407) from culture-confirmed PTB (N = 1,828) and other respiratory diseases (N = 2,652). At specie level, 2.56%, 10.73%, and 16.49% of NTM-infected patients were infected by Mycobacterium kansasii, M. abscessus, and with M. avmm-intracellulare complex (MAC), respectively. Valid analyses of T-SPOT.TB (ESAT-6, CFP-10) and QFT-G were available for 37.03% and 85.79% in NTM-infected patients, including zero and 100% (36/36) of M. kansasii infection, 21.85% (33/151) and 92.05% (139/151) of M. abscessus infection, and 17.67% (41/232) and 91.24% (211/232) of MAC infection. Based on means comparisons and further ROC analysis, T-SPOT.TB and QFT-G performed moderate accuracy when discriminating NTM from PTB at modified cut-off values (ESAT-6 < 4 SFCs, CFP-10 < 3 SFCs, and QFT-G < 0.667 IU/ml), with corresponding AUC values of 0.7560, 0.7699, and 0.856. At species level of NTM, QFT-G effectively distinguished between MAC (AUC=0.8778), M. kansasii (AUC=0.8834) or M. abscessus (AUC=0.8783) than T-SPOT.TB. No significant differences in discriminatory power of these three IGRA tools were observed when differentiating NTM and Controls. Our results demonstrated that T-SPOT.TB and QFT-G were both efficient methods for differentiating NTM disease from PTB, and QFT-G possessed sufficient discriminatory power to distinguish infections by different NTM species.

Signal transducer and activator of transcription-3 (STAT3) plays an important role in biological balance. Our and others previous studies implied that STAT3 had a great effect on fast-acting innate immunity against tuberculosis (TB). We hypothesized that stat3 SNP down-regulation of STAT3 leads to a change in susceptibility to TB in humans. To test this hypothesis, we investigated STAT3 SNPs using SNP scan™ technique in a case-control study of TB patients (n = 470) and HC subjects (n = 356), and then conducted functional studies of them using cellular models. We found that SNPs in STAT3 3`-UTR of rs1053004 TT and rs1053005 AA genotypes or T-A haplotype were associated with susceptibility to TB or TB severity. While the TT/AA genotype correlated with the low constitutive expression of stat3 and IL-17A in PBMC, the variant stat3 of rs1053004-rs1053005 T-A haplotype indeed reduced stat3 expression in reporter assays. Interestingly, host PBMC expressing the rs1053005 AA genotype and low constitutive stat3 exhibited the reduced ability to mount fast-acting innate immunity against mycobacterial infection in cellular models. Finally, mechanistic experiments showed that the STAT3 down-regulation broadly depressed STAT3 downstream anti-mycobacterial activities involving VDR-related CAMP pathway as well as IL-32, iNOS and autophagy mechanisms, leading to an enhanced mycobacterial infection. The findings of this study suggest that low constitutive stat3 derived from the TT/AA genotype/T-A haplotype acts to down-regulate STAT3, depressing multiple anti-mycobacterial pathways/mechanisms downstream, which leads to an enhanced mycobacterial infection or TB in high-risk individuals.