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BackgroundPerturbed iron homeostasis is a risk factor for tuberculosis (TB) progression and an indicator of TB treatment failure and mortality. Few studies have evaluated iron homeostasis as a TB diagnostic biomarker.MethodsWe recruited participants with TB, latent TB infection (LTBI), cured TB (RxTB), pneumonia (PN) and healthy controls (HCs). We measured serum levels of three iron biomarkers including serum iron, ferritin and transferrin, then established and validated our prediction model.ResultsWe observed and verified that the three iron biomarker levels correlated with patient status (TB, HC, LTBI, RxTB or PN) and with the degree of lung damage and bacillary load in patients with TB. We then built a TB prediction model, neural network (NNET), incorporating the data of the three iron biomarkers. The model showed good performance for diagnosis of TB, with 83% (95% CI 77 to 87) sensitivity and 86% (95% CI 83 to 89) specificity in the training data set (n=663) and 70% (95% CI 58 to 79) sensitivity and 92% (95% CI 86 to 96) specificity in the test data set (n=220). The area under the curves (AUCs) of the NNET model to discriminate TB from HC, LTBI, RxTB and PN were all >0.83. Independent validation of the NNET model in a separate cohort (n=967) produced an AUC of 0.88 (95% CI 0.85 to 0.91) with 74% (95% CI 71 to 77) sensitivity and 92% (95% CI 87 to 96) specificity.ConclusionsThe established NNET TB prediction model discriminated TB from HC, LTBI, RxTB and PN in a large cohort of patients. This diagnostic assay may augment current TB diagnostics.

Tuberculosis (TB) remains a major global public health problem. New immunization methods against TB are urgently needed. Plasmid DNA with a microneedle patch is a potentially attractive strategy to improve the immune effect. A DNA vaccine encoding the secreted protein Ag85B of Mycobacterium tuberculosis was immunized in the skin using microneedles, which can improve protective immunity compared to conventional intramuscular (IM) injection. There is no significant difference between microneedle patch (MNP) and IM immunization when the immunizing dose is low (4.2 μg). However, the results for detecting humoral immunity showed MNP immunization could better provoke an antibody response than IM when the dose is high (12.6 μg). A similar result was observed in cellular immune responses by measuring the cytokines in splenocytes. The effective protection of MNP can also be demonstrated by counting bacteria and analyzing the survival rate. This study indicated that DNA vaccination in the skin using dissolving microneedles may provide a new strategy against TB.

The role of MAIT cells in immunity against Mycobacterium tuberculosis infection in humans is still largely unexplored. In this study, we investigated the functional role of 4-1BB on MAIT cells. We found that 4-1BB was highly up-regulated on MAIT cells from tuberculous pleural effusions following Mtb antigen stimulation and its level of expression correlated with IFN-γ and IL-17 production. 4-1BB expression on MAIT cells in response to Mtb antigens was partially dependent on IL-2 and was associated with common γ chain receptor. By transcriptome sequencing, we identified numerous differentially expressed genes between 4-1BB- and 4-1BB+ MAIT cells. GO enrichment and KEGG pathway analysis of differentially expressed genes identified enriched pathways that included T-cell receptor and NF-κB signaling pathways. It is concluded that 4-1BB has the potential to be used as a biomarker to identify MAIT cells with enhanced IFN-γ and IL-17 responses that might be associated with tuberculosis infection control.

Polysaccharide nucleic acid fractions of bacillus Calmette–Guérin, termed BCG-PSN, have traditionally been used as immunomodulators in the treatment of dermatitis and allergic diseases. While the sales of injectable BCG-PSN have shown steady growth in recent years, no reports of using BCG-PSN powder or its immunotherapeutic effects exist. Here, BCG-PSN powder was applied directly to the skin to evaluate the immunotherapeutic effects on mice infected with Mycobacterium tuberculosis (MTB). In total, 34 μg of BCG-PSN powder could be loaded into a microneedle patch (MNP). Mice receiving BCG-PSN powder delivered via MNP exhibited significantly increased IFN-γ and TNF-α production in peripheral blood CD4 + T cells and improved pathological changes in their lungs and spleens compared to control group mice. The immunotherapeutic effect of BCG-PSN powder delivered via MNP was better than that delivered via intramuscular injection to some extent. Furthermore, MNPs eliminate the side effects of syringes, and this study demonstrated that BCG-PSN can be clinically administrated in powder form.

Diagnosis of cervical tuberculous lymphadenitis (CTL), the most commonly occurring form of extrapulmonary tuberculosis, remains as a challenge in clinic. Detection of the presence of Mycobacterium tuberculosis (Mtb) in fine needle aspiration cytology (FNAC) samples is one golden criterion to confirm the CTL diagnosis. Due to the non-specific clinical presentation, CTL might be confused with other lymph node enlargement diseases; therefore empirical treatment with non-anti-TB antibiotics is often initially administered. However, it is still unclear whether this diagnostic antibiotic treatment affects the positivity of Mtb detection in FNAC. The demographics and clinical characteristics of 732 lymph node enlargement patients who had underwent FNAC were retrospectively analyzed and 605 (82.65%) of them were diagnosed as CTL. A total of 279 CTL cases (279/605, 46.11%) with completion of three Mtb tests (AFB, NAAT, and Mtb culture) in FNAC samples were selected for analyzing the effect of empirical antibiotic treatment on the positivity of Mtb tests. Compared to CTL patients without antibiotic treatment prior to FNAC, patients received empirical non anti-TB treatment had significantly lower positivity for acid fast bacilli staining (adjusted OR 0.11, 95% CI 0.06-0.21), nucleic acid amplification test (NAAT) (adjusted OR 0.38, 95% CI 0.21-0.71), and Mtb culture (adjusted OR 0.11, 95% CI 0.06-0.19). In conclusion, this study demonstrated that empirical non anti-TB antibiotic treatment reduced the opportunity to confirm CTL by microbiological analysis. Patients with cervical lymph node enlargement should undergo FNAC for Mtb tests prior to initiation of empirical non anti-TB treatment.