Abstract
This study identified urinary biomarkers for tuberculosis (TB) diagnosis. The urine proteomic profiles of 45 pulmonary tuberculosis patients prior to anti-TB treatment and 45 healthy controls were analyzed and compared using two-dimensional electrophoresis with matrix-assisted laser desorption/ionization time of flight mass spectrometry. Nineteen differentially expressed proteins were identified preliminarily, and western blotting and qRT-PCR were performed to confirm these changes at the translational and transcriptional levels, respectively, using samples from 122 additional pulmonary tuberculosis patients and 73 additional healthy controls. Two proteins, mannose-binding lectin 2 and a 35-kDa fragment of inter-α-trypsin inhibitor H4, exhibited the highest differential expression. We constructed a protein-microRNA interaction network that primarily involved complement and inflammatory responses. Eleven microRNAs from microRNA-target protein interactions were screened and validated using qRT-PCR with some of the above samples, including 97 pulmonary tuberculosis patients and 48 healthy controls. Only miR-625-3p exhibited significant differential expression (p < 0.05). miR-625-3p was increased to a greater extent in samples of smear-positive than smear-negative patients. miR-625-3p was predicted to target mannose-binding lectin 2 protein. A binary logistic regression model based on miR-625-3p, mannose-binding lectin 2, and inter-α-trypsin inhibitor H4 was further established. This three-biomarker combination exhibited better performance for tuberculosis diagnosis than individual biomarkers or any two-biomarker combination and generated a diagnostic sensitivity of 85.87% and a specificity of 87.50%. These novel urine biomarkers may significantly improve tuberculosis diagnosis.
Highlights
Tuberculosis (TB) is a chronic infectious zoonotic disease caused by Mycobacterium tuberculosis (Mtb), and it is a significant worldwide threat to human health
Statistical comparisons indicated that 19 spots were significantly different between pulmonary TB (PTB) patients and healthy controls, and these spots were identified using the default criteria of fold changes >1.50 and p < 0.05, which indicate a significant upregulation in PTB patients using an unpaired two-tailed t-test in the statistical tools built into the PD Quest software (Fig. 1)
A total of 173 and 102 protein spots were identified from the urine samples of PTB patients and healthy controls, respectively, using 2-DE on the immobilized pH gradient (IPG) strip
Summary
Tuberculosis (TB) is a chronic infectious zoonotic disease caused by Mycobacterium tuberculosis (Mtb), and it is a significant worldwide threat to human health. The. World Health Organization (WHO) reported that ~10.4 million people developed TB in 2016, and 1.7 million died, which makes TB the ninth leading cause of human death worldwide and the leading cause from a single-infectious agent[1]. TB diagnosis generally depends on clinical manifestations, tuberculin skin testing, smear microscopy, and bacterial culture isolation. Isolation of Mtb from clinical samples is the gold standard for TB diagnosis. This test is a very low sensitivity and time-consuming methodology, and it requires a significant biosafety requirement for facilities[3]. The most commonly used clinical assay uses blood to detect cell-mediated immunity (e.g., interferon gamma production).
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