DOUBLE-STEP MULTIPLEX REAL TIME PCR WITH MELTING CURVE ANALYSIS FOR DETECTION AND DIFFERENTIATION OF MYCOBACTERIA IN SPUTUM.

Mycobacterium tuberculosis (M. tb) is a causative agent of tuberculosis, a worldwide public health problem. In recent years, the incidence of human mycobacterial infection due to species other than M. tb has increased. However, the lack of specific, rapid, and inexpensive methods for identification of mycobacterial species remains a pressing problem. A diagnostic test was developed for mycobacterial strain differentiation utilizing a double-step multiplex real time PCR together with melting curve analysis for identifying and distinguishing among M. tb, M. bovis BCG, other members of M. tb. complex, M. avium, and non-tuberculosis mycobacteria. The assay was tested using 167 clinical sputum samples in comparison with acid-fast staining and culturing. Using only the first step (step A) the assay achieved sensitivity and specificity of 81% and 95%, respectively. The detection limit was equivalent to 50 genome copies.

Levels of anti-topoisomerase I antibody correlated with short onset of cardiopulmonary involvement in Thai systemic sclerosis patients.

Anti-topoisomerase-I antibody (ATA) is associated with disease severity and internal organ involvement in patients with systemic sclerosis (SSc). The correlation between ATA levels and the clinical course of SSc is unclear. We aimed to determine the correlation between ATA level and survival time and the onset of internal organ fibrosis in SSc patients. This historical cohort study was conducted in adult SSc patients with quantitative tests of ATA between January 2019 and December 2022. Patients with overlap syndrome and no quantitative ATA test were excluded. According to the sample size calculation, and 10% compensated for missing data, a total of 153 patients were needed. The respective mean age on the study date and median ATA level was 59.9 ± 11.3 years and 370 U/mL (range 195-652). Most cases (107 cases; 69.9%) were the diffuse cutaneous SSc subset. According to a multivariable analysis, the ATA titer had a negative correlation with the onset of cardiac involvement (Rho - 0.47, p = 0.01), and had a positive correlation with skin thickness progression (Rho 0.39, p = 0.04). Eleven cases exhibited ATA levels < 7 U/mL and outlier ATA levels were excluded, 142 cases were included in the sensitivity analysis, and multivariable analysis showed the correlation between early onset of ILD and cardiac involvement (Rho - 0.43, p = 0.03 and Rho - 0.51, p = 0.01, respectively). The ATA level was correlated with neither the survival time nor the onset of renal crisis in both analyses. High ATA levels were correlated with a short onset of ILD and cardiac involvement and the presence of extensive skin tightness. Quantitative tests of ATA could serve as an effective tool for identifying patients at risk of an unfavorable prognosis.