Aptamer-Based TB Antigen Tests for the Rapid Diagnosis of Pulmonary Tuberculosis: Potential Utility in Screening for Tuberculosis

Abstract

Pulmonary tuberculosis is the most common manifestation of tuberculosis, and to this day, sputum smear microscopy remains the most widely used diagnostic test in resource-limited settings despite its suboptimal sensitivity. Here we report the development of two DNA aptamer-based diagnostic tests, namely aptamer linked immobilized sorbent assay (Aptamer ALISA) and electrochemical sensor (ECS), for the direct detection of a TB biomarker HspX in sputum. First we compared the performance of Aptamer ALISA with anti-HspX polyclonal antibody-based enzyme linked immunosorbent assay (Antibody ELISA) in a blinded study of 314 sputum specimens. Aptamer ALISA displayed a high sensitivity of 94.1% (95% CI 86.8–98%) as compared to 68.2% sensitivity (95% CI 57.2–77.9%) of Antibody ELISA (p-value < 0.05) using culture as the reference standard without compromising test specificity of 100%. Out of nine smear-negative culture-positive samples, six were positive by Aptamer ALISA and only two were detected by Antibody ELISA. ALISA detected as positive 80 of 85 culture-positive TB as compared to 57 of 81 diagnosed as TB by X-ray (p-value < 0.0001). These findings demonstrate the superiority of the aptamer-based test over smear microscopy, antibody-based ELISA, and chest X-ray for TB detection (p-value < 0.0001 for all). Further, we have developed a ∼30 min point-of-care ECS test that discriminates between tuberculous and nontuberculous sputum with a sensitivity of ∼92.3% and specificity of 91.2%. The tests developed in the current study cost ∼$1–3/test and have potential utility in active case finding in high-risk groups and screening for pulmonary TB among presumptive TB subjects.

Advances in detection of infectious agents by aptamer-based technologies

“…Tuberculosis [9] H63SL2-M6 HspX Aptamer linked immobilized sorbent assay and electrochemical sensor Tuberculosis [18] Thiolated MPT64 aptamer MPT64 Combination of aptamers and electrochemical impedance spectroscopy Tuberculosis [19] MAA I and MAA II MPT64 A sandwich-type electrochemical aptasensor Salmonella [10] LA27 LPS Truncated aptamer LA27 combined with GO-based FP assays Salmonella [20] Apt-E. coli Apt-S. typ E. coli S. typ Aptamer modified by fluorescent magnetic multifunctional nanoprobe (APT-FMNPs) S. aureus [21] Chimeric [33] HCVcp ssDNA aptamer HCVcp A plasmonic nanoplatform Human papillomavirus [34] HPV16 L1 ssRNA aptamer HPV16 L1 protein The laser desorption/ionization mass spectrometry method Human papillomavirus [35] HPV-07 HPV16 virus-like particle (VLP) Slow abscission rate-modified DNA aptamer Influenza [37] NP-specific aptamer, CTnI-specific aptamer Influenza nucleoprotein (NP) and cardiac troponin I (cTnI).…”

“…Specific aptamers for infectious microorganisms in the event of new epidemic outbreaks can be quickly obtained through efficient SELEX technology, such as the recent specific aptamers of SARS-CoV-2 have been obtained rapidly [8]. Once the specific aptamers of pathogenic microorganisms or their toxic molecules are identified, a variety of detection schemes can be designed based on the aptamers, including chemiluminescent, electrochemical or fluorescent detection according to the different detection signals [9][10][11][12]. In order to improve the sensitivity of aptamer-based detection, several research teams tried to develop signal amplification strategies.…”

<p>Differentiation of Intraspinal Tuberculosis and Metastatic Cancer Using Magnetic Resonance Imaging</p>

“…11 Although the lungs are the most common sites of primary cancer, the breast ranked first in the frequency of occurrence locations for primary cancer in this study, which may be due to the small sample size. 5,11,12 As for pathologic mechanisms for intraspinal metastasis, three probable mechanisms have been suggested: (1) hematogenous spread via arteries and/ or the vertebral venous plexus (Batson's venous plexus), (2) infiltration along with involved nerve roots, and (3) direct infiltration through the CSF. 11 Hematogenous spread is generally believed to be responsible for most cases.…”

“…Although the lungs are most commonly involved, extrapulmonary tuberculosis accounted for 15% of all tuberculosis cases according to the WHO in 2016. 1,2 As a common type of extrapulmonary tuberculosis, intraspinal tuberculosis occurs increasingly, although its accurate incidence is unknown, which is in the same situation as intraspinal metastasis. 3 Both intraspinal tuberculosis and metastatic cancer may be rare, but they are difficult to differentiate because of their similar presentations and the high negative incidence of microbiological test for tuberculosis in cerebrospinal fluid (CSF) and their prognoses are completely different.…”

<p>Structural switching electrochemical DNA aptasensor for the rapid diagnosis of tuberculous meningitis</p>

“…In recent years, functional nucleic acids known as aptamers (DNA or RNA) have demonstrated the ability to potentially overcome the limitations associated with antibodies. [21][22][23][24][25][26] Owing to these attributes, aptamers are regarded as a strong chemical rival of antibodies, with an ability to replace antibodies in all possible diagnostic formats. 22,27 Furthermore, in the last few years, aptamers have demonstrated their utility in the detection of diverse analytes ranging from small molecules to antigens to enzymes to hormones and even whole bacterial or cancer cells.…”