Progress toward Developing Sensitive Non-Sputum-Based Tuberculosis Diagnostic Tests: the Promise of Urine Cell-Free DNA

MacLean, Emily; Nathavitharana, Ruvandhi R.

doi:10.1128/jcm.00706-21

Cited by 4 publications

(4 citation statements)

References 31 publications

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“…Another form of promising urine-based tests, albeit at an earlier development stage, is cell-free DNA detection. These are Mtb fragments released into the blood (where they can also be detected) and subsequently excreted in urine [ 107 , 108 ]; however, no commercial products exist.…”

Section: Confirmatory Toolsmentioning

confidence: 99%

Diagnosing Tuberculosis: What Do New Technologies Allow Us to (Not) Do?

Abdulgader¹,

Ojo-Okunola²,

Ndlangalavu³

et al. 2022

Respiration

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New tuberculosis (TB) diagnostics are at a crossroads: their development, evaluation, and implementation is severely damaged by resource diversion due to COVID-19. Yet several technologies, especially those with potential for non-invasive non-sputum-based testing, hold promise for efficiently triaging and rapidly confirming TB near point-of-care. Such tests are, however, progressing through the pipeline slowly and will take years to reach patients and health workers. Compellingly, such tests will create new opportunities for difficult-to-diagnose populations, including primary care attendees (all-comers in high burden settings irrespective of reason for presentation) and community members (with early stage disease or risk factors like HIV), many of whom cannot easily produce sputum. Critically, all upcoming technologies have limitations that implementers and health workers need to be cognizant of to ensure optimal deployment without undermining confidence in a technology that still offers improvements over the status quo. In this state-of-the-art review, we critically appraise such technologies for active pulmonary TB diagnosis. We highlight strengths, limitations, outstanding research questions, and how current and future tests could be used in the presence of these limitations and uncertainties. Among triage tests, CRP (for which commercial near point-of-care devices exist) and computer-aided detection software with digital chest X-ray hold promise, together with late-stage blood-based assays that detect host and/or microbial biomarkers; however, aside from a handful of prototypes, the latter category has a shortage of promising late-stage alternatives. Furthermore, positive results from new triage tests may have utility in people without TB; however, their utility for informing diagnostic pathways for other diseases is under-researched (most sick people tested for TB do not have TB). For confirmatory tests, few true point-of-care options will be available soon; however, combining novel approaches like tongue swabs with established tests like Ultra have short-term promise but first require optimizations to specimen collection and processing procedures. Concerningly, no technologies yet have compelling evidence of meeting the World Health Organization optimal target product profile performance criteria, especially for important operational criteria crucial for field deployment. This is alarming as the target product profile criteria are themselves almost a decade old and require urgent revision, especially to cater for technologies made prominent by the COVID-19 diagnostic response (e.g., at-home testing and connectivity solutions). Throughout the review, we underscore the importance of how target populations and settings affect test performance and how the criteria by which these tests should be judged vary by use case, including in active case finding. Lastly, we advocate for health workers and researchers to themselves be vocal proponents of the uptake of both new tests and those – already available tests that remain suboptimally utilized.

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Section: Confirmatory Toolsmentioning

confidence: 99%

Diagnosing Tuberculosis: What Do New Technologies Allow Us to (Not) Do?

Abdulgader¹,

Ojo-Okunola²,

Ndlangalavu³

et al. 2022

Respiration

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“…14 Atas dasar tersebut, beberapa peneliti mengembangkan pemeriksaan TB yang tidak menggunakan sputum melainkan urin. [15][16][17][18] Dalam urin penderita TB dapat ditemukan transrenal cell free DNA (tr-cfDNA) dari MTB. DNA tersebut dihasilkan dari pemecahan sel MTB kemudian masuk ke dalam aliran darah dan melewati glomerulus.…”

Section: Pendahuluanunclassified

“…Selama ini metode LAMP diaplikasikan untuk mendeteksi gen gyrB dari spesimen sputum, padahal beberapa penelitian melaporkan bahwa urin dapat dijadikan alternatif spesimen pemeriksaan. [15][16][17][18] Pada urin terdapat tr-cfDNA yang bisa dideteksi dengan metode berbasis molekuler dan dijadikan penanda diagnostik untuk pengobatan TB paru atau ekstra paru. Penelitian sebelumnya melaporkan bahwa pemeriksaan molekuler TB yang memanfaatkan tr-cfDNA memiliki nilai sensitivitas sebesar 83,7% dan spesifisitas sebesar 100%.…”

Section: Pembahasanunclassified

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OPTIMASI METODE LOOP MEDIATED ISOTHERMAL AMPLIFICATION UNTUK DETEKSI GEN gyrB Mycobacterium tuberculosis PADA SPESIMEN URIN

Naully,

Hidashanti,

Wasdili

2023

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Salah satu metode alternatif untuk pemeriksaan tuberkulosis (TB) adalah Loop Mediated Isothermal Amplification (LAMP) dengan gen gyrB sebagai penandanya. Spesimen yang umumnya digunakan untuk pemeriksaan tersebut adalah sputum, namun proses pengambilan dan penanganannya cukup sulit. Maka dari itu, beberapa penelitian menyarankan menggunakan spesimen urin. Tujuan penelitian ini adalah menentukan kondisi optimum metode LAMP dalam mengamplifikasi gen gyrB dari Mycobacterium tuberculosis (MTB) pada spesimen urin. Spesimen urin yang digunakan dalam penelitian ini berasal dari penderita TB di Puskesmas Cimahi Selatan. Penelitian diawali dengan isolasi deoxyribonucleic acid (DNA) dari urin, lalu gen gyrB diamplifikasi dengan metode LAMP. Amplifikasi dilakukan pada berbagai varian suhu yaitu 59-62 o C dan konsentrasi DNA template sebesar 25 ng/µL, 50 ng/µL, 100 ng/µL serta 200 ng/ µL. Amplikon divisualisasi dengan metode elektroforesis agarose. Hasil penelitian menunjukkan bahwa gen gyrB yang berukuran 320 bp hanya berhasil diamplifikasi dari DNA template dengan konsentrasi sebesar 100 ng/µL dan 200 ng/µL. Pita amplikon yang paling jelas dan tebal terlihat pada konsentrasi 200 ng/µL. Berdasarkan optimasi suhu amplifikasi diketahui bahwa gen gyrB berhasil diamplifikasi pada suhu 59-62 o C, namun pita yang paling tebal terbentuk pada suhu 61 o C. Berdasarkan hasil tersebut dapat disimpulkan bahwa gen gyrB dapat diamplifikasi secara optimum menggunakan metode LAMP pada suhu 61 o C dan dengan konsentrasi DNA template sebesar 200 ng/µL.

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