Estimating Limit of Detection of M. Tuberculosis DNA as an Analyte Released from a Chemically Coated Solid Matrix and Using a WHO-approved CB-NAAT Platform

Abstract

Krishna H. Goyani and Pratap N. Mukhopadhyaya

The limit of detection of Mtb (Mycobacterium tuberculosis) bacilli was evaluated by detection of released DNA from a chemically treated cellulose matrix. Loopful of the Mtb cells were suspended in phosphate buffer (pH 6.8), DNA was extracted from it and quantified using a commercial Mtb real time PCR quantitation kit. Synthetic sputum, prepared as the matrix of choice was mixed with defined number of Mtb bacilli and spotted onto the cellulose matrix. Mtb DNA from the matrix was released into a DNA-re- lease buffer and detected using a WHO-approved CB-NAAT detection platform. One hundred and fifty, 500 and 1000 but not 10 Mtb bacilli/mL of synthetic sputum could be detected by this protocol. The result underlined the potential of cellulose matrix as a potent, room temperature, Mtb-infected sputum transportation, storage and DNA-release device for sensitive detection of Mtb using Mtb Xpert Ultra CB-NAAT as the WHO-approved Mtb test of choice.

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