Ergonomic Diagnostic Tool based on Chip Mini RT-PCR for Diagnosis of Pulmonary and Extra Pulmonary Tuberculosis

Abstract

Diagnosis of Tuberculosis (TB) is a challenging problem with the currently available conventional methods such as microscopy for Acid Fast Bacilli (AFB) and culture techniques.The diagnosis of Extra pulmonary tuberculosis (EPTB) is still more challenging due to the low yield of bacilli in the clinical specimens. Recently, a chip based Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) was introduced in India. This study was conducted for finding out the sensitivity and specificity of TrueNAT RT-PCR for the diagnosis of Pulmonary and EPTB at a Tertiary Care Hospital in Southern India. A total of 145 samples including both Pulmonary (80) and EPTB (65) were examined by Smear microscopy, culture on Lowenstein Jensen (LJ) medium and TruNat RT PCR. All the positive samples were confirmed by conventional PCR technique as well. Out of 80 Pulmonary samples such as sputum, bronchial wash and tracheal aspirate 41(51.25%) samples were positive in RT-PCR, 22 (27.5%) were positive in microscopy and 29 (36.25%) was positive for culture on LJ medium. Among the 65 EPTB samples such as pus, pleural fluid, Cerebro spinal fluid (CSF), ascitic fluid, Tissue, Peritoneal fluid, Pericardial fluid, Urine, synovial fluid, Fine needle aspiration cytology (FNAC) 38 (58.46%) were positive in RT-PCR, 08 (12.30%) were positive in microscopy and 31(47.69%) were positive for culture on Lowenstein Jensen (LJ) medium. The sensitivity and specificity of TrueNat RT-PCR for the diagnosis of Pulmonary tuberculosis (PTB) is 93.1% and 72.5 % and for EPTB is 96.77 %and 76.4 %, respectively. The newer diagnostic tool has a Turn Around Time (TAT) of less than 2 hours, can detect rifampicin resistance, longer shelf life, cost effective and can work from 2°C to 40°C. It is portable and comes with an inbuilt rechargeable battery which makes it a compatible equipment for any health care setup and out reach programs to detect and treat patients even in remote villages.

A prospective multicentre diagnostic accuracy study for the Truenat tuberculosis assays

“…Ltd. (Bangalore, India) developed three assays that utilize chip-based real-time micro PCR, two for detection of M. tuberculosis: the Truenat™ MTB (including the nrdB single copy target) and MTB Plus (including nrdZ and multi-copy IS6110 targets) assays; and one for the detection of RIF resistance: the MTB-RIF Dx reflex assay targeting the rpoB gene [11,12]. These assays can be run from the same DNA eluate [13][14][15][16], obtained from the automated bead-based Trueprep ® DNA extraction device that uses a universal cartridge-based system to extract DNA from 0.5 mL of sputum in under 20 minutes. The DNA eluate is loaded onto the chipbased Truelab ™ micro PCR device to detect the presence of M. tuberculosis DNA in approximately 40 minutes.…”

Comparative Study of Fast Plaque Assay and Real Time PCR for Detection of Mycobacterium tuberculosis in Pulmonary Samples

“…A better technique for diagnosis can be rapid, accurate and highly sensitive. RT-PCR has been proven to be the more useful molecular technique to diagnose pulmonary and extra-pulmonary TB due to the efficient processing of fluid and tissue samples of patients (Mangayarkarasi, et al, 2019). The Positivity rate in pulmonary samples is much higher in the case of Real-Time PCR Technique which is 44.44% when compared to Fast Plaque Assay which is 13.33% which tells that the Real-Time PCR Technique is a much better technique than Fast Plaque Assay.…”

Point-of-care diagnostics for infectious diseases: From methods to devices

“…Up to date, the diagnosis of tuberculosis is still a challenge in the clinical medicine by using available conventional methods and it is more difficult to detect extra pulmonary tuberculosis (EPTB) due to the low yield of bacilli in the clinical specimens. For the purpose of addressing this challenge, a chip based TrueNAT device using RT-PCR was introduced for the diagnosis of pulmonary tuberculosis (PTB) and EPTB [454] . A systematic evaluation of the device was conducted using a total of 145 clinical samples including both PTB (80) and EPTB (65).…”

“…All the results demonstrated that the sensitivity and specificity of TrueNAT device is 93.1% and 72.5% for the diagnosis of PTB and 96.77% and 76.4% for EPTB, respectively. Besides, the device was able to achieve fast detection of PTB and EPTB as well as rifampicin resistance within 2 h, which is portable and compatible for POC detection of Mycobacterium tuberculosis [454] . Moreover, an integrated continuous flow PCR (CF-PCR) and electrophoresis biochip based all-in-one microfluidic device was recently developed by Li and his coworkers for the detection of Porphyromonas gingivalis , Treponema denticola and Tannerella forsythia [455] .…”