Usefulness of truenat-derived DNA from extrapulmonary specimens in direct detection of drug resistant tuberculosis by line probe assay

Abstract

Objectives: Isoniazid (INH) and second-line drug resistance (DR) detection through line probe assay (LPA) takes long in extrapulmonary (EP) specimens because culture growth needs to be obtained to perform deoxyribonucleic acid (DNA) extraction due to the paucibacillary nature of these specimens. Knowing the DR pattern at the earliest is key to success of the treatment. Delay in appropriate tuberculosis (TB) treatment in EP TB patients runs the risk of DR amplification, significant disease damage, and patient loss to follow-up. Here, LPA was attempted on truenat-derived DNA elute from EP specimens, which, in routine, is discarded after the truenat test, to determine drug sensitivity test (DST) for INH and, where necessary, for second-line drugs (Fluoroquinolones, Kanamycin, amikacin, and capreomycin). Material and Methods: Truenat, acid-fast bacilli culture, and fluorescent microscopy were performed on all EP samples that were received at the laboratory during June–September 2022. DNA elute that was left over from 59 truenat Mycobacterium tuberculosis (MTB) positive EP samples were subjected to Genotype MTBDR plus Ver 2.0 assay. Results: MTBDR plus assay (DNA elute) detected MTB and rifampicin (RIF) and INH DST in 47 samples (79.6%) having truenat MTB count of 7.8 × 102 colony-forming unit/milliliter and above. It also detected RIF DST in 65.2% truenat RIF indeterminate samples and DST for both RIF and INH in 60% of culture negative EP specimens. DST results by LPA (DNA elute) completely concorded with standard indirect LPA (on 21 culture isolates from smear-negative specimens). The MTBDRsl yield was however relatively low (11.1%), although second line LPA (SLLPA) was performed only on 9 first-line DR samples. Conclusions: Left-over truenat-derived DNA elute is a significant sample by-product that can significantly speed up and increase the yield of determination of MTB DST in EP samples for RIF and INH, the most critical drugs for TB treatment.