Table 2:

Summary of assays currently available for genotypic DST in tuberculosis, and their genetic targets

	Xpert MTB/RIF (Cepheid, Sunnyvale, CA, USA)	Xpert MTB/RIF Ultra (Cepheid, Sunnyvale, CA, USA)	Truenat MTB-Rif Dx (MolBio Diagnostics, Goa, India)	GenoType MTBDRplus VER 2.0 (HAIN Lifescience/Bruker, Nehren, Germany)	Nipro NTM+MDRTB detection kit 2 (Nipro Corporation, Osaka, Japan)	GenoType MTBDRsl VER 2.0 (HAIN Lifescience/Bruker, Nehren, Germany)	Xpert MTB/XDR (Cepheid, Sunnyvale, CA, USA)	Nipro Genoscholar PZA-TB II (Nipro Corporation, Osaka, Japan)	Deeplex Myc-TB* (Genoscreen, Lille, France)	Next Gen-RDST* (Illumina, San Diego, CA, USA)	AmpliSeq TB*
Technology	Cartridge-based real-time PCR and molecular beacons	Cartridge-based real-time PCR and sloppy molecular beacons	Chip-based real-time PCR	Line probe assay	Line probe assay	Line probe assay	Cartridge-based real-time PCR and sloppy molecular beacons	Line probe assay	tNGS	tNGS	tNGS
Genotypic DST approach	Negative hybridisation of wild-type probes	Mutation-specific melting temperature shifts analysis	Melting temperature shifts analysis	Negative hybridisation of wild-type probes or positive hybridisation of specific mutation probes	Negative hybridisation of wild-type probes or positive hybridisation of specific mutation probes	Negative hybridisation of wild-type probes or positive hybridisation of specific mutation probes	Mutation-specific melting temperature shifts analysis	Negative hybridisation of wild-type probes	Specific identification of mutations by sequencing	Single molecule-overlapping reads	Specific identification of mutations by sequencing
First-line
Rifampicin (RR)	rpoB RRDR	rpoB RRDR	rpoB RRDR	rpoB RRDR	rpoB RRDR	..	..	..	rpoB fragment (incl. cod. 170,491, and RRDR)	rpoB RRDR	rpoB full gene
Isoniazid (MDR)	..	..	..	katG codon 315; inhA promoter region	katG codon 315 region; inhA promoter region	..	katG codon 315 region; inhA promoter region; ahpC promoter region; fabG1 codon 203 region	..	katG fragment (including codon 315); ahpC fragment (including promoter region); inhA fragment (including promoter region, and codon 94); fabG1 fragment (including promoter region)	katG fragment (including codon 315); inhA promoter region	katG full gene; inhA full gene
Group A
Levofloxacin and moxifloxacin (pre-XDR)	..	..	..	..	..	gyrA QRDR; gyrB QRDR	gyrA QRDR; gyrB QRDR	..	gyrA QRDR; gyrB QRDR	gyrA QRDR	gyrA full gene
Bedaquiline (XDR)	..	..	..	..	..	..	..	..	Rv0678 full gene	..	..
Linezolid (XDR)	..	..	..	..	..	..	..	..	rplC fragment; rrl fragment	..	..
Group B
Clofazimine	..	..	..	..	..	..	..	..	Rv0678 full gene	..	..
Cycloserine	..	..	..	..	..	..	..	..	..	..	..
Terizidone	..	..	..	..	..	..	..	..	..	..	..
Group C
Ethambutol	..	..	..	..	..	..	..	..	embB fragment (including codon 306)	..	embB full gene
Delamanid	..	..	..	..	..	..	..	..	..	..	..
Pretoman id	..	..	..	..	..	..	..	..	..	..	..
Pyrazinamide	..	..	..	..	..	..	..	pncA full gene	pncA full gene	..	pncA full gene
Imipenem-cilastatin	..	..	..	..	..	..	..	..	..	..	..
Meropenem	..	..	..	..	..	..	..	..	..	..	..
Amikacin	..	..	..	..	..	rrs position 1401, 1402, and 1484	rrs 1400 region	..	rrs full gene	rrs 1400 and 1484 regions	..
Streptomycin	..	..	..	..	..	..	..	..	rpsL fuIl gene; gidB full gene	..	rpsL full gene
Ethionamide and prothionamide	..	..	..	inhA promoter region	..	..	inhA promoter region	..	inhA fragment (including promoter region, and codon 94); fabG1 fragment (including promoter region); ethA full gene	..	..
P-aminosalicylic acid	..	..	..	..	..	..	..	..	..	..	..
No longer recommended
Kanamycin	..	..	..	..	..	rrs position 1401, 1402, and 1484; eis promoter region	rrs 1400 region; eis promoter region	..	rrs full gene; eis fragment (including promoter region)	rrs 1400 and 1484 regions; eis promoter region	eis full gene
Capreomycin	..	..	..	..	..	rrs position 1401, 1402, and 1484	rrs 1400 region	..	rrs full gene; tlyA full gene	rrs 1400 and 1484 regions	..

The table focuses on WHO-endorsed methods, and main tNGS assays currently available. Drugs are grouped according to WHO.⁷⁴ MDR, pre-XDR, and XDR tuberculosis are defined according to WHO.³ Target regions are indicative; for detailed positions covered please refer to the specific manufacturers’ instruction manuals. tNGS=targeted next-generation sequencing. DST=drug susceptibility testing. RR=rifampicin resistance. MDR=multidrug-resistant, pre-XDR=pre-extensively drug-resistant. XDR=extensively drug-resistant. RRDR=rifampicin resistance-determining region. QRDR=quinolone resistance-determining region.

^*None of the targeted next-generation sequencing approaches are currently endorsed by a WHO recommendation.