Genome Medicine (May 2022)

Rapid molecular diagnostics of tuberculosis resistance by targeted stool sequencing

  • Doctor B. Sibandze,
  • Alexander Kay,
  • Viola Dreyer,
  • Welile Sikhondze,
  • Qiniso Dlamini,
  • Andrew DiNardo,
  • Godwin Mtetwa,
  • Bhekumusa Lukhele,
  • Debrah Vambe,
  • Christoph Lange,
  • Muyalo Glenn Dlamini,
  • Tara Ness,
  • Rojelio Mejia,
  • Barbara Kalsdorf,
  • Jan Heyckendorf,
  • Martin Kuhns,
  • Florian P. Maurer,
  • Sindisiwe Dlamini,
  • Gugu Maphalala,
  • Stefan Niemann,
  • Anna Mandalakas

DOI
https://doi.org/10.1186/s13073-022-01054-6
Journal volume & issue
Vol. 14, no. 1
pp. 1 – 11

Abstract

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Abstract Background Stool is an important diagnostic specimen for tuberculosis in populations who struggle to provide sputum, such as children or people living with HIV. However, the culture of Mycobacterium tuberculosis (M. tuberculosis) complex strains from stool perform poorly. This limits the opportunity for phenotypic drug resistance testing with this specimen. Therefore, reliable molecular methods are urgently needed for comprehensive drug resistance testing on stool specimens. Methods We evaluated the performance of targeted next-generation sequencing (tNGS, Deeplex® Myc-TB) for the detection of mutations associated with M. tuberculosis complex drug resistance on DNA isolated from stool specimens provided by participants from a prospective cohort of patients treated for tuberculosis in Eswatini (n = 66; 56 with and 10 participants without M. tuberculosis complex DNA detected in stool by real-time quantitative PCR), and an independent German validation cohort of participants with culture-confirmed tuberculosis (n = 21). Results The tNGS assay detected M. tuberculosis complex DNA in 38 of 56 (68%) samples; for 28 of 38 (74%) samples, a full M. tuberculosis complex drug resistance prediction report was obtained. There was a high degree of concordance with sputum phenotypic drug susceptibility results (κ = 0.82). The ability to predict resistance was concentration-dependent and successful in 7/10 (70%), 18/25 (72%), and 3/21 (14%) of samples with stool PCR concentration thresholds of > 100 femtogram per microliter (fg/μl), 1 to 100 fg/μl, and < 1 fg/μl, respectively (p = 0.0004). The German cohort confirmed these results and demonstrated a similarly high concordance between stool tNGS and sputum phenotypic drug susceptibility results (κ = 0.84). Conclusions tNGS can identify drug resistance from stool provided by tuberculosis patients. This affords the opportunity to obtain critical diagnostic information for tuberculosis patients who struggle to provide respiratory specimens.

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