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Development and in-use evaluation of a novel Luminex MicroPlex microsphere-based (TRIOL) assay for simultaneous identification of Mycobacterium tuberculosis and detection of first-line and second-line anti-tuberculous drug resistance in China
  1. Feifei Yin1,2,
  2. Jasper Fuk-Woo Chan3,4,5,6,
  3. Qixuan Zhu1,2,
  4. Ruijia Fu1,2,
  5. Jonathan Hon-Kwan Chen4,
  6. Garnet Kwan-Yue Choi4,
  7. Kah-Meng Tee4,
  8. Lihua Li1,2,
  9. Shiuyun Qian1,2,
  10. Wing-Cheong Yam3,4,5,6,
  11. Gang Lu1,2,
  12. Kwok-Yung Yuen3,4,5,6
  1. 1Key Laboratory of Translation Medicine Tropical Diseases, Department of Ministry of Education, Hainan Medical University, Haikou, Hainan, China
  2. 2Department of Pathogen Biology, Hainan Medical University, Haikou, Hainan, China
  3. 3State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China
  4. 4Department of Microbiology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China
  5. 5Research Centre of Infection and Immunology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China
  6. 6Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China
  1. Correspondence to Professor Kwok-Yung Yuen, State Key Laboratory of Emerging Infectious Diseases, Department of Microbiology, Carol Yu Centre for Infection, The University of Hong Kong, University Pathology Building, Queen Mary Hospital, Pokfulam, Hong Kong, Hong Kong; kyyuen{at}hku.hk or Professor Gang Lu, Key Laboratoy of Translation Medicine Tropical Diseases, Hainan Medical University, Ministry of Education, Haikou, Hainan, 571101, China; luganghn{at}aliyun.com

Abstract

Aims Rapid and accurate diagnostic assays with simultaneous microbial identification and drug resistance detection are essential for optimising treatment and control of tuberculosis.

Methods We developed a novel multiplex (TRIOL, Tuberculosis-Rifampicin-Isoniazid-Ofloxacin-Luminex) assay using the Luminex xMAP system that simultaneously identifies Mycobacterium tuberculosis and detects resistance to first-line and second-line anti-tuberculous drugs, and compared its performance with that by PCR sequencing, using phenotypic drug susceptibility testing as the gold standard.

Results Identification of M. tuberculosis by the TRIOL assay was highly sensitive (100%) and specific (100%). The overall drug-specific specificities were excellent (100%). The overall sensitivity of the TRIOL assay was lower than that of the PCR-sequencing assays (72.4% vs 82.8%) because of a lower sensitivity of detecting rifampicin resistance (71.4% vs 92.9%). The sensitivity of detecting isoniazid and ofloxacin resistance was as good as the PCR-sequencing assays. Importantly, the TRIOL assay did not miss any mutations that were included in the assay. All of the resistant isolates that were missed had uncommon mutations or unknown resistance mechanisms that were not included in the assay.

Conclusions The TRIOL assay has higher throughput, lower cost and is less labour intensive than the PCR-sequencing assays. The TRIOL assay is advantageous in having the capability to detect resistance to multiple drugs and an open-architecture system that allows addition of more specific primers to detect uncommon mutations. Inclusion of additional primers for the identification of non-tuberculous mycobacteria, spoligotyping and improvement of rifampicin resistance detection would enhance the use of the TRIOL assay in future clinical and epidemiological studies.

  • TUBERCULOSIS
  • MICROBIOLOGY
  • LABORATORY TESTS
  • INFECTIONS

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