title: Development and in-use evaluation of a novel Luminex MicroPlex microsphere-based (TRIOL) assay for simultaneous identification ofMycobacterium tuberculosisand detection of first-line and second-line anti-tuberculous drug resistance in China.
abstract: OBJECTIVE
  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 identifiesMycobacterium tuberculosisand 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 ofM. tuberculosisby 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.
authors.affiliation: Key Laboratory of Translation Medicine Tropical Diseases, Department of Ministry of Education, Hainan Medical University, Haikou, Hainan, China.|Department of Pathogen Biology, Hainan Medical University, Haikou, Hainan, China.
authors.forename: Feifei
authors.initials: F
authors.lastname: Yin
authors.affiliation: State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Department of Microbiology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Research Centre of Infection and Immunology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.
authors.forename: Jasper Fuk-Woo
authors.initials: JF
authors.lastname: Chan
authors.affiliation: Key Laboratory of Translation Medicine Tropical Diseases, Department of Ministry of Education, Hainan Medical University, Haikou, Hainan, China.|Department of Pathogen Biology, Hainan Medical University, Haikou, Hainan, China.
authors.forename: Qixuan
authors.initials: Q
authors.lastname: Zhu
authors.affiliation: Key Laboratory of Translation Medicine Tropical Diseases, Department of Ministry of Education, Hainan Medical University, Haikou, Hainan, China.|Department of Pathogen Biology, Hainan Medical University, Haikou, Hainan, China.
authors.forename: Ruijia
authors.initials: R
authors.lastname: Fu
authors.affiliation: Department of Microbiology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.
authors.forename: Jonathan Hon-Kwan
authors.initials: JH
authors.lastname: Chen
authors.affiliation: Department of Microbiology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.
authors.forename: Garnet Kwan-Yue
authors.initials: GK
authors.lastname: Choi
authors.affiliation: Department of Microbiology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.
authors.forename: Kah-Meng
authors.initials: KM
authors.lastname: Tee
authors.affiliation: Key Laboratory of Translation Medicine Tropical Diseases, Department of Ministry of Education, Hainan Medical University, Haikou, Hainan, China.|Department of Pathogen Biology, Hainan Medical University, Haikou, Hainan, China.
authors.forename: Lihua
authors.initials: L
authors.lastname: Li
authors.affiliation: Key Laboratory of Translation Medicine Tropical Diseases, Department of Ministry of Education, Hainan Medical University, Haikou, Hainan, China.|Department of Pathogen Biology, Hainan Medical University, Haikou, Hainan, China.
authors.forename: Shiuyun
authors.initials: S
authors.lastname: Qian
authors.affiliation: State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Department of Microbiology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Research Centre of Infection and Immunology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.
authors.forename: Wing-Cheong
authors.initials: WC
authors.lastname: Yam
authors.affiliation: Key Laboratory of Translation Medicine Tropical Diseases, Department of Ministry of Education, Hainan Medical University, Haikou, Hainan, China.|Department of Pathogen Biology, Hainan Medical University, Haikou, Hainan, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Lu
authors.affiliation: State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Department of Microbiology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Research Centre of Infection and Immunology, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.|Carol Yu Centre for Infection, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong, China.
authors.forename: Kwok-Yung
authors.initials: KY
authors.lastname: Yuen
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002648:Child; D002681:China; D005260:Female; D006801:Humans; D008297:Male; D008826:Microbial Sensitivity Tests / Q000379:methods*; D008863:Microspheres; D008875:Middle Aged; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D018088:Tuberculosis, Multidrug-Resistant / Q000175:diagnosis*; D055815:Young Adult
keywords: INFECTIONS; LABORATORY TESTS; MICROBIOLOGY; TUBERCULOSIS