Validate a two-step multiplex PCR in detecting pathogens of vertebral osteomyelitis. Focus on Mycobacterium tuberculosis

Chishih Chu, Tsung-Jen Huang, Paul Yann Lin, Yen-Yao Li, Meng-Huang Wu, Tsung-Han Yang, Chin-Chang Cheng, Ching-Yu Lee

Research output: Contribution to journalArticle

Abstract

Background: The key to achieve better clinical outcomes in treating vertebral osteomyelitis (VO) is to obtain a prompt and reliable bacterial culture. Detecting Mycobacterium tuberculosis (M. tuberculosis) in spine infection is difficult owing to slow growth of mycobacteria and the presence of fewer mycobacteria at the affected sites. The aim of this study was to validate the effectiveness of a 2-step multiplex polymerase chain reaction (M-PCR) assay in detecting M. tuberculosis in formalin-fixed paraffin-embedded (FFPE) vertebral specimens of VO. Methods: Through the Spine Operation Registry of the authors' institution, 43 patients with a diagnosis of VO and undergoing open spine surgery at the authors' hospital were retrospectively reviewed. Thirty-two patients, 12 TB- and 20 non-TB VO, who had a confirmed spine microbial culture at the affected site and a compatible histo-pathological finding were enrolled for study. Bacterial DNAs were extracted from vertebral FFPE blocks, the 2-step M-PCR assay was developed to identify bacterial or fungal infection first and then to identify mainly M. tuberculosis and Staphylococcus aureus of Gram-positive bacteria. All the FFPE vertebral specimens were blinded prior to the 2-step M-PCR study. Results: The non-TB VO cases were used as a control group. The current 2-step M-PCR assay in detecting M. tuberculosis was positive in 11 out of 12 TB VO and was positive in 2 out of 20 non-TB cases. The detection of M. tuberculosis was false negative in 1/12 (8.3%) and false positive in 2/20 (10%). It indicated a 91.7% of the sensitivity, 90% of the specificity, 84.6% of the positive predictive value, and 94.7% of the negative predictive value. The accuracy rate was 90.1%. Conclusion: Our results indicated that the current 2-step M-PCR assay is sensitive and effective in detecting M. tuberculosis in FFPE vertebral specimen from patients with VO. Potentially, the 2-step M-PCR can be applied as a quick and valuable supplementary tool for increasing the diagnostic accuracy of TB spondylitis.
Original languageEnglish
Pages (from-to)66-74
Number of pages9
JournalFormosan Journal of Musculoskeletal Disorders
Volume8
Issue number2
DOIs
Publication statusPublished - 2017

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Multiplex Polymerase Chain Reaction
Osteomyelitis
Mycobacterium tuberculosis
Paraffin
Formaldehyde
Spine
Mycobacterium
Spondylitis
Bacterial DNA
Spinal Dysraphism
Mycoses
Gram-Positive Bacteria
Bacterial Infections
Registries
Staphylococcus aureus
Sensitivity and Specificity
Control Groups
Growth
Infection

Keywords

  • formalin-fixed paraffin-embedded
  • multiplex polymerase chain reaction
  • tuberculosis
  • vertebral osteomyelitis

Cite this

Validate a two-step multiplex PCR in detecting pathogens of vertebral osteomyelitis. Focus on Mycobacterium tuberculosis. / Chu, Chishih; Huang, Tsung-Jen; Lin, Paul Yann; Li, Yen-Yao; Wu, Meng-Huang; Yang, Tsung-Han; Cheng, Chin-Chang; Lee, Ching-Yu.

In: Formosan Journal of Musculoskeletal Disorders, Vol. 8, No. 2, 2017, p. 66-74.

Research output: Contribution to journalArticle

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abstract = "Background: The key to achieve better clinical outcomes in treating vertebral osteomyelitis (VO) is to obtain a prompt and reliable bacterial culture. Detecting Mycobacterium tuberculosis (M. tuberculosis) in spine infection is difficult owing to slow growth of mycobacteria and the presence of fewer mycobacteria at the affected sites. The aim of this study was to validate the effectiveness of a 2-step multiplex polymerase chain reaction (M-PCR) assay in detecting M. tuberculosis in formalin-fixed paraffin-embedded (FFPE) vertebral specimens of VO. Methods: Through the Spine Operation Registry of the authors' institution, 43 patients with a diagnosis of VO and undergoing open spine surgery at the authors' hospital were retrospectively reviewed. Thirty-two patients, 12 TB- and 20 non-TB VO, who had a confirmed spine microbial culture at the affected site and a compatible histo-pathological finding were enrolled for study. Bacterial DNAs were extracted from vertebral FFPE blocks, the 2-step M-PCR assay was developed to identify bacterial or fungal infection first and then to identify mainly M. tuberculosis and Staphylococcus aureus of Gram-positive bacteria. All the FFPE vertebral specimens were blinded prior to the 2-step M-PCR study. Results: The non-TB VO cases were used as a control group. The current 2-step M-PCR assay in detecting M. tuberculosis was positive in 11 out of 12 TB VO and was positive in 2 out of 20 non-TB cases. The detection of M. tuberculosis was false negative in 1/12 (8.3{\%}) and false positive in 2/20 (10{\%}). It indicated a 91.7{\%} of the sensitivity, 90{\%} of the specificity, 84.6{\%} of the positive predictive value, and 94.7{\%} of the negative predictive value. The accuracy rate was 90.1{\%}. Conclusion: Our results indicated that the current 2-step M-PCR assay is sensitive and effective in detecting M. tuberculosis in FFPE vertebral specimen from patients with VO. Potentially, the 2-step M-PCR can be applied as a quick and valuable supplementary tool for increasing the diagnostic accuracy of TB spondylitis.",
keywords = "formalin-fixed paraffin-embedded, multiplex polymerase chain reaction, tuberculosis, vertebral osteomyelitis",
author = "Chishih Chu and Tsung-Jen Huang and Lin, {Paul Yann} and Yen-Yao Li and Meng-Huang Wu and Tsung-Han Yang and Chin-Chang Cheng and Ching-Yu Lee",
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T1 - Validate a two-step multiplex PCR in detecting pathogens of vertebral osteomyelitis. Focus on Mycobacterium tuberculosis

AU - Chu, Chishih

AU - Huang, Tsung-Jen

AU - Lin, Paul Yann

AU - Li, Yen-Yao

AU - Wu, Meng-Huang

AU - Yang, Tsung-Han

AU - Cheng, Chin-Chang

AU - Lee, Ching-Yu

PY - 2017

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N2 - Background: The key to achieve better clinical outcomes in treating vertebral osteomyelitis (VO) is to obtain a prompt and reliable bacterial culture. Detecting Mycobacterium tuberculosis (M. tuberculosis) in spine infection is difficult owing to slow growth of mycobacteria and the presence of fewer mycobacteria at the affected sites. The aim of this study was to validate the effectiveness of a 2-step multiplex polymerase chain reaction (M-PCR) assay in detecting M. tuberculosis in formalin-fixed paraffin-embedded (FFPE) vertebral specimens of VO. Methods: Through the Spine Operation Registry of the authors' institution, 43 patients with a diagnosis of VO and undergoing open spine surgery at the authors' hospital were retrospectively reviewed. Thirty-two patients, 12 TB- and 20 non-TB VO, who had a confirmed spine microbial culture at the affected site and a compatible histo-pathological finding were enrolled for study. Bacterial DNAs were extracted from vertebral FFPE blocks, the 2-step M-PCR assay was developed to identify bacterial or fungal infection first and then to identify mainly M. tuberculosis and Staphylococcus aureus of Gram-positive bacteria. All the FFPE vertebral specimens were blinded prior to the 2-step M-PCR study. Results: The non-TB VO cases were used as a control group. The current 2-step M-PCR assay in detecting M. tuberculosis was positive in 11 out of 12 TB VO and was positive in 2 out of 20 non-TB cases. The detection of M. tuberculosis was false negative in 1/12 (8.3%) and false positive in 2/20 (10%). It indicated a 91.7% of the sensitivity, 90% of the specificity, 84.6% of the positive predictive value, and 94.7% of the negative predictive value. The accuracy rate was 90.1%. Conclusion: Our results indicated that the current 2-step M-PCR assay is sensitive and effective in detecting M. tuberculosis in FFPE vertebral specimen from patients with VO. Potentially, the 2-step M-PCR can be applied as a quick and valuable supplementary tool for increasing the diagnostic accuracy of TB spondylitis.

AB - Background: The key to achieve better clinical outcomes in treating vertebral osteomyelitis (VO) is to obtain a prompt and reliable bacterial culture. Detecting Mycobacterium tuberculosis (M. tuberculosis) in spine infection is difficult owing to slow growth of mycobacteria and the presence of fewer mycobacteria at the affected sites. The aim of this study was to validate the effectiveness of a 2-step multiplex polymerase chain reaction (M-PCR) assay in detecting M. tuberculosis in formalin-fixed paraffin-embedded (FFPE) vertebral specimens of VO. Methods: Through the Spine Operation Registry of the authors' institution, 43 patients with a diagnosis of VO and undergoing open spine surgery at the authors' hospital were retrospectively reviewed. Thirty-two patients, 12 TB- and 20 non-TB VO, who had a confirmed spine microbial culture at the affected site and a compatible histo-pathological finding were enrolled for study. Bacterial DNAs were extracted from vertebral FFPE blocks, the 2-step M-PCR assay was developed to identify bacterial or fungal infection first and then to identify mainly M. tuberculosis and Staphylococcus aureus of Gram-positive bacteria. All the FFPE vertebral specimens were blinded prior to the 2-step M-PCR study. Results: The non-TB VO cases were used as a control group. The current 2-step M-PCR assay in detecting M. tuberculosis was positive in 11 out of 12 TB VO and was positive in 2 out of 20 non-TB cases. The detection of M. tuberculosis was false negative in 1/12 (8.3%) and false positive in 2/20 (10%). It indicated a 91.7% of the sensitivity, 90% of the specificity, 84.6% of the positive predictive value, and 94.7% of the negative predictive value. The accuracy rate was 90.1%. Conclusion: Our results indicated that the current 2-step M-PCR assay is sensitive and effective in detecting M. tuberculosis in FFPE vertebral specimen from patients with VO. Potentially, the 2-step M-PCR can be applied as a quick and valuable supplementary tool for increasing the diagnostic accuracy of TB spondylitis.

KW - formalin-fixed paraffin-embedded

KW - multiplex polymerase chain reaction

KW - tuberculosis

KW - vertebral osteomyelitis

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DO - 10.6492/FJMD.20160412

M3 - Article

VL - 8

SP - 66

EP - 74

JO - Formosan Journal of Musculoskeletal Disorders

JF - Formosan Journal of Musculoskeletal Disorders

SN - 2210-7940

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ER -