Clinical metagenomic sequencing for diagnosis of pulmonary tuberculosis
Introduction
Tuberculosis (TB), which is caused by the bacillus Mycobacterium tuberculosis (MTB), remains a leading cause of death from infectious disease. According to the 2019 World Health Organization global report, TB was one of the top 10 causes of death worldwide, ranking above acquired immune deficiency syndrome. There were an estimated 10.0 million newly diagnosed TB cases worldwide in 2018, with a high incidence and mortality in developing countries.1,2 Although effective treatment has been available for decades, the difficulty faced by physicians has been the issue of rapid and reliable diagnosis. Currently, the Xpert MTB/RIF assay (Cepheid, Sunnyvale, CA. USA) has been endorsed by the WHO for the initial diagnosis of TB suspects.3 However, the presumptive diagnosis of TB mainly relies on an examination of clinical symptoms, radiological findings, and acid-fast staining (AFS). The nonspecific symptoms and signs of pulmonary TB often overlap with common pulmonary infections, which may delay the diagnosis of pulmonary TB and increase TB-related death.4,5 Therefore, alternative rapid and hypothesis-free diagnostic approaches are urgently required to accurately discriminate pulmonary TB from other common pulmonary infections in the early stages of the diagnostic process.
Recent advances and lower costs of next-generation sequencing (NGS) technologies have allowed NGS to be applied to diagnostic microbiology, with the advantages of hypothesis-free, culture-independent and unbiased pathogen detection directly from clinical specimens.6 To date, clinical metagenomics next-generation sequencing (mNGS) has been applied in diagnosis of pathogens causing a variety of infectious syndromes, including respiratory tract infection,7,8 bloodstream infection,9,10 meningitis and encephalitis.11,12 However, published reports describing the usefulness of mNGS in pulmonary TB suspects are limited to individual patients or small, retrospective studies.13, 14, 15, 16 The question remains whether the diagnostic performance of clinical mNGS testing for pulmonary TB justifies its wider adoption by the healthcare community.
Here, we performed a prospective, single-center study involving hospitalized patients with suspicion of active pulmonary TB infections. This study was designed to evaluate the clinical performance and effect of the mNGS assay in comparison with conventional microbiological testing in patient-care scenarios in which the tests, including Xpert, solid media culture, acid-fast bacilli staining (AFS), are likely to be used.
Section snippets
Study population and specimen
As a tertiary hospital, the Infectious Diseases Hospital of Soochow University admitted suspected tuberculosis patients. We enrolled 110 patients with suspected tuberculosis who underwent bronchoscopy and were tested for both Xpert and mNGS of alveolar lavage fluid from June 1, 2019, to January 31, 2020.
The patient's attending physician judged the patient with suspected tuberculosis based on clinical manifestations and imaging tests. Patients' sputum test methods included AFS (Ziehl–Neelsen
Diagnosis and demographic baseline
A total of 116 patients initially suspected of having active TB infection underwent bronchoscopy during hospitalization and received clinical lavage fluid or other tissue specimens were obtained for Xpert and mNGS tests. Six patients, with 9 specimens total, were excluded, including 4 endobronchial ultrasound (EBUS) tissue specimens, 1 cerebrospinal fluid specimen, 1 pleural fluid specimen and 3 tissue specimens. Finally, 110 patients were enrolled, and 48 patients were eventually diagnosed
Discussion
In this study, we enrolled a cohort of 110 patients with suspicious pulmonary TB infection. Forty-eight patients were diagnosed with PTB, and the rest were non-TB patients. Compared with the non-tuberculosis patients, the patients with pulmonary tuberculosis had no specific clinical features. The utility of clinical symptoms for screening PTB is hard to characterize, with rather low sensitivity and unsatisfactory specificity.23 TSPOT-TB is a widely used indicator for detecting tuberculosis
Declaration of Competing Interest
We declare that we do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted.
Acknowledgments
We thank Meng Xiang from Shanghai Genskey Technology Co., Ltd for the technical support with NGS sample processing and Library construction.
This work was supported by grants from the Science and Technology Plan of Suzhou, China (SYS201778, X.Y.; SYSD2018193, C.S.; SYS2019111, M.W.; and SS201880, J.Z.), and High-level Health Talents in Jiangsu Province (LGY2019014, P.T.). Our research sponsors had no role in the research design, data collection, data analysis, data interpretation, or report
References (33)
- et al.
The WHO 2014 global tuberculosis report–further to go
Lancet Glob Health
(2015) WHO global progress report on tuberculosis elimination
Lancet Respir Med
(2020)- et al.
Next-generation sequencing combined with routine methods to detect the pathogens of encephalitis/meningitis from a Chinese tertiary pediatric neurology center
J Infect
(2019) - et al.
Community-acquired pneumonia and tuberculosis: differential diagnosis and the use of fluoroquinolones
Int J Infect Dis
(2014) Roadmap for rolling out Xpert MTB/RIF for rapid diagnosis of TB and MDR-TB
(2010)- et al.
A population-based survey of tuberculosis symptoms: how atypical are atypical presentations?
Clin Infect Dis
(2000) - et al.
Pulmonary tuberculosis presenting as community-acquired pneumonia
Respirology
(2006) - et al.
Clinical metagenomics
Nat Rev Genet
(2019) - et al.
Metagenomic sequencing detects respiratory pathogens in hematopoietic cellular transplant patients
Am J Respir Crit Care Med
(2018) - et al.
Viral pathogen detection by metagenomics and pan-viral group polymerase chain reaction in children with pneumonia lacking identifiable etiology
J Infect Dis
(2017)
Analytical and clinical validation of a microbial cell-free DNA sequencing test for infectious disease
Nat Microbiol
Evaluation of plasma microbial cell-free DNA sequencing to predict bloodstream infection in pediatric patients with relapsed or refractory cancer
JAMA Oncol
Clinical metagenomic sequencing for diagnosis of meningitis and encephalitis
N Engl J Med
Clinical evaluation of diagnosis efficacy of active mycobacterium tuberculosis complex infection via metagenomic next-generation sequencing of direct clinical samples
Front Cell Infect Microbiol
Same-day diagnostic and surveillance data for tuberculosis via whole-genome sequencing of direct respiratory samples
J Clin Microbiol
Rapid whole-genome sequencing of mycobacterium tuberculosis isolates directly from clinical samples
J Clin Microbiol
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These authors contributed equally to this work.