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Celik M, Gurbuz E, Cicek Y, Buyuktuna SA, Gundag O, Gulderen Kuscu E, Mermutluoglu C, Alkan S, Yuruk Atasoy P, Yuksekkaya E, Sahinoglu MS, Sahin A, Parlak E, Akgul F, Dindar Demiray EK, Oz M, Ciftci EZ, Kirik Y, Arslan Y, Ceylan MR, Mert A. Demographic, clinical and laboratory characteristics of extrapulmonary tuberculosis: Eight-year results of a multicenter retrospective study in Turkey. J Investig Med 2025; 73:206-217. [PMID: 39508290 DOI: 10.1177/10815589241299367] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2024]
Abstract
Extrapulmonary tuberculosis (EPTB) is an important public health problem due to its diverse clinical presentations, diagnostic complexities, and significant impact on patient outcomes and public health. Our study aimed to understand the sociodemographic, clinical, and laboratory characteristics as well as diagnostic and treatment modalities of adult patients with EPTB. This is a multicentric retrospective study that covers patients with EPTB cases followed up from January 2015 to December 2022 among tuberculosis (TB) dispensaries and Infectious Diseases and Clinical Microbiology clinics of 15 hospitals located in various regions of Turkey. The study included 64.6% women with a mean age of 44 years and a mortality rate of 3.5% within 1 year of diagnosis. Initial constitutional symptoms were predominantly fatigue (57%) and anorexia (53.7%). The most commonly affected sites were the lymph nodes (49.1%) and pleura (9.7%). The lumbar region was particularly involved in cases with spinal TB. Diagnostic findings included acid-fast bacilli positivity in 27.5% of cases, tuberculosis polymerase chain reaction positivity in 41%, elevated adenosine deaminase levels in 91.2% (especially in pleural and peritoneal fluids), and mycobacterial culture positivity in 40.9%. Pathology slides showed granulomatous inflammation in 97.7%. Increased C-reactive protein (CRP) levels correlated with the number of organs affected. Anti-TB treatment-related hepatotoxicity was detected in 8.9% of patients. In this study, it is important to note that the lumbar region is predominantly affected with involvement in spinal region. CRP level was consistent with the number of organ involvements and was one of the most critical results of this study.
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Affiliation(s)
- Mehmet Celik
- Department of Infectious Diseases and Clinical Microbiology, University of Harran Faculty of Medicine, Sanliurfa, Turkey
| | - Esra Gurbuz
- Department of Infectious Diseases and Clinical Microbiology, SBU Van Training and Research Hospital, Van, Turkey
| | - Yeliz Cicek
- Department of Infectious Diseases and Clinical Microbiology, Bingol State Hospital, Bingol, Turkey
| | - Seyit Ali Buyuktuna
- Department of Infectious Diseases and Clinical Microbiology, Sivas Cumhuriyet University, Sivas, Turkey
| | - Omur Gundag
- Department of Infectious Diseases and Clinical Microbiology, Elazığ Fethi Sekin City Hospital, Elazıg, Turkey
| | - Evrim Gulderen Kuscu
- Department of Infectious Disease and Clinical Microbiology, Faculty of Medicine, Kahramanmaras Sütcü Imam University, Kahramanmaras, Turkey
| | - Cigdem Mermutluoglu
- Department of Infectious Disease and Clinical Microbiology, Faculty of Medicine, Dicle University, Diyarbakir, Turkey
| | - Sevil Alkan
- Department of Infectious Diseases and Clinical Microbiology, Canakkale Onsekiz Mart University Faculty of Medicine, Canakkale, Turkey
| | - Pınar Yuruk Atasoy
- Department of Infectious Diseases and Clinical Microbiology, Ankara City Hospital, Ankara, Turkey
| | - Esra Yuksekkaya
- Department of Infectious Diseases and Clinical Microbiology, Mehmet Akif İnan Training and Research Hospital, Sanliurfa, Turkey
| | - Mustafa Serhat Sahinoglu
- Department of Infectious Diseases and Clinical Microbiology, Manisa City Hospital, Manisa, Turkey
| | - Ahmet Sahin
- Department of Infectious Diseases and Clinical Microbiology, Dr. Ersin Arslan Training and Research Hospital, Gaziantep, Turkey
| | - Emine Parlak
- Department of Infectious Diseases and Clinical Microbiology, Erzurum Ataturk University, Erzurum, Turkey
| | - Fethiye Akgul
- Department of Infectious Diseases and Clinical Microbiology, Batman Training and Research Hospital, Batman, Turkey
| | | | - Murtaza Oz
- Department of Infectious Diseases and Clinical Microbiology, Sivas Numune Hospital, Sivas, Turkey
| | - Elif Zelal Ciftci
- Department of Infectious Diseases and Clinical Microbiology, Diyarbakir Dagkapi State Hospital, Diyarbakir, Turkey
| | - Yasemin Kirik
- Department of Infectious Diseases and Clinical Microbiology, Elazıg Fethi Sekin City Hospital, Elazıg, Turkey
| | - Yusuf Arslan
- Department of Infectious Diseases and Clinical Microbiology, Batman Training and Research Hospital, Batman, Turkey
| | - Mehmet Resat Ceylan
- Department of Infectious Diseases and Clinical Microbiology, University of Harran Faculty of Medicine, Sanliurfa, Turkey
| | - Ali Mert
- Department of Infectious Diseases and Clinical Microbiology, University of Medipol, Faculty of Medicine, Istanbul, Turkey
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Paul SK, Ahmed S, Chakrabortty R, Paul SK, Rahman MA. Miliary tuberculosis in an immune-competent Bangladeshi man-A case report. Clin Case Rep 2023; 11:e7516. [PMID: 37305888 PMCID: PMC10256868 DOI: 10.1002/ccr3.7516] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2022] [Revised: 03/11/2023] [Accepted: 05/28/2023] [Indexed: 06/13/2023] Open
Abstract
Miliary tuberculosis is a disseminated and active form of tuberculosis caused by Mycobacterium tuberculosis. It frequently affects immunocompromised patients. However, immune-competent hosts are reported rarely. Herein, we reported a case of miliary tuberculosis of a 40-year-old immune-competent Bangladeshi man presented with pyrexia of unknown origin.
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Affiliation(s)
- Susanta Kumar Paul
- Department of Respiratory MedicineBangabandhu Sheikh Mujib Medical UniversityDhakaBangladesh
| | - Shamim Ahmed
- Department of Respiratory MedicineBangabandhu Sheikh Mujib Medical UniversityDhakaBangladesh
| | - Rajashish Chakrabortty
- Department of Respiratory MedicineBangabandhu Sheikh Mujib Medical UniversityDhakaBangladesh
| | - Shamrat Kumar Paul
- Department of Physics and AstronomyClemson UniversityClemsonSouth CarolinaUSA
| | - Mohammed Atiqur Rahman
- Department of Respiratory MedicineBangabandhu Sheikh Mujib Medical UniversityDhakaBangladesh
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Campelo TA, Cardoso de Sousa PR, Nogueira LDL, Frota CC, Zuquim Antas PR. Revisiting the methods for detecting Mycobacterium tuberculosis: what has the new millennium brought thus far? Access Microbiol 2021; 3:000245. [PMID: 34595396 PMCID: PMC8479963 DOI: 10.1099/acmi.0.000245] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2021] [Accepted: 06/17/2021] [Indexed: 01/07/2023] Open
Abstract
Tuberculosis (TB) affects around 10 million people worldwide in 2019. Approximately 3.4 % of new TB cases are multidrug-resistant. The gold standard method for detecting Mycobacterium tuberculosis, which is the aetiological agent of TB, is still based on microbiological culture procedures, followed by species identification and drug sensitivity testing. Sputum is the most commonly obtained clinical specimen from patients with pulmonary TB. Although smear microscopy is a low-cost and widely used method, its sensitivity is 50-60 %. Thus, owing to the need to improve the performance of current microbiological tests to provide prompt treatment, different methods with varied sensitivity and specificity for TB diagnosis have been developed. Here we discuss the existing methods developed over the past 20 years, including their strengths and weaknesses. In-house and commercial methods have been shown to be promising to achieve rapid diagnosis. Combining methods for mycobacterial detection systems demonstrates a correlation of 100 %. Other assays are useful for the simultaneous detection of M. tuberculosis species and drug-related mutations. Novel approaches have also been employed to rapidly identify and quantify total mycobacteria RNA, including assessments of global gene expression measured in whole blood to identify the risk of TB. Spoligotyping, mass spectrometry and next-generation sequencing are also promising technologies; however, their cost needs to be reduced so that low- and middle-income countries can access them. Because of the large impact of M. tuberculosis infection on public health, the development of new methods in the context of well-designed and -controlled clinical trials might contribute to the improvement of TB infection control.
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Affiliation(s)
- Thales Alves Campelo
- Faculdade de Medicina, Departamento de Patologia e Medicina Legal, Federal University of Ceará, Fortaleza, Brazil
| | | | - Lucas de Lima Nogueira
- Faculdade de Medicina, Departamento de Patologia e Medicina Legal, Federal University of Ceará, Fortaleza, Brazil
| | - Cristiane Cunha Frota
- Faculdade de Medicina, Departamento de Patologia e Medicina Legal, Federal University of Ceará, Fortaleza, Brazil
| | - Paulo Renato Zuquim Antas
- Laboratório de Imunologia Clínica, Instituto Oswaldo Cruz, Oswaldo Cruz Foundation, Rio de Janeiro, Brazil
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Kim C, Lee S, Kim J. Spinal epidural abscess due to coinfection of bacteria and tuberculosis: A case report. World J Clin Cases 2021; 9:4072-4080. [PMID: 34141768 PMCID: PMC8180206 DOI: 10.12998/wjcc.v9.i16.4072] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/16/2021] [Revised: 02/23/2021] [Accepted: 03/24/2021] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Spinal epidural abscess (SEA) is a rare condition that mostly results from infection with either bacteria or tuberculosis. However, coinfection with bacteria and tuberculosis is extremely rare, and it results in delays in diagnosis and antimicrobial treatment causing unfavorable outcomes.
CASE SUMMARY A 75-year-old female visited the hospital with low back pain, and magnetic resonance imaging (MRI) revealed an SEA at the lumbosacral segment. Staphylococcus hominis and methicillin-resistant Staphylococcus epidermidis were identified from preoperative blood culture and intraoperative abscess culture, respectively. Thus, the patient underwent treatment with vancomycin medication for 9 wk after surgical drainage of the SEA. However, the low back pain recurred 2 wk after vancomycin treatment. MRI revealed an aggravated SEA in the same area in addition to erosive destruction of vertebral bodies. Second surgery was performed for SEA removal and spinal instrumentation. The microbiological study and pathological examination confirmed Mycobacterium tuberculosis as the pathogen concurrent with the bacterial SEA. The patient improved completely after 12 mo of antitubercular medication.
CONCLUSION We believe that the identification of a certain pathogen in SEAs does not exclude coinfection with other pathogens. Tubercular coinfection should be suspected if an SEA does not improve despite appropriate antibiotics for the identified pathogen.
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Affiliation(s)
- Choonghyo Kim
- Department of Neurosurgery, Kangwon National University School of Medicine, Chuncheon 24341, Gangwon, South Korea
- Department of Neurosurgery, Kangwon National University Hospital, Chuncheon 24289, Gangwon, South Korea
| | - Seungkoo Lee
- Department of Anatomic Pathology, Kangwon National University School of Medicine, Chuncheon 24341, Gangwon, South Korea
- Department of Anatomic Pathology, Kangwon National University Hospital, Chuncheon 24289, Gangwon, South Korea
| | - Jiha Kim
- Department of Neurosurgery, Kangwon National University School of Medicine, Chuncheon 24341, Gangwon, South Korea
- Department of Neurosurgery, Kangwon National University Hospital, Chuncheon 24289, Gangwon, South Korea
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Natarajan A, Beena PM, Devnikar AV, Mali S. A systemic review on tuberculosis. Indian J Tuberc 2020; 67:295-311. [PMID: 32825856 DOI: 10.1016/j.ijtb.2020.02.005] [Citation(s) in RCA: 123] [Impact Index Per Article: 24.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2018] [Revised: 09/07/2019] [Accepted: 02/18/2020] [Indexed: 12/17/2022]
Abstract
Tuberculosis (TB), which is caused by bacteria of the Mycobacterium tuberculosis complex, is one of the oldest diseases known to affect humans and a major cause of death worldwide. Tuberculosis continues to be a huge peril disease against the human population and according to WHO, tuberculosis is a major killer of the human population after HIV/AIDS. Tuberculosis is highly prevalent among the low socioeconomic section of the population and marginalized sections of the community. In India, National strategic plan (2017-2025) has a national goal of elimination of tuberculosis by 2025. It requires increased awareness and understanding of Tuberculosis. In this review article history, taxonomy, epidemiology, histology, immunology, pathogenesis and clinical features of both pulmonary tuberculosis (PTB) and extra-pulmonary tuberculosis (EPTB) has been discussed. A great length of detailed information regarding diagnostic modalities has been explained along with diagnostic algorithm for PTB and EPTB. Treatment regimen for sensitive, drug resistant and extensive drug resistant tuberculosis has been summarized along with newer drugs recommended for multi drug resistant tuberculosis. This review article has been written after extensive literature study in view of better understanding and to increase awareness regarding tuberculosis, as a sincere effort that will help eliminate tuberculosis off the face of the earth in near future.
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MESH Headings
- Humans
- Algorithms
- Culture Techniques
- Extensively Drug-Resistant Tuberculosis
- History, 15th Century
- History, 16th Century
- History, 17th Century
- History, 18th Century
- History, 19th Century
- History, 20th Century
- History, Ancient
- Interferon-gamma Release Tests
- Mycobacterium tuberculosis
- Nucleic Acid Amplification Techniques
- Polymerase Chain Reaction
- Tuberculin Test
- Tuberculosis/diagnosis
- Tuberculosis/epidemiology
- Tuberculosis/history
- Tuberculosis/immunology
- Tuberculosis, Multidrug-Resistant
- Tuberculosis, Pulmonary/diagnosis
- Tuberculosis, Pulmonary/epidemiology
- Tuberculosis, Pulmonary/history
- Tuberculosis, Pulmonary/immunology
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Affiliation(s)
- Arvind Natarajan
- Department of Microbiology, Sri Devaraj Urs Medical College, Sri Devaraj Urs Academy of Higher Education and Research, Kolar, India
| | - P M Beena
- Department of Microbiology, Sri Devaraj Urs Medical College, Sri Devaraj Urs Academy of Higher Education and Research, Kolar, India
| | - Anushka V Devnikar
- Department of Microbiology, S Nijalingappa Medical College, Bagalkot, India
| | - Sagar Mali
- SDM Narayanaya Heart Centre, Sri Dharmasthala Manjunatheshwara Medical College, Sri Dharmasthala Manjunatheshwara University, Dharwad, India.
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Li QH, Zhang Y, Zhao MM, Gu Y, Hu Y, Su YL, Zhang F, Shen L, Zhou Y, Li HP. Simultaneous amplification and testing method forMycobacterium tuberculosisrRNA to differentiate sputum-negative tuberculosis from sarcoidosis. Am J Physiol Lung Cell Mol Physiol 2019; 316:L519-L524. [PMID: 30652492 DOI: 10.1152/ajplung.00172.2018] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
We use the simultaneous application and testing method to detect Mycobacterium tuberculosis rRNA (SAT-TB) with the endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) biopsy specimens to differentiate sputum-negative tuberculosis from sarcoidosis. In the first part, we validated the SAT-TB on the bronchial or EBUS-TBNA biopsy specimens from sputum smear-positive pulmonary tuberculosis. In the second part, all EBUS-TBNA specimens for sputum smear-negative intrathoracic tuberculous lymphadenopathies or sarcoidosis were tested with the SAT-TB, acid-fast bacilli smear, and culture. In the 16 sputum-positive tuberculosis cases, 5 showed negative SAT (2 nontuberculous mycobacteria and 3 had anti-tuberculosis therapies previously); the remaining 11 were positive. Of the 41 sputum-negative tuberculosis cases in the second part, five other diseases were negative. In the remaining 36 cases, 27 sarcoidosis cases were negative; 7 in 9 with sputum-negative tuberculosis were positive (77.78%). In these 36 patients, the sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of the SAT method were 77.78, 100, 100, 93.10, and 94.44%, respectively. The SAT distinguished sputum-negative tuberculosis from sarcoidosis significantly ( P < 0.0001) and identified cases with active M. tuberculosis as accurately as the conventional methods (κ = 0.912, P < 0.0001). We conclude that the SAT-TB may be an effective method for using biopsy specimens to differentiate sputum-negative tuberculosis from sarcoidosis.
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Affiliation(s)
- Qiu-Hong Li
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Yuan Zhang
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Meng-Meng Zhao
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Ye Gu
- Department of Endoscope, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Yang Hu
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Yi-Liang Su
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Fen Zhang
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Li Shen
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Ying Zhou
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
| | - Hui-Ping Li
- Department of Respiratory Medicine, Shanghai Pulmonary Hospital, Tongji University, School of Medicine, Shanghai, China
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Mohd Bakhori N, Yusof NA, Abdullah J, Wasoh H, Md Noor SS, Ahmad Raston NH, Mohammad F. Immuno Nanosensor for the Ultrasensitive Naked Eye Detection of Tuberculosis. SENSORS 2018; 18:s18061932. [PMID: 29899214 PMCID: PMC6022021 DOI: 10.3390/s18061932] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/12/2018] [Revised: 05/08/2018] [Accepted: 05/20/2018] [Indexed: 12/23/2022]
Abstract
In the present study, a beneficial approach for the ultrasensitive and affordable naked eye detection and diagnosis of tuberculosis (TB) by utilizing plasmonic enzyme-linked immunosorbent assay (ELISA) via antibody-antigen interaction was studied. Here, the biocatalytic cycle of the intracellular enzymes links to the formation and successive growth of the gold nanoparticles (GNPs) for ultrasensitive detection. The formation of different colored solutions by the plasmonic nanoparticles in the presence of enzyme labels links directly to the existence or non-existence of the TB analytes in the sample solutions. For disease detection, the adapted protocol is based mainly on the conventional ELISA procedure that involves catalase-labeled antibodies, i.e., the enzymes consume hydrogen peroxide and further produce GNPs with the addition of gold (III) chloride. The amount of hydrogen peroxide remaining in the solution determines whether the GNPs solution is to be formed in the color blue or the color red, as it serves as a confirmation for the naked eye detection of TB analytes. However, the conventional ELISA method only shows tonal colors that need a high concentration of analyte to achieve high confidence levels for naked eye detection. Also, in this research, we proposed the incorporation of protein biomarker, Mycobacterium tuberculosis ESAT-6-like protein esxB (CFP-10), as a means of TB detection using plasmonic ELISA. With the use of this technique, the CFP-10 detection limit can be lowered to 0.01 µg/mL by the naked eye. Further, our developed technique was successfully tested and confirmed with sputum samples from patients diagnosed with positive TB, thereby providing enough evidence for the utilization of our technique in the early diagnosis of TB disease.
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Affiliation(s)
- Noremylia Mohd Bakhori
- Institute of Advanced Technology, Universiti Putra Malaysia, 43400 UPM Serdang, Malaysia.
| | - Nor Azah Yusof
- Institute of Advanced Technology, Universiti Putra Malaysia, 43400 UPM Serdang, Malaysia.
- Department of Chemistry, Faculty of Science, Universiti Putra Malaysia, 43400 UPM Serdang, Malaysia.
| | - Jaafar Abdullah
- Department of Chemistry, Faculty of Science, Universiti Putra Malaysia, 43400 UPM Serdang, Malaysia.
| | - Helmi Wasoh
- Faculty of Biotechnology and Biomolecule Science, Universiti Putra Malaysia, 43400 UPM Serdang, Malaysia.
| | - Siti Suraiya Md Noor
- School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian, 16150 Kelantan, Malaysia.
| | - Nurul Hanun Ahmad Raston
- School of Biosciences and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan, Malaysia, 43600 UKM Bangi, Malaysia.
| | - Faruq Mohammad
- Surfactant Research Chair, Department of Chemistry, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia.
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Kanniappan P, Ahmed SA, Rajasekaram G, Marimuthu C, Ch'ng ES, Lee LP, Raabe CA, Rozhdestvensky TS, Tang TH. RNomic identification and evaluation of npcTB_6715, a non-protein-coding RNA gene as a potential biomarker for the detection of Mycobacterium tuberculosis. J Cell Mol Med 2017; 21:2276-2283. [PMID: 28756649 PMCID: PMC5618688 DOI: 10.1111/jcmm.13148] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2016] [Accepted: 02/01/2017] [Indexed: 01/15/2023] Open
Abstract
Technological advances in RNA biology greatly improved transcriptome profiling during the last two decades. Besides the discovery of many small RNAs (sRNA) that are involved in the physiological and pathophysiological regulation of various cellular circuits, it becomes evident that the corresponding RNA genes might also serve as potential biomarkers to monitor the progression of disease and treatment. sRNA gene candidate npcTB_6715 was previously identified via experimental RNomic (unpublished data), and we report its application as potential biomarker for the detection of Mycobacterium tuberculosis (MTB) in patient samples. For proof of principle, we developed a multiplex PCR assay and report its validation with 500 clinical cultures, positive for Mycobacteria. The analysis revealed 98.9% sensitivity, 96.1% specificity, positive and negative predictive values of 98.6% and 96.8%, respectively. These results underscore the diagnostic value of the sRNA gene as diagnostic marker for the specific detection of MTB in clinical samples. Its successful application and the general ease of PCR‐based detection compared to standard bacterial culture techniques might be the first step towards ‘point‐of‐care’ diagnostics of Mycobacteria. To the best of our knowledge, this is the first time for the design of diagnostic applications based on sRNA genes, in Mycobacteria.
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Affiliation(s)
- Priyatharisni Kanniappan
- Advanced Medical & Dental Institute (AMDI), Universiti Sains Malaysia, Kepala Batas, Penang, Malaysia.,Department of Pathology, Johor Bahru General Hospital, Johor, Malaysia
| | - Siti Aminah Ahmed
- Advanced Medical & Dental Institute (AMDI), Universiti Sains Malaysia, Kepala Batas, Penang, Malaysia
| | | | - Citartan Marimuthu
- Advanced Medical & Dental Institute (AMDI), Universiti Sains Malaysia, Kepala Batas, Penang, Malaysia
| | - Ewe Seng Ch'ng
- Advanced Medical & Dental Institute (AMDI), Universiti Sains Malaysia, Kepala Batas, Penang, Malaysia
| | - Li Pin Lee
- Advanced Medical & Dental Institute (AMDI), Universiti Sains Malaysia, Kepala Batas, Penang, Malaysia
| | - Carsten A Raabe
- Institute of Experimental Pathology (ZMBE), University of Muenster, Münster, Germany.,Institute of Evolutionary and Medical Genomics, Brandenburg Medical School (MHB), Neuruppin, Germany.,Institute of Medical Biochemistry, Centre for Molecular Biology of Inflammation (ZMBE), University of Muenster, Münster, Germany
| | | | - Thean Hock Tang
- Advanced Medical & Dental Institute (AMDI), Universiti Sains Malaysia, Kepala Batas, Penang, Malaysia
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Ghosh GC, Sharma B, Gupta BB. CSF ADA Determination in Early Diagnosis of Tuberculous Meningitis in HIV-Infected Patients. SCIENTIFICA 2016; 2016:5820823. [PMID: 27144055 PMCID: PMC4837278 DOI: 10.1155/2016/5820823] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/12/2015] [Accepted: 03/20/2016] [Indexed: 05/17/2023]
Abstract
Tuberculous and Cryptococcal meningitis are common in HIV patients. A highly specific and sensitive rapid test for diagnosis of Tuberculous meningitis especially in setting of HIV is not available in developing countries where the burden of disease is high. We measured ADA (adenosine deaminase) levels using spectrophotometric method in the CSF of HIV patients with meningitis to differentiate Tuberculous meningitis from meningitis due to other causes. Kruskal-Wallis test was used to compare ADA values between tuberculous meningitis (TBM) and nontuberculous (non-TB) meningitis patients and a receiver-operating characteristic (ROC) analysis curve was drawn from these values. Levels of ADA in the CSF of patients with TBM were significantly higher than those in patients with meningitis due to other causes. CSF ADA level determination with a cut-off value of 6 IU/L was found to be highly specific and fairly sensitive test for the diagnosis of TBM in HIV positive patients.
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Affiliation(s)
- Gopal Chandra Ghosh
- Hospital Annexe, Christian Medical College, Hospital Campus, Room No. 310, Vellore 632004, India
- *Gopal Chandra Ghosh:
| | - Brijesh Sharma
- PGIMER and Dr. RML Hospital, Main Block, New Delhi 100001, India
| | - B. B. Gupta
- PGIMER and Dr. RML Hospital, Main Block, New Delhi 100001, India
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10
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Tiwari D, Haque S, Tiwari RP, Jawed A, Govender T, Kruger HG. Fast and efficient detection of tuberculosis antigens using liposome encapsulated secretory proteins of Mycobacterium tuberculosis. JOURNAL OF MICROBIOLOGY, IMMUNOLOGY, AND INFECTION = WEI MIAN YU GAN RAN ZA ZHI 2015; 50:189-198. [PMID: 26231299 DOI: 10.1016/j.jmii.2015.05.014] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/15/2015] [Revised: 04/19/2015] [Accepted: 05/28/2015] [Indexed: 11/25/2022]
Abstract
PURPOSE A rapid and efficient diagnostic test was developed for the detection of Mycobacterium tuberculosis antigens in serum samples of active tuberculosis (TB) and extrapulmonary TB patients via a liposomal agglutination-based method. METHODS A rapid card test has been developed to facilitate the recognition of high-affinity binding rabbit raised purified culture filtrate protein antibodies coupled on the surface of activated liposomal preparation. In the presence of TB antigens, the polyclonal antibodies bound to the liposomal particles demonstrate a visible agglutination reaction. RESULTS The developed assay was simple, rapid, reliable, sensitive, and specific as a diagnostic test for the detection of antigens in serum samples of clinically confirmed cases of TB within 4-5 minutes' duration. The test was evaluated at different hospitals, medical colleges, and pathology centers, and involved 1483 participants. This investigation was conducted to detect the presence of these antigens during the period of active growth of the microorganism in serum samples for pulmonary TB and processed tissue biopsy for other extrapulmonary TB. Results obtained using this test were compared with acid-fast bacilli smear and culture results. CONCLUSION Our study demonstrated that the newly developed liposome tuberculosis antigen card test detected antigens in our study population with approximately 97.48% sensitivity and 95.79% specificity. This is the first study to report the liposomal encapsulation of culture filtrate proteins from M. tuberculosis for diagnostic application.
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Affiliation(s)
- Dileep Tiwari
- Catalysis and Peptide Research Unit, School of Health Sciences, University of KwaZulu-Natal, Durban, South Africa; School of Environmental Biology and Centre for Biotechnology Studies, University of Awdhesh Pratap Singh, Rewa 486001, Madhya Pradesh, India.
| | - Shafiul Haque
- Department of Biosciences, Jamia Millia Islamia, New Delhi 110025, India; Research and Scientific Studies Unit, College of Nursing and Allied Health Sciences, Jazan University, Jazan 45142, Saudi Arabia
| | - Ram P Tiwari
- Department of Biotechnology, Immunodiagnostic Division, Vanguard Diagnostic Pvt. Ltd., Delhi 110020, India
| | - Arshad Jawed
- Research and Scientific Studies Unit, College of Nursing and Allied Health Sciences, Jazan University, Jazan 45142, Saudi Arabia
| | - Thavendran Govender
- Catalysis and Peptide Research Unit, School of Health Sciences, University of KwaZulu-Natal, Durban, South Africa
| | - Hendrik G Kruger
- Catalysis and Peptide Research Unit, School of Health Sciences, University of KwaZulu-Natal, Durban, South Africa
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Hu X, Shang M, Chen X, Xie Y, Ye Y, Zhou J, Song X, Lu X, Ying B, Wang L. Evaluation of three rapid assays for Mycobacterium tuberculosis complex detection in a comprehensive hospital from West China. Clin Biochem 2014; 48:79-84. [PMID: 25444951 DOI: 10.1016/j.clinbiochem.2014.10.003] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2014] [Revised: 09/26/2014] [Accepted: 10/01/2014] [Indexed: 02/05/2023]
Abstract
OBJECTIVES To assess the capacity of rapid and accurate confirmation of the Mycobacterium tuberculosis complex (MTBC) in a Chinese clinical laboratory. DESIGN AND METHODS This prospective study investigated three rapid assays, the Amplified Mycobacterium Tuberculosis Direct (MTD) test, real-time PCR, and acid-fast bacilli (AFB) smear, for direct detection of MTBC in a large consecutive series of different clinical specimens. Performance parameters were estimated and compared overall and for separate specimen categories using a combined reference gold standard. RESULTS The overall sensitivities were similar for MTD and real-time PCR (62.26% vs. 58.49%), significantly higher than those of AFB smear (31.13%). Among three assays, MTD had a satisfactory sensitivity in respiratory specimen (73.33%) and a nearly perfect detection for smear-positive samples (96.97%). Real-time PCR showed a high positive rate (58.97%) in regard to nonrespiratory specimen. A combination of molecular assays with conventional methods reached marked additive diagnostic values (sensitivity up to 76.42%), higher than each method individually. All detection systems showed excellent specificities (>96.00%). CONCLUSIONS The present study indicated that our lab had a moderate diagnostic performance for tuberculosis. Quality guarantee for specimen pretreatment, as well as combination analysis, will enable these assays to better incorporate into the routine laboratory workflow in China.
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Affiliation(s)
- Xuejiao Hu
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Mengqiao Shang
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Xuerong Chen
- Division of Pulmonary Disease, Department of Respiratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Yi Xie
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Yuanxin Ye
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Juan Zhou
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Xingbo Song
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Xiaojun Lu
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China
| | - Binwu Ying
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China.
| | - Lanlan Wang
- Department of Laboratory Medicine, West China Hospital, Sichuan University, Chengdu 610041, PR China.
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12
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A multicenter study of Cross-Priming Amplification for tuberculosis diagnosis at peripheral level in China. Tuberculosis (Edinb) 2014; 94:428-33. [DOI: 10.1016/j.tube.2014.04.006] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2014] [Revised: 04/18/2014] [Accepted: 04/26/2014] [Indexed: 11/19/2022]
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13
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Ou X, Li Q, Xia H, Pang Y, Wang S, Zhao B, Song Y, Zhou Y, Zheng Y, Zhang Z, Zhang Z, Li J, Dong H, Zhang J, Kam KM, Chi J, Huan S, Chin DP, Zhao Y. Diagnostic accuracy of the PURE-LAMP test for pulmonary tuberculosis at the county-level laboratory in China. PLoS One 2014; 9:e94544. [PMID: 24788724 PMCID: PMC4006777 DOI: 10.1371/journal.pone.0094544] [Citation(s) in RCA: 41] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2013] [Accepted: 03/17/2014] [Indexed: 01/31/2023] Open
Abstract
Background Early and effective detection of Mycobacterium tuberculosis (MTB), particularly in smear-negative tuberculosis (TB), is a priority for global TB control. Loop-mediated isothermal amplification with a procedure for ultra rapid DNA extraction (PURE-LAMP) can detect TB in sputum samples rapidly and with high sensitivity and specificity. However, the PURE-LAMP test has not been effectively evaluated, especially in resource-limited laboratories. In this study, we evaluated the performance of the PURE-LAMP test for TB detection in TB suspects from two county-level TB dispensaries in China. Methodology/Principal Findings From April 2011 to February 2012, patients with suspected TB were continuously enrolled from two county-level TB laboratories in China. Three sputum samples (spot, night, and morning sputum) were collected from each recruited patient. Detection of MTB by PURE-LAMP was compared to a reference standard L-J culture. The results showed that the sensitivity of the PURE-LAMP test based on spot sputum for MTB detection was 70.67%, while the sensitivity of the PURE-LAMP test based on spot sputum for MTB detection in smear positive and culture positive patients and smear negative and culture positive patients was 92.12% and 53.81%, respectively. The specificity of PURE-LAMP based on spot sputum for MTB detection was 98.32%. The sensitivity and specificity of the PURE-LAMP test based on three sputa combination for MTB detection was 88.80% and 96.86%, respectively. The results also showed that the PURE-LAMP test had a significantly lower contamination rate than did solid culture. Conclusions/Significance The study suggested that, in peripheral-level TB laboratories in China, the PURE-LAMP test showed high sensitivity and specificity for TB detection in TB suspects, making it a more effective, rapid, and safe method worthy of broader use in the future.
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Affiliation(s)
- Xichao Ou
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Qiang Li
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Hui Xia
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Yu Pang
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Shengfen Wang
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Bing Zhao
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Yuanyuan Song
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Yang Zhou
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Yang Zheng
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
| | - Zhijian Zhang
- Respiratory Diseases Department of Nanlou, Chinese People’s Liberation Army General Hospital, Beijing, P. R. China
| | | | | | | | | | - Kai Man Kam
- Stanley Ho Centre for Emerging Infectious Diseases, Faculty of Medicine, Chinese University of Hong Kong, Hong Kong, P. R. China
| | - Junying Chi
- Bill & Melinda Gates Foundation, China Office, Beijing, China
| | - Shitong Huan
- Bill & Melinda Gates Foundation, China Office, Beijing, China
| | - Daniel P. Chin
- Bill & Melinda Gates Foundation, China Office, Beijing, China
- * E-mail: (Y. Zhao); (DPC)
| | - Yanlin Zhao
- National Center for Tuberculosis Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, P. R. China
- * E-mail: (Y. Zhao); (DPC)
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The predictive value of Gen-Probe's amplified Mycobacterium tuberculosis direct test compared with culturing in paraffin-embedded lymph node tissue exhibiting granulomatous inflammation and negative acid fast stain. J Infect Public Health 2014; 7:251-6. [PMID: 24602771 DOI: 10.1016/j.jiph.2013.11.002] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2013] [Revised: 10/22/2013] [Accepted: 11/08/2013] [Indexed: 11/21/2022] Open
Abstract
BACKGROUND AND OBJECTIVES The diagnosis of granulomatous inflammation with possible tuberculosis (TB) infection in histopathology is often difficult. There is a need for a rapid and reliable diagnostic test. Thus, we evaluated the performance of the Mycobacterium tuberculosis direct (MTD) test in specimens with granulomatous lymphadenitis and negative acid fast stains. METHODS The M. tuberculosis direct (MTD) test by Gen-Probe was performed on 45 formalin-fixed paraffin-embedded tissue samples including 34 lymph nodes. We measured the predictive values of the MTD test in specimens with granulomatous lymphadenitis and negative acid fast stains. RESULTS The overall test sensitivity was 73.9%, and specificity was 95.4%. The MTD test sensitivity and specificity for lymph node tissue were 72.7% and 91.67%, respectively. In the presence of granulomatous inflammation, the MTD test sensitivity and specificity were higher than those for all tissue samples, at 75% and 100%, respectively. CONCLUSION Based on this study, the MTD test should be used as a supportive test in addition to conventional histochemical or immunological staining methods when evaluating lymph node tissue with a granulomatous inflammation to deliver stronger evidence to support clinical decisions at a much earlier time than a culture would allow.
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15
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Bensi EPA, Panunto PC, Ramos MDC. Incidence of tuberculous and non-tuberculous mycobacteria, differentiated by multiplex PCR, in clinical specimens of a large general hospital. Clinics (Sao Paulo) 2013; 68:179-84. [PMID: 23525313 PMCID: PMC3584283 DOI: 10.6061/clinics/2013(02)oa10] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/05/2012] [Accepted: 10/22/2012] [Indexed: 11/18/2022] Open
Abstract
OBJECTIVE To determine the incidence of Mycobacterium tuberculosis complex and non-tuberculous mycobacterial isolates in the routine setting of a large general hospital using an "in-house" multiplex polymerase chain reaction method and to establish a paradigm for the definitive identification of mycobacteria isolated using semi-automated equipment. METHODS Established tests, including polymerase chain reaction restriction enzyme analysis, PNB, and NAP inhibition tests as the gold standard, showed 100% agreement with an IS6110/hsp65 multiplex polymerase chain reaction when used to identify stock strains (n = 117). RESULTS In a subsequent study, 8,790 clinical specimens producing 476 isolates were evaluated with multiplex PCR and also showed 100% agreement in identification using PRA-polymerase chain reaction as the gold standard. The application of this technique to routine analysis was demonstrated in this study. A method was established with the initial application of multiplex PCR for all positive liquid cultures and the subsequent identification of non-tuberculous mycobacteria by polymerase chain reaction restriction enzyme analysis. In total, 77% of isolates belonged to the Mycobacterium tuberculosis complex, and 23% were non-tuberculous mycobacteria. CONCLUSIONS Several non-tuberculous mycobacterial species were identified, primarily M. avium, but other potentially pathogenic species were also frequently observed, including M. fortuitum, M. abscessus, and M. kansasii. The expeditious communication of these data to the clinical staff was fundamental for the diagnosis of clinical cases. Even in settings where tuberculosis is of major importance, the incidence of non-tuberculous mycobacteria infection is substantial.
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16
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Bi A, Nakajima C, Fukushima Y, Tamaru A, Sugawara I, Kimura A, Kawahara R, Hu Z, Suzuki Y. A rapid loop-mediated isothermal amplification assay targeting hspX for the detection of Mycobacterium tuberculosis complex. Jpn J Infect Dis 2012; 65:247-51. [PMID: 22627308 DOI: 10.7883/yoken.65.247] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
A rapid, simple, and low-cost diagnostic tool for tuberculosis (TB) detection is urgently needed in countries with a high TB burden. Here, we report a novel loop-mediated isothermal amplification (LAMP) assay targeting the hspX gene for the rapid detection of Mycobacterium tuberculosis, M. bovis, M. africanum, and M. microti. The specificity of this assay was evaluated using 4 reference strains of Mycobacterium tuberculosis complex (MTC), 22 species of non-tuberculous mycobacteria (NTM), 7 non-mycobacterial species, and 50 clinical M. tuberculosis isolates. All the reference MTC strains and M. tuberculosis clinical isolates were successfully detected by this method, and there were no false-positive results with NTM or non-mycobacterial species, which demonstrates the high specificity of this assay for MTC. The detection limit was 10 copies of MTC genome within 27 min, and the detection speed of this assay was higher than that of any other isothermal methods reported so far. Because of its speed, simplicity, sensitivity, specificity, and inexpensiveness, the TB hspX LAMP assay is a potential gene diagnostic method for TB detection in developing countries with a high TB burden.
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Affiliation(s)
- Aixiao Bi
- Shanghai Key Laboratory of Tuberculosis, Shanghai Pulmonary Hospital, Tongji University School of Medicine, Shanghai, China
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17
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Anochie PI, Onyeneke EC, Ogu AC, Onyeozirila AC, Aluru S, Onyejepu N, Zhang J, Efere L, Adetunji MA, Sánchez JGB. Recent advances in the diagnosis of Mycobacterium tuberculosis. Germs 2012; 2:110-20. [PMID: 24432271 PMCID: PMC3882855 DOI: 10.11599/germs.2012.1021] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2012] [Accepted: 09/01/2012] [Indexed: 02/06/2023]
Abstract
Molecular technologies offer the greatest potential for laboratories in resource-rich countries because they have the highest sensitivity and specificity. Continued use of new technologies will be crucial in elucidating the true epidemiology and pathogenesis of a disease, including the less well studied diseases. Continued development of affordable, sensitive, and specific diagnostic tools will be required for use in resource-poor settings, where the incidence of disease is highest.
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Affiliation(s)
| | | | | | | | | | - Nneka Onyejepu
- Nigerian Institute of Medical Research, Yaba, Lagos, Nigeria
| | - Jian Zhang
- Ada Technologies Inc. Denver, Colorado, USA
| | - Lauretta Efere
- Nigerian Institute of Medical Research, Yaba, Lagos, Nigeria
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18
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Palomino JC. Current developments and future perspectives for TB diagnostics. Future Microbiol 2012; 7:59-71. [PMID: 22191447 DOI: 10.2217/fmb.11.133] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022] Open
Abstract
TB persists as a global epidemic with high morbidity and mortality, especially in low-income countries. It is the only infectious disease ever declared as a global emergency by the WHO. The HIV pandemic and the emergence of drug resistance represent two additional obstacles to better control of the disease. Important progress has been made in the last decade in TB diagnostics. Major needs still exist, such as the availability of a real point-of-care test, a better diagnosis of TB in immune-compromised populations and in children, and the possibility to predict progression to disease in latently infected people. This review will summarize the current developments in TB diagnostics and the perspectives for future developments in the field.
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Affiliation(s)
- Juan Carlos Palomino
- Mycobacteriology Unit, Institute of Tropical Medicine, Nationalestraat 155, 2000, Antwerp, Belgium.
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Abubakar I, Stagg HR, Cohen T, Mangtani P, Rodrigues LC, Pimpin L, Watson JM, Squire SB, Zumla A. Controversies and unresolved issues in tuberculosis prevention and control: a low-burden-country perspective. J Infect Dis 2012; 205 Suppl 2:S293-300. [PMID: 22448025 DOI: 10.1093/infdis/jir886] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/14/2022] Open
Abstract
Despite declining incidence in most high-income countries, tuberculosis shows no signs of disappearing in the near future. Although surveillance data from most Western European countries show relatively stable declines in the rate of tuberculosis over the past several decades, some have reported either an increasing rate or a decelerating pace of reduction in recent years. The burden of disease now disproportionately affects high-risk groups such as migrants, homeless persons, and prisoners. In view of the concentration of cases in urban areas and high-risk deprived groups, interventions that may not be efficient when applied to the general population may be highly cost effective when targeted at high-risk groups. In this article, we examine some controversial elements of tuberculosis prevention and control in low-burden countries and recommend issues for further research. In particular, we assess current evidence on the duration of protection by BCG vaccine, the screening of migrants and hard-to-reach groups, and the use of preventive therapy for contacts of cases of infectious multidrug-resistant tuberculosis. This analysis is presented from the perspective of low-tuberculosis-burden, high-income countries attempting to eliminate tuberculosis.
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Affiliation(s)
- Ibrahim Abubakar
- Respiratory Diseases Department, Health Protection Services Colindale, Health Protection Agency, London, UK.
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20
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Novel real-time simultaneous amplification and testing method to accurately and rapidly detect Mycobacterium tuberculosis complex. J Clin Microbiol 2012; 50:646-50. [PMID: 22205804 DOI: 10.1128/jcm.05853-11] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
The aim of this study was to establish and evaluate a simultaneous amplification and testing method for detection of the Mycobacterium tuberculosis complex (SAT-TB assay) in clinical specimens by using isothermal RNA amplification and real-time fluorescence detection. In the SAT-TB assay, a 170-bp M. tuberculosis 16S rRNA fragment is reverse transcribed to DNA by use of Moloney murine leukemia virus (M-MLV) reverse transcriptase, using specific primers incorporating the T7 promoter sequence, and undergoes successive cycles of amplification using T7 RNA polymerase. Using a real-time PCR instrument, hybridization of an internal 6-carboxyfluorescein-4-[4-(dimethylamino)phenylazo] benzoic acid N-succinimidyl ester (FAM-DABCYL)-labeled fluorescent probe can be used to detect RNA amplification. The SAT-TB assay takes less than 1.5 h to perform, and the sensitivity of the assay for detection of M. tuberculosis H37Rv is 100 CFU/ml. The TB probe has no cross-reactivity with nontuberculous mycobacteria or other common respiratory tract pathogens. For 253 pulmonary tuberculosis (PTB) specimens and 134 non-TB specimens, the SAT-TB results correlated with 95.6% (370/387 specimens) of the Bactec MGIT 960 culture assay results. The sensitivity, specificity, and positive and negative predictive values of the SAT-TB test for the diagnosis of PTB were 67.6%, 100%, 100%, and 62.0%, respectively, compared to 61.7%, 100%, 100%, and 58.0% for Bactec MGIT 960 culture. For PTB diagnosis, the sensitivities of the SAT-TB and Bactec MGIT 960 culture methods were 97.6% and 95.9%, respectively, for smear-positive specimens and 39.2% and 30.2%, respectively, for smear-negative specimens. In conclusion, the SAT-TB assay is a novel, simple test with a high specificity which may enhance the detection rate of TB. It is therefore a promising tool for rapid diagnosis of M. tuberculosis infection in clinical microbiology laboratories.
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Hong M, Zha L, Fu W, Zou M, Li W, Xu D. A modified visual loop-mediated isothermal amplification method for diagnosis and differentiation of main pathogens from Mycobacterium tuberculosis complex. World J Microbiol Biotechnol 2011; 28:523-31. [PMID: 22806847 DOI: 10.1007/s11274-011-0843-y] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2010] [Accepted: 07/08/2011] [Indexed: 11/25/2022]
Abstract
This study was aimed to rapidly identify and differentiate two main pathogens of the Mycobacterium tuberculosis complex: Mycobacterium tuberculosis subsp. tuberculosis and Mycobacterium bovis by a modified loop-mediated isothermal amplification (LAMP) assay. The reaction results could be evaluated by naked eye with two optimized closed tube detection methods as follows: adding the modified fluorescence dye in advance into the reaction mix so as to observe the color changes or putting a tinfoil in the tube and adding the SYBR Green I dye on it, then making the dye drop into the bottom of the tube by centrifuge after reaction. The results showed that the two groups of primers used jointly in this assay could successfully identify and differentiate Mycobacterium tuberculosis subsp. tuberculosis and Mycobacterium tuberculosis bovis. Sensitivity test displayed that the modified LAMP assay with the closed tube system could determine the minimal template concentration of 1 copy/μl, which was more sensitive than that of routine PCR. The advantages of this LAMP method for detection of the Mycobacterium tuberculosis complex included high specificity, high sensitivity, simplicity, and superiority in avoidance of aerosol contamination. The modified LAMP assay would provide a potential for clinical diagnosis and therapy of tuberculosis in the developing countries and the resource-limited areas.
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Affiliation(s)
- Ming Hong
- Department of Genome Engineering, Beijing Institute of Basic Medical Sciences, Taiping Road 27, Beijing, 100850, China
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22
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Gupta BK, Bharat A, Debapriya B, Baruah H. Adenosine Deaminase Levels in CSF of Tuberculous Meningitis Patients. J Clin Med Res 2011; 2:220-4. [PMID: 21629544 PMCID: PMC3104661 DOI: 10.4021/jocmr429w] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 08/12/2010] [Indexed: 11/12/2022] Open
Abstract
Background Tuberculosis kills five lakh patients in India every year, out of which 7-12 % are with meningeal involvement. Delay in its diagnosis and in initiation of treatment results in poor prognosis and sequlae in up to 25% of cases. The aim of the present study is to look for a simple, rapid, cost effective, non-invasive and fairly specific test in differentiating tubercular etiology from other causes. Methods Forty patients between the age of 6 - 24 months attending hospital with symptoms and signs of meningitis were selected and divided into two groups: tubercular and non-tubercular, depending upon the accepted criteria. CSF was drawn and ADA estimated. Results Out of 19 tubercular patients, 18 had CSF ADA at or above the cutoff value while one had below. Out of 21 non-tuberculous patients, two had ADA levels at or above the cutoff value while 19 had below this value. Results of our study indicate that ADA level estimation in CSF is not only of considerable value in the diagnosis of TBM, CSF ADA level 10 U/L as a cutoff value exhibited 94.73% sensitivity and 90.47% specificity in differentiating tuberculous from non-tuberculous meningitis; it also has 90.00% positive predictive value and 95.00% negative predictive value. Conclusions It can be concluded that ADA estimation in CSF is not only simple, inexpensive and rapid but also fairly specific method for making a diagnosis of tuberculous etiology in TBM, especially when there is a dilemma of differentiating the tuberculous etiology from non-tuberculous ones. For this reason ADA estimation in TBM may find a place as a routine investigation. Keywords Cerebrospinal fluid; Adenosine deaminase; Tuberculous meningitis
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Affiliation(s)
- Bharat Kumar Gupta
- Department of Biochemistry, Subharti Medical College, S. V. S. University, Meerut- 250005, India
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Endobronchial ultrasound increases the diagnostic yields of polymerase chain reaction and smear for pulmonary tuberculosis. J Thorac Cardiovasc Surg 2010; 139:1554-60. [PMID: 20494195 DOI: 10.1016/j.jtcvs.2010.02.019] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/19/2009] [Revised: 11/22/2009] [Accepted: 02/09/2010] [Indexed: 11/23/2022]
Abstract
OBJECTIVES Our objective was to determine the contribution of endobronchial ultrasound in the diagnostic yields of acid-fast bacillus smear, nucleic acid amplification tests, and culture in bronchoalveolar lavage fluid for pulmonary tuberculosis. METHODS During a 1-year interval, 99 patients who had initial sputum-negative acid-fast bacillus smears or no sputum but were later proven to have a positive culture for Mycobacterium tuberculosis in their sputum or bronchoalveolar lavage fluid were retrospectively studied. Among them, 56 patients underwent bronchoscopy with endobronchial ultrasound (EBUS group) and 43 patients received conventional bronchoscopy for bronchoalveolar lavage (non-EBUS group). RESULTS The diagnostic yields of the nucleic acid amplification tests (89.3%, 50/56; P = .006), acid-fast bacillus smear (30.4%, 17/56; P = .013), and M tuberculosis culture in bronchoalveolar lavage fluid (67.9%, 38/56; P = .041) were significantly higher in the EBUS group of patients. The results of those who underwent conventional bronchoscopy were 65.1% (28/43), 9.3% (4/43), and 46.5% (20/43), respectively. Combining bronchoalveolar lavage fluid smear and nucleic acid amplification tests, we made a rapid diagnosis of pulmonary tuberculosis in 51 (91.1%) of the 56 EBUS patients and 29 (67.4%; P = .004) of the 43 non-EBUS patients. CONCLUSIONS The introduction of endobronchial ultrasound increases the diagnostic yield of the nucleic acid amplification tests, acid-fast bacillus smear, and M tuberculosis culture from bronchioalveolar lavage fluid in patients with pulmonary tuberculosis who have negative sputum smear or no sputum production.
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Palomino JC. Molecular detection, identification and drug resistance detection inMycobacterium tuberculosis: Table 1. ACTA ACUST UNITED AC 2009; 56:103-11. [DOI: 10.1111/j.1574-695x.2009.00555.x] [Citation(s) in RCA: 64] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
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Sun JR, Lee SY, Perng CL, Lu JJ. Detecting Mycobacterium tuberculosis in Bactec MGIT 960 cultures by inhouse IS6110-based PCR assay in routine clinical practice. J Formos Med Assoc 2009; 108:119-25. [PMID: 19251547 DOI: 10.1016/s0929-6646(09)60042-5] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/20/2022] Open
Abstract
BACKGROUND/PURPOSE Diagnosis of tuberculosis is challenging because the current methods are time-consuming and laborious. We have developed a method combining the Bactec MGIT 960 rapid culture system with the IS6110-based PCR for rapid diagnosis of tuberculosis. METHODS A total of 1745 samples from 712 patients treated at the Tri-Service General Hospital, Taipei, Taiwan between June and August 2005 were tested. An aliquot of positive Bactec MGIT 960 culture fluids was Kinyoun stained, and the samples positive for Kinyoun staining were directly assayed by the IS6110-based PCR. The same samples were also examined by the conventional methods for identification of Mycobacterium tuberculosis complex. RESULTS One hundred and four samples from 62 patients were positive according to the Bactec MGIT 960 system. Among these, 59 (56.7%) were positive and 45 (43.3%) were negative according to the IS6110-based PCR. Compared with the conventional identification methods, the IS6110-based PCR assay correctly identified all 59 M. tuberculosis isolates and gave negative results for all nontuberculosis mycobacteria. The mean turnaround time for M. tuberculosis identification by this combined method was 6.41 days for smear-positive and 14.33 days for smear-negative specimens. The sensitivity of the IS6110-based PCR assay was determined to be 5 cells or 0.1 pg of mycobacterial DNA. CONCLUSION The combined use of the automated Bactec MGIT 960 system and the IS6110-based PCR assay is sensitive and rapid for the detection of M. tuberculosis complex, and we recommend that this method be used routinely for identification of mycobacteria in clinical laboratories.
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Affiliation(s)
- Jun-Ren Sun
- Division of Clinical Pathology, Department of Pathology, Tri-Service General Hospital and National Defense Medical Center, Taipei, Taiwan
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Diagnostic accuracy of in-house PCR for pulmonary tuberculosis in smear-positive patients: meta-analysis and metaregression. J Clin Microbiol 2009; 47:569-76. [PMID: 19144797 DOI: 10.1128/jcm.02051-08] [Citation(s) in RCA: 40] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
In-house PCR (hPCR) could speed differential diagnosis between tuberculosis (TB) and nontuberculous mycobacterial disease in patients with positive smears and pulmonary infiltrates, but its reported accuracy fluctuates across studies. We conducted a systematic review and meta-analysis of hPCR sensitivity and specificity for smear-positive TB diagnosis, using culture as the reference standard. After searching English language studies in MEDLINE and EMBASE, we estimated cumulative accuracy by means of summary receiver operating characteristic analysis. The possible influence of hPCR procedures and study methodological features on accuracy was explored by univariate metaregression, followed by multivariate adjustment of items selected as significant. Thirty-five articles (1991 to 2006) met the inclusion criteria. The pooled estimates of the diagnostic odds ratio, sensitivity, and specificity (random-effect model) were, respectively, 60 (confidence interval [CI], 29 to 123), 0.96 (CI, 0.95 to 0.97), and 0.81 (CI, 0.78 to 0.84), but significant variations (mainly in specificity) limit their clinical applicability. The quality of the reference test, the detection method, and real-time PCR use explained some of the observed heterogeneity. Probably due to the limited study power of our meta-analysis and to the wide differences in both laboratory techniques and methodological quality, only real-time PCR also displayed a positive impact on accuracy in the multivariate model. Currently, hPCR can be confidently used to exclude TB in smear-positive patients, but its low specificity could lead to erroneous initiation of therapy, isolation, and contact investigation. As the inclusion of samples from treated patients could have artificially reduced specificity, future studies should report mycobacterial-culture results for each TB and non-TB sample analyzed.
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Cross-priming amplification for rapid detection of Mycobacterium tuberculosis in sputum specimens. J Clin Microbiol 2008; 47:845-7. [PMID: 19116359 DOI: 10.1128/jcm.01528-08] [Citation(s) in RCA: 112] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Cross-priming amplification (CPA) technology for tuberculosis diagnosis from sputum specimens was evaluated. The sensitivity of CPA from smear- and liquid culture-positive specimens was 96.9%, and that from smear-negative and liquid culture-positive specimens was 87.5%. The specificity of CPA in culture-negative specimens was 98.8%.
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Kim MH, Yang HY, Suh JT, Lee HJ. Comparison of in-house PCR with conventional techniques and Cobas Amplicor M. tuberculosis kit for detection of Mycobacterium tuberculosis. Yonsei Med J 2008; 49:537-44. [PMID: 18729295 PMCID: PMC2615280 DOI: 10.3349/ymj.2008.49.4.537] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/27/2022] Open
Abstract
PURPOSE Polymerase chain reaction (PCR) assay, introduced as a fast and sensitive diagnostic method, is useful in detecting Mycobacterium tuberculosis. The purpose of this study was to evaluate the usefulness of in-house PCR assay in the detection of Mycobacterium tuberculosis by comparing PCR results with conventional diagnostic techniques and Cobas Amplicor M. tuberculosis kit. MATERIALS AND METHODS We retrospectively assessed the diagnostic yield of in-house PCR method employed for the amplification IS6110 sequences in 2,973 specimens. We also compared in-house PCR with Cobas Amplicor M. tuberculosis kit in 120 specimens collected from June to July 2006. Routine acid-fast stain (AFS) and culture assay were also performed and analyzed. RESULTS Of 2,973 cases, 2,832 cases (95.3%) showed consistent results between in house PCR, AFS and culture methods, whereas 141 (4.7%) displayed inconsistent results. The sensitivities, specificities, and positive and negative predictive values of each method were as follows: 77.5%, 99.7%, 95.5%, and 98.0%, respectively for PCR; 49.2%, 100%, 100%, and 95.7%, respectively, for AFS method; and 80.7%, 100%, 100%, and 98.3%, respectively, for culture assay. Consistent results between PCR and Cobas Amplicor M. tuberculosis kit were shown in 109 cases (90.8%). The sensitivities, specificities, and positive and negative predictive values of each method were as follows: 81.3%, 98.9%, 96.3%, and 93.5% respectively for PCR and 71.9%, 100%, 100%, and 90.7%, respectively, for Cobas Amplicor kit. CONCLUSION In-house PCR and Cobas Amplicor kit show high sensitivity and specificity, and are reliable tests in the diagnosis of tuberculosis.
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Affiliation(s)
- Myeong-Hee Kim
- Department of Laboratory Medicine, Kyung Hee University College of Medicine, Seoul, Korea
| | - Hee-Young Yang
- Department of Laboratory Medicine, Kyung Hee University College of Medicine, Seoul, Korea
| | - Jin-Tae Suh
- Department of Laboratory Medicine, Kyung Hee University College of Medicine, Seoul, Korea
| | - Hee Joo Lee
- Department of Laboratory Medicine, Kyung Hee University College of Medicine, Seoul, Korea
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Ling DI, Flores LL, Riley LW, Pai M. Commercial nucleic-acid amplification tests for diagnosis of pulmonary tuberculosis in respiratory specimens: meta-analysis and meta-regression. PLoS One 2008; 3:e1536. [PMID: 18253484 PMCID: PMC2212137 DOI: 10.1371/journal.pone.0001536] [Citation(s) in RCA: 151] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2007] [Accepted: 01/06/2008] [Indexed: 11/22/2022] Open
Abstract
Background Hundreds of studies have evaluated the diagnostic accuracy of nucleic-acid amplification tests (NAATs) for tuberculosis (TB). Commercial tests have been shown to give more consistent results than in-house assays. Previous meta-analyses have found high specificity but low and highly variable estimates of sensitivity. However, reasons for variability in study results have not been adequately explored. We performed a meta-analysis on the accuracy of commercial NAATs to diagnose pulmonary TB and meta-regression to identify factors that are associated with higher accuracy. Methodology/Principal Findings We identified 2948 citations from searching the literature. We found 402 articles that met our eligibility criteria. In the final analysis, 125 separate studies from 105 articles that reported NAAT results from respiratory specimens were included. The pooled sensitivity was 0.85 (range 0.36–1.00) and the pooled specificity was 0.97 (range 0.54–1.00). However, both measures were significantly heterogeneous (p<.001). We performed subgroup and meta-regression analyses to identify sources of heterogeneity. Even after stratifying by type of commercial test, we could not account for the variability. In the meta-regression, the threshold effect was significant (p = .01) and the use of other respiratory specimens besides sputum was associated with higher accuracy. Conclusions/Significance The sensitivity and specificity estimates for commercial NAATs in respiratory specimens were highly variable, with sensitivity lower and more inconsistent than specificity. Thus, summary measures of diagnostic accuracy are not clinically meaningful. The use of different cut-off values and the use of specimens other than sputum could explain some of the observed heterogeneity. Based on these observations, commercial NAATs alone cannot be recommended to replace conventional tests for diagnosing pulmonary TB. Improvements in diagnostic accuracy, particularly sensitivity, need to be made in order for this expensive technology to be worthwhile and beneficial in low-resource countries.
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Affiliation(s)
- Daphne I. Ling
- Division of Epidemiology, School of Public Health, University of California, Berkeley, California, United States of America
| | - Laura L. Flores
- Division of Pulmonary and Critical Care Medicine, San Francisco General Hospital, San Francisco, California, United States of America
| | - Lee W. Riley
- Division of Epidemiology, School of Public Health, University of California, Berkeley, California, United States of America
- Division of Infectious Diseases, School of Public Health, University of California, Berkeley, California, United States of America
| | - Madhukar Pai
- Department of Epidemiology, Biostatistics and Occupational Health, McGill University, Montreal, Quebec, Canada
- * To whom correspondence should be addressed. E-mail:
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Bramante CT, Talbot EA, Rathinam SR, Stevens R, Zegans ME. Diagnosis of ocular tuberculosis: a role for new testing modalities? Int Ophthalmol Clin 2007; 47:45-62. [PMID: 17667275 DOI: 10.1097/iio.0b013e318074de79] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/16/2023]
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Tiwari RP, Hattikudur NS, Bharmal RN, Kartikeyan S, Deshmukh NM, Bisen PS. Modern approaches to a rapid diagnosis of tuberculosis: promises and challenges ahead. Tuberculosis (Edinb) 2006; 87:193-201. [PMID: 17029964 DOI: 10.1016/j.tube.2006.07.005] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2006] [Revised: 06/05/2006] [Accepted: 07/20/2006] [Indexed: 11/20/2022]
Abstract
The limitations of the conventional methods for diagnosing tuberculosis have spurred multi-faceted research activities in this field throughout the world. Chromatographic methods appear promising but may not be widely available in the developing countries. Immuno-diagnostic methods using combinations ("cocktails") of antigens have high sensitivity and specificity and can easily be applied in the peripheral laboratories and in the field settings. Though expensive, molecular methods for diagnosis of tuberculosis have advantages of speed, sensitivity, and specificity. Adequate training of the eligible personnels in molecular methods and prevention of laboratory-dependent contamination may help reduce false positive results. Although, there are no clear guidelines, so far on how to make out the best from the gene amplification methods, yet their use may be encouraged with adequate quality controls, because of the inherent ingenuity and promises of these methods. Phage-based molecular methods provide rapid results in susceptibility tests for anti-tubercular drugs. In future, many sophisticated techniques are expected to hit the market for a rapid diagnosis of tuberculosis. In the developing countries, it is necessary to evaluate availability of suitable infrastructure and trained personnels before adopting modern diagnostic methods.
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Affiliation(s)
- Ram Pramod Tiwari
- Diagnostic Division, Nicholas Piramal India Limited, Pawane, Navi Mumbai 400 705, India
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32
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Angela DP, Giuseppina C, Tony FV, Bijo B, Fatmira S, Giuseppina T. Detection of Mycobacterium tuberculosis complex in milk using polymerase chain reaction (PCR). Food Control 2006. [DOI: 10.1016/j.foodcont.2005.04.019] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2022]
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Soo PC, Horng YT, Hsueh PR, Shen BJ, Wang JY, Tu HH, Wei JR, Hsieh SC, Huang CC, Lai HC. Direct and Simultaneous Identification of Mycobacterium tuberculosis complex (MTBC) and Mycobacterium tuberculosis (MTB) by Rapid Multiplex nested PCR-ICT assay. J Microbiol Methods 2006; 66:440-8. [PMID: 16516314 DOI: 10.1016/j.mimet.2006.01.010] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2005] [Revised: 01/18/2006] [Accepted: 01/23/2006] [Indexed: 11/24/2022]
Abstract
The Mycobacterium tuberculosis (MTB) shows different virulence and host infection range from other members of the M. tuberculosis complex (MTBC). Differential identification of MTB from MTBC is thus important in certain occasions. The currently commercially available molecular assays which use either IS6110 or 16S rDNA fragment as identification targets are mainly designed for identifying MTBC but not for MTB. Comparative genomic DNA analysis has provided valuable information on regions of difference (RD) present in MTB but not in other members of the MTBC. RD9 region is further suggested to be a potential target for differential identification of MTB from MTBC. In this study, using IS6110 and Rv3618 (belong to RD9) as the specific identification targets for MTBC and MTB, respectively, we developed and tested a multiplex nested PCR-ICT (immuno-chromatography test) assay for simultaneously and directly detecting not only MTBC but also MTB from 1500 clinical sputum specimens. The results were compared with traditional culture and biochemical identification results together with patients' clinical assessments. This assay showed a 95.5% sensitivity, 97.9% specificity, 2.1% false positive rate and 4.5% false negative rate towards detection of MTBC, and a 93.0% sensitivity, 99.8% specificity, 0.2% false positive rate and 7.0% false negative rate for detection of MTB. This detection system shows great potential in clinical application.
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Affiliation(s)
- Po-Chi Soo
- Department of Clinical Laboratory Sciences and Medical Biotechnology, National Taiwan University College of Medicine, No. 1 Chan-Der Street, Taipei 100, Taiwan, ROC
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Balamurugan R, Venkataraman S, John KR, Ramakrishna BS. PCR amplification of the IS6110 insertion element of Mycobacterium tuberculosis in fecal samples from patients with intestinal tuberculosis. J Clin Microbiol 2006; 44:1884-6. [PMID: 16672431 PMCID: PMC1479206 DOI: 10.1128/jcm.44.5.1884-1886.2006] [Citation(s) in RCA: 52] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023] Open
Abstract
PCR amplification of insertion element IS6110 of Mycobacterium tuberculosis in fecal samples was evaluated in the diagnosis of intestinal tuberculosis (ITB). The numbers of samples that tested positive by PCR with SalI digestion were 16/18 untreated-ITB samples, 0/8 treated-ITB samples, 12/14 smear-positive pulmonary tuberculosis samples, and 0/30 control samples. The sensitivity, specificity, positive predictive value, and negative predictive value of fecal PCR were 88.8%, 100%, 100%, and 93.7%, respectively.
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Affiliation(s)
- Ramadass Balamurugan
- Department of Gastrointestinal Sciences, Christian Medical College, Ida Scudder Road, Vellore 632004, Tamil Nadu, India
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Abstract
PURPOSE OF REVIEW This article will review some of the recent developments for the rapid diagnosis and detection of drug resistance in tuberculosis. RECENT FINDINGS Tuberculosis remains one of the major causes of death from a single infectious agent worldwide. Of great concern for tuberculosis control is the emergence of drug resistance since there is no cure for some multidrug-resistant strains of M. tuberculosis, and there is concern that they may spread around the world, stressing the need for additional control measures such as new diagnostics and better drugs for treatment. Recent advances in molecular biology and a better understanding of the molecular basis of drug resistance have provided new tools for rapid tuberculosis diagnosis. Other non-conventional diagnostic approaches have also been proposed. Nucleic acid amplification techniques, both commercial and in-house, and non-molecular methods are being evaluated. The overall accuracy of most of these tests is promising and some of them can be easily implemented in clinical mycobacteriology laboratories. SUMMARY New genotypic and phenotypic methods for rapid diagnosis and detection of drug resistance have been developed and tested both in M. tuberculosis strains as well as in clinical samples. Further controlled evaluations are necessary in high-endemic countries for their eventual implementation in the routine diagnostic systems.
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Yeom JS, Bae WT, Park ES, Seo JH, Lim JY, Park CH, Woo HO, Youn HS. A clinical study on the etiology of parapneumonic effusion in children. KOREAN JOURNAL OF PEDIATRICS 2006. [DOI: 10.3345/kjp.2006.49.1.56] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
Affiliation(s)
- Jung-Sook Yeom
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
| | - Won-Tae Bae
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
| | - Eun-Sil Park
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
| | - Ji-Hyun Seo
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
| | - Jae-Young Lim
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
| | - Chan-Hoo Park
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
| | - Hyang-Ok Woo
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
| | - Hee-Shang Youn
- Departments of Pediatrics, Gyeongsang National University College of Medicine, Jinju, Korea
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Shamputa IC, Rigouts And L, Portaels F. Molecular genetic methods for diagnosis and antibiotic resistance detection of mycobacteria from clinical specimens. APMIS 2004; 112:728-52. [PMID: 15638836 DOI: 10.1111/j.1600-0463.2004.apm11211-1203.x] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
Mycobacteria comprise a diverse group of bacteria that are widespread in nature, some of which cause significant disease in humans. Members of the Mycobacterium tuberculosis complex (MTBC) are the most important human pathogens of the genus Mycobacterium. Traditional methods for detection and identification of mycobacteria include microscopy, culture and phenotypic tests. These methods either lack sensitivity, specificity, or are time consuming. Advances in the field of molecular biology have provided rapid diagnostic tools that have reduced the turnaround times for detecting MTBC and drug resistance in cultures and directly in clinical specimens from weeks to days. This review discusses the molecular genetic techniques for detecting and identifying MTBC as well as drug resistance of mycobacteria in clinical specimens.
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Affiliation(s)
- I C Shamputa
- Mycobacteriology Unit, Department of Microbiology, Institute of Tropical Medicine, Antwerp, Belgium
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Ogusku MM, Salem JI. Análise de diferentes primers utilizados na PCR visando ao diagnóstico da tuberculose no Estado do Amazonas. J Bras Pneumol 2004. [DOI: 10.1590/s1806-37132004000400008] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
INTRODUÇÃO: Há diferentes primers sendo testados para a detecção do DNA do Mycobacterium tuberculosis. A acuidade da reação em cadeia da polimerase (PCR) depende da existência da seqüência alvo no bacilo e de os testes serem realizados em cepas isoladas ou em amostras clínicas. OBJETIVO: Verificar a presença das seqüências de DNA alvo mais relatadas na literatura para o diagnóstico da tuberculose em amostras clínicas usando como controle positivo as respectivas cepas de M. tuberculosis isoladas. MÉTODO: Oitenta e uma amostras clínicas de pacientes com suspeita de tuberculose foram submetidas à baciloscopia e cultivo. A técnica de PCR foi realizada nas amostras clínicas e cepas isoladas com primers específicos para os seguintes alvos: IS6110, 65 kDa, 38 kDa e MPB64. RESULTADOS: Em 24 amostras com baciloscopia e cultivo negativos, a PCR também foi negativa com todos os primers testados. Em 19 amostras com baciloscopia positiva e nas cepas isoladas obteve-se 100% de resultados positivos nas PCR, exceto nas PCR em amostras clínicas com os primers para a seqüência MPB64 (89,4%). Em 38 amostras com baciloscopia negativa e cultivo positivo, as PCR tiveram resultados variáveis, sendo que os primers específicos que amplificam o fragmento de 123 pb da seqüência IS6110 foram os que forneceram os maiores percentuais de positividade (92,1%), concordância diagnóstica (0,9143), co-positividade (94,7%) e co-negatividade (100%). CONCLUSÃO: As seqüências IS6110, 38 kDa, MPB64 e 65 kDa foram encontradas no genoma de todas as cepas de M. tuberculosis isoladas desses pacientes do Estado do Amazonas. O protocolo utilizado no processamento das amostras clínicas e os primers específicos utilizados para amplificação do fragmento de 123 pb da seqüência IS6110 demonstraram maior eficiência no diagnóstico da tuberculose pulmonar (paucibacilar) em comparação com a literatura.
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Laifer G, Widmer AF, Frei R, Zimmerli W, Fluckiger U. Polymerase Chain Reaction for Mycobacterium tuberculosis. Chest 2004; 125:981-6. [PMID: 15006957 DOI: 10.1378/chest.125.3.981] [Citation(s) in RCA: 18] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/01/2022] Open
Abstract
STUDY OBJECTIVES Screening for pulmonary tuberculosis (TB) in war refugees entering low-prevalence countries for TB is a common policy, but workup strategies are difficult and expensive. DESIGN Prospective screening of war refugees for TB by chest radiograph and evaluation of the impact of additional polymerase chain reaction (PCR) testing for Mycobacterium tuberculosis complex (MTB) on clinical management in case of pulmonary infiltrates suspicious for TB. SETTING Academic university medical center. PATIENTS A total of 3,119 adult war refugees from the Kosovo war were screened by chest radiograph on arrival. Refugees with pulmonary infiltrates suspicious for TB were hospitalized, and a standardized diagnostic workup was performed. MEASUREMENTS AND RESULTS Of 3,119 adult war refugees screened for TB, 29 patients (0.9%) were identified with pulmonary infiltrates suspicious for TB; 103 specimens (76 sputa; 27 BAL fluids) were collected for acid-fast smear (AFS), PCR, and culture. The prevalence of culture-proven TB infection in this population was 27.6%. Sensitivity for PCR was higher compared with AFS for all specimens (64% vs 20%; p < 0.01) and also for each refugee with at least one positive specimen finding (100% vs 37.5%; p = 0.025). More important, the negative predictive value for three consecutive PCRs (in two sputa and one BAL) was 100%. CONCLUSIONS Repeated PCR testing for MTB in a population of asymptomatic war refugees with pulmonary infiltrates highly suggestive of TB is significantly more sensitive than AFS. Three negative PCR results allow discharge from isolation, thus reducing the economic burden of isolation strategies.
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Affiliation(s)
- Gerd Laifer
- Division of Infectious Diseases, University Hospitals Basel, Switzerland.
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Valdivieso-Garcia A, Desruisseau A, Riche E, Fukuda S, Tatsumi H. Evaluation of a 24-hour bioluminescent enzyme immunoassay for the rapid detection of Salmonella in chicken carcass rinses. J Food Prot 2003; 66:1996-2004. [PMID: 14627274 DOI: 10.4315/0362-028x-66.11.1996] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
A bioluminescent enzyme immunoassay (BEIA), using Salmonella-specific monoclonal antibody M183 for capture and biotinylated monoclonal antibody M183 for detection, was developed with InteLite AB streptavidin-biotinylated firefly luciferase complex as a reporter. Salmonella cultures were preenriched in buffered peptone water with shaking for 6 h at 37 degrees C and then selectively enriched in Muller-Kauffmann tetrathionate (MKTT) broth and modified semisolid Rappaport-Vassiliadis (MSRV) medium for 16 h at 42 degrees C. After enrichment, the total test time for the BEIA was 1.5 h. The analytical sensitivity of the BEIA ranged from 6.0 x 10(2) CFU/ml to 1.2 x 10(5) CFU/ml in MKTT and from 1.4 x 10(5) to 2.3 x 10(6) CFU/ml in MSRV using six Salmonella serovars prevalent in Canada. With enrichment cultures, the BEIA detected 1 CFU of Salmonella Typhimurium and Salmonella Enteritidis in 25 ml of chicken rinses. Representative strains of 10 Salmonella serovars were detected, and cross-reactivity was not observed with 25 non-Salmonella foodborne bacteria. The BEIA performance was assessed by testing 420 poultry samples, which were analyzed in parallel with the standard MSRV culture method. The BEIA detected 117 (27.88%) Salmonella-positive samples, whereas the standard MSRV culture method detected 124 (29.5%). The BEIA had a sensitivity of 64.5% and a specificity of 87.5% compared to the standard MSRV culture method. However, similar specificities and sensitivities were obtained when the standard MSRV culture method was compared to the BEIA (sensitivity = 68.4% and specificity = 85.5%). Neither method detected 100% of the Salmonella found in the samples tested, and statistical analyses indicated no significant difference between the two methods. In summary, the BEIA offers another alternative for the detection of Salmonella, with the additional advantage of providing a 24-h test for detecting Salmonella in chicken carcass rinses. The results obtained in this research indicate that tests are still needed for the isolation and detection of Salmonella that will establish the true prevalence of Salmonella in chicken samples.
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Affiliation(s)
- A Valdivieso-Garcia
- Laboratory for Foodborne Zoonoses, Population and Public Health Branch, Health Canada, 110 Stone Road West, Guelph, Ontario, Canada N1G 3W4.
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The Use of Molecular Methods to Determine the Cause of Mycobacterial Infections. PATHOLOGY CASE REVIEWS 2003. [DOI: 10.1097/01.pcr.0000076495.47026.5b] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
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Sarmiento OL, Weigle KA, Alexander J, Weber DJ, Miller WC. Assessment by meta-analysis of PCR for diagnosis of smear-negative pulmonary tuberculosis. J Clin Microbiol 2003; 41:3233-40. [PMID: 12843069 PMCID: PMC165327 DOI: 10.1128/jcm.41.7.3233-3240.2003] [Citation(s) in RCA: 113] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023] Open
Abstract
We conducted a meta-analysis to assess the performance of PCR for the diagnosis of smear-negative pulmonary tuberculosis (SPT) and to identify factors that account for differences in the diagnostic accuracy of different studies. Studies published before February 2002 were included if sensitivity and specificity of PCR in smear-negative respiratory or gastric-aspirate specimens could be calculated. Analysis was conducted by using summary receiver operating characteristics models. Sensitivity and specificity ranged from 9 to 100% and from 25 to 100%, respectively. Fewer than 40% of the 50 studies reported results by number of patients, reported clinical characteristics of patients, or used as a reference standard combined culture and clinical criteria. Studies that included bronchial specimens showed higher accuracy than studies that evaluated only sputum specimens or included gastric aspirates. Studies that did not report that tests were applied blindly showed higher accuracy than those reporting blind testing. Increased sensitivity due to the use of DNA purification methods was associated with decreased specificity. Studies published after 1995, using Amplicor or dUTP-UNG, were associated with an increase in specificity at the expense of lower sensitivity. We concluded that PCR is not consistently accurate enough to be routinely recommended for the diagnosis of SPT. However, PCR of bronchial specimens could be useful in highly suspicious SPT cases. Studies not reporting blind testing are likely to overestimate accuracy of PCR. Future evaluation of PCR accuracy should be conducted by patient and type of respiratory specimen, blindly, by using a reference standard that combines culture and clinical criteria and addresses the issue of how patient characteristics affect PCR accuracy.
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Affiliation(s)
- Olga L Sarmiento
- Department of Epidemiology, School of Public Health, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599, USA
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Iwamoto T, Sonobe T, Hayashi K. Loop-mediated isothermal amplification for direct detection of Mycobacterium tuberculosis complex, M. avium, and M. intracellulare in sputum samples. J Clin Microbiol 2003; 41:2616-22. [PMID: 12791888 PMCID: PMC156570 DOI: 10.1128/jcm.41.6.2616-2622.2003] [Citation(s) in RCA: 450] [Impact Index Per Article: 20.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Loop-mediated isothermal amplification (LAMP) is a novel nucleic acid amplification method in which reagents react under isothermal conditions with high specificity, efficiency, and rapidity. We used LAMP for detection of Mycobacterium tuberculosis complex, Mycobacterium avium, and Mycobacterium intracellulare directly from sputum specimens as well as for detection of culture isolates grown in a liquid medium (MGIT; Nippon Becton Dickinson Co., Ltd., Tokyo, Japan) or on a solid medium (Ogawa's medium). Species-specific primers were designed by targeting the gyrB gene, and their specificities were validated on 24 mycobacterial species and 7 nonmycobacterial species. The whole procedure is quite simple, starting with the mixing of all reagents in a single tube, followed by an isothermal reaction during which the reaction mixture is held at 63 degrees C. The resulting amplicons are visualized by adding SYBR Green I to the reaction tube. The only equipment needed for the amplification reaction is a regular laboratory water bath or heat block that furnishes a constant temperature of 63 degrees C. The assay had a detection limit of 5 to 50 copies of purified DNA with a 60-min incubation time. The reaction time could be shortened to 35 min for the species identification of M. tuberculosis complex, M. avium, and M. intracellulare from a solid-medium culture. Residual DNA lysates prepared for the Amplicor assay (Roche Diagnostics GmbH) from 66 sputum specimens were tested in the LAMP assay. Although the sample size used for the latter assay was small, 2.75 micro l of the DNA lysates, it showed a performance comparable with that of the Amplicor assay, which required 50 micro l of the lysates. This LAMP-based assay is simple, rapid, and sensitive; a result is available in 35 min for a solid-medium culture and in 60 min for a liquid-medium culture or for a sputum specimen that contains a corresponding amount of DNA available for testing.
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Affiliation(s)
- Tomotada Iwamoto
- Department of Bacteriology. Department of Parasitic Agents, Kobe Institute of Health, 4-6 Minatojima-nakamachi, Chuo-ku, Kobe 650-0046, Japan.
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Abstract
The culture of viable microorganisms from the blood or from cardiac tissue is currently the most important test for diagnosis of IE. This is followed by phenotypic identification methods used for taxonomic positioning of isolates. However, in those cases where the invading microorganism is difficult or impossible to culture (including instances of prior antimicrobial treatment), molecular methods provide the best means for detection. Molecular identification methods, either nucleic acid target or signal amplification alone or in combination with sequence analysis can offer a more specific and in some cases a more rapid alternative to the phenotypic methods. We propose revised Duke criteria of IE, including positive identification of an organism by molecular biology methods.
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Affiliation(s)
- Annette Moter
- Charité – Universitätsmedizin Berlin, Institut für Mikrobiologie und Hygiene, Dorotheenstr. 96, 10117 Berlin, Germany
| | - Michele Musci
- Deutsches Herzzentrum Berlin, Augustenburger Platz 1, 13353 Berlin, Germany
| | - Dinah Schmiedel
- Charité – Universitätsmedizin Berlin, Institut für Mikrobiologie und Hygiene, Dorotheenstr. 96, 10117 Berlin, Germany
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Tønjum T, Klintz L, Bergan T, Baann J, Furuberg G, Cristea M, Petrini B, Hoffner S. Direct detection of Mycobacterium tuberculosis in respiratory samples from patients in Scandinavia by polymerase chain reaction. Clin Microbiol Infect 2002; 2:127-131. [PMID: 11866830 DOI: 10.1111/j.1469-0691.1996.tb00218.x] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
OBJECTIVE: To investigate the use of DNA amplification by the polymerase chain reaction (PCR) for the detection of Mycobacterium tuberculosis directly in human respiratory specimens. METHODS: The PCR assay employed was the Amplicor M. tuberculosis Test (Roche Diagnostics, Switzerland), which uses the 16S rDNA as the target template. Nine hundred and sixty samples from 741 patients in two clinical microbiology laboratories in Norway and Sweden were processed by routine culture analysis and PCR. RESULTS: Of the 56 specimens containing cultivatable M. tuberculosis, 49 (87.5%) were detected by PCR. Among the 904 culture-negative specimens, 897 samples were negative also by PCR and seven (0.8%) were positive by PCR. In comparison with culture, the sensitivity, specificity, and positive and negative predictive values of PCR were 91.7%, 99.6%, 94.2% and 99.4% for laboratory 1 and 80.0%, 98.7%, 76.2% and 99.0% for laboratory 2, respectively. For both laboratories combined the values were 87.5%, 99.2%, 87.5% and 99.2%. CONCLUSIONS: These results indicate that multiple (two or three) respiratory samples from each patients should be tested, to allow sufficient accuracy in detecting M. tuberculosis in the specimens. Still, the labor-intensive format of this test necessitates strong clinical indications and patient prioritization to provide a service feasible within the current limits of routine laboratories.
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Affiliation(s)
- Tone Tønjum
- Institute of Microbiology, University of Oslo, Rikshospitalet (National Hospital), Oslo, Norway
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Yeboah-Manu D, Yates MD, Wilson SM. Application of a simple multiplex PCR to aid in routine work of the mycobacterium reference laboratory. J Clin Microbiol 2001; 39:4166-8. [PMID: 11682550 PMCID: PMC88507 DOI: 10.1128/jcm.39.11.4166-4168.2001] [Citation(s) in RCA: 44] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
A PCR specific for spacer regions 33 and 34 of the direct repeat region of the Mycobacterium tuberculosis complex was developed to complement the biochemical differentiation of M. tuberculosis, Mycobacterium bovis, M. bovis BCG, and Mycobacterium africanum subtypes I and II. In addition, this approach was incorporated into a multiplex PCR that included primers specific for IS6110 and the 65-kDa antigen gene in order to differentiate members of the M. tuberculosis complex from atypical mycobacteria.
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Affiliation(s)
- D Yeboah-Manu
- Noguchi Memorial Institute for Medical Research, University of Ghana, Legon, Ghana
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Abstract
Abstract
Background: Nucleic acid amplification technologies such as PCR are revolutionizing the detection of infectious pathogens such as tuberculosis (TB). Amplification technology offers the potential for the diagnosis of TB in a few hours with a high degree of sensitivity and specificity. However, molecular assays neither replace nor reduce the need for conventional smear and culture, speciation, and antibiotic sensitivity assays.
Methods: We undertook prospective studies of sputum samples to assess the performance of two PCR-based assays for the detection of TB as well as the impact of more rapid availability of test results on patient care.
Results: The sensitivity of both the in-house and Amplicor PCR assays was 100% for smear-positive sputa. For smear-negative sputa (two sputum samples collected during the first 24 h of hospitalization), the sensitivity was 85% for our in-house PCR assay and 74% for the Roche PCR assay. Approximately 10% of the smear- and culture-negative sputa yielded positive PCR results; however, more than one-half of these were positive with both the in-house and Amplicor assays, suggesting the presence of TB DNA or organisms. Several of these came from patients whose other samples grew Mycobacterium tuberculosis during the same admission, and others came from patients who had previously treated TB. Overall, the specificities of the in-house and Amplicor PCR assays in smear-negative patients were 86% and 93%, respectively.
Conclusions: Molecular detection of slow-growing pathogens such as M. tuberculosis have the potential to improve clinical care through a dramatic reduction in the time required for detection and may provide substantial savings in the overall cost of care of a patient compared with conventional smear, culture, and speciation alone, despite the fact that conventional assays must still be performed for speciation of nontuberculous mycobacteria and for full assessment of antibiotic sensitivity.
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Affiliation(s)
- Karen L Kaul
- Department of Pathology, Evanston Northwestern Healthcare, 2650 Ridge Ave., Evanston, IL 60201. Fax 847-570-1938; e-mail
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Sakallah SA. Molecular diagnostics of infectious diseases: state of the technology. BIOTECHNOLOGY ANNUAL REVIEW 2001; 6:141-61. [PMID: 11193293 DOI: 10.1016/s1387-2656(00)06021-x] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
In this review, the basic technologies and procedures currently used in clinical laboratories performing molecular diagnostics are described. Special emphasis on specimen processing has been made since it is one of the most challenging steps involved in molecular testing. Representative examples are given for each type of technology, especially tests that are currently available in the market. The types of hybridization-based and amplification-based procedures are detailed. Finally, current problems and future developments are discussed.
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Affiliation(s)
- S A Sakallah
- Department of Health and Human Services, Public Health Laboratories, State of New Hampshire, 6 Hazen Drive, Concord, NH 03301, USA.
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Abstract
OBJECTIVE To assess the clinical utility of the commercial nucleic acid amplification (NAA) tests (ie, Amplified Mycobacterium Tuberculosis Direct Test, Gen-Probe, Inc and AMPLICOR Mycobacterium tuberculosis Test, Roche Molecular Systems, Inc) for direct detection of Mycobacterium tuberculosis complex. DATA SOURCES Review of the English-language literature. CONCLUSIONS The performance of both NAA tests is excellent (sensitivity, > or = 95%; specificity, 100%) when testing respiratory specimens that are smear-positive for acid-fast bacilli (AFB). Only the Gen-Probe assay is approved for testing respiratory specimens regardless of the AFB smear result. Data from 3 studies showed that the sensitivity of the Mycobacterium Tuberculosis Direct Test in smear-negative patients ranged from 83% to 85%, and that the specificity was 99%. Both NAA tests have been used to test nonrespiratory specimens; in some studies, the performance was comparable to the performance obtained for respiratory specimens, whereas in others, it was lower. The NAA tests also appear to be reliable tools for rapid detection of M tuberculosis complex in positive broth cultures of all specimen types (except blood). The impact of the NAA tests on patient outcome varies based on the result of the AFB smear. In smear-positive patients, public health and hospital infection-control resources are predominantly affected. The potential for influencing patient outcome is much greater when the AFB smear is negative. In smear-negative patients, the NAA test could provide more rapid diagnosis of tuberculosis and subsequent initiation of therapy; eliminate the need for invasive diagnostic procedures, which are both costly and pose an added risk to the patient; and allow earlier discharge of hospitalized patients. Prospective studies concerning the cost-effectiveness of the NAA tests are needed.
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Affiliation(s)
- G L Woods
- Department of Pathology, University of Texas Medical Branch-Galveston, Galveston, Tex 77555-0740, USA.
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Al Zahrani K, Al Jahdali H, Poirier L, René P, Gennaro ML, Menzies D. Accuracy and utility of commercially available amplification and serologic tests for the diagnosis of minimal pulmonary tuberculosis. Am J Respir Crit Care Med 2000; 162:1323-9. [PMID: 11029339 DOI: 10.1164/ajrccm.162.4.9912115] [Citation(s) in RCA: 52] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
Diagnosis of patients with minimal active tuberculosis (TB) is difficult, as there is no single test with high sensitivity and specificity. The yield and clinical utility of a combination of diagnostic tests were prospectively studied among 500 consecutive patients referred for sputum induction for diagnosis of possible active TB. Patients underwent sputum induction, chest X-ray, tuberculin testing, and had blood drawn for serologic testing (Detect-TB test; Biochem ImmunoSystems). Sputum was examined with fluorescent microscopy and PCR (Amplicor MTB-Roche) and cultured for mycobacteria using liquid (BACTEC) and solid media. For the diagnosis of the 60 cases of active TB, sensitivity and specificity, respectively, of the following diagnostic tests were mycobacterial culture, 73% and 100%; PCR, 42% and 100%; chest X-ray, 67-77% and 66-76%; tuberculin testing, 94% and 20%; and serology, 33% and 87%. After consideration of PCR and radiographic and clinical characteristics, a positive serologic test was independantly associated with diagnosis of active disease (adjusted odds of disease if positive, 2.6; 95% confidence limits, 1.1,6.1). No currently available test has sensitivity and specificity high enough for the accurate diagnosis of minimal pulmonary TB. Utilization of a combination of tests, together with consideration of key clinical characteristics, could improve diagnostic accuracy.
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Affiliation(s)
- K Al Zahrani
- Montreal Chest Institute, Respiratory Epidemiology Unit, Department of Microbiology of the Royal Victoria Hospital, McGill University; Hopital Maisonneuve Rosemont, University of Montreal, Montreal, Quebec, Canada
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