scholarly journals Molecular Detection of Mycobacterium tuberculosis in Oral Mucosa from Patients with Presumptive Tuberculosis

2020 ◽  
Vol 9 (12) ◽  
pp. 4124
Author(s):  
Barbara Molina-Moya ◽  
Nelly Ciobanu ◽  
Marta Hernandez ◽  
Cristina Prat-Aymerich ◽  
Valeriu Crudu ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Oral Mucosa ◽  
The Elderly ◽  
Storage Conditions ◽  
Molecular Diagnostic ◽  
Smear Microscopy ◽  
X Rays ◽  
Rt Pcr ◽  
Tuberculosis Complex ◽  
Tb 34

Tuberculosis (TB) diagnosis is increasingly based on the detection of Mycobacterium tuberculosis complex (MTBC) DNA in sputum using molecular diagnostic tests as the first test for diagnosis. However, sputum can be difficult to obtain in children, patients without productive cough, and the elderly and approaches testing non-sputum samples are needed. We evaluated whether TB can be detected from the oral mucosa of patients with TB. Adults with presumptive TB were examined using culture, Xpert MTB/RIF, smear microscopy and X-Rays. Oral mucosa swabs collected on PrimeStore-MTM, stored at room temperature if tested within 30 days or at −20 °C if examined at a later time. RT-PCR was performed to detect M. tuberculosis DNA. Eighty patients had bacteriologically-confirmed TB, 34 had bacteriologically-negative TB (negative tests but abnormal X-rays) and 152 were considered not to have TB (not TB). Oral swabs RT-PCR were positive in 29/80 (36.3%) bacteriologically-confirmed, 9/34 (26.5%) bacteriologically-negative and 29/152 (19.1%) not TB. The yield varied among samples stored for less and more than 30 days (p = 0.013) from 61% (11/18) and 29% (18/62) among bacteriologically confirmed, and 30.8% (4/13) and 23.8% (5/21) among bacteriologically-negative participants. Among not TB patients, the specificity was 80.9% (123/152), being 78.3% (18/23) among samples stored less than 30 days and 81.4% (105/129) among samples stored for more than 30 days (p = 0.46). The detection of M. tuberculosis in oral mucosa samples is feasible, but storage conditions may affect the yield.

scholarly journals Role of GenoType® Mycobacterium Common Mycobacteria/Additional Species Assay for Rapid Differentiation between Mycobacterium tuberculosis Complex and Different Species of Non-tuberculous Mycobacteria

2013 ◽  
Vol 5 (02) ◽  
pp. 083-089 ◽  
Author(s):  
Amresh Kumar Singh ◽  
Anand Kumar Maurya ◽  
Jyoti Umrao ◽  
Surya Kant ◽  
Ram Awadh Singh Kushwaha ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Mycobacterium Tuberculosis Complex ◽  
Clinical Isolates ◽  
Molecular Diagnostic ◽  
Mycobacterium Fortuitum ◽  
Tuberculosis Complex ◽  
Accurate Identification ◽  
Additional Species ◽  
Culture Positive

ABSTRACT Background: Mycobacterium tuberculosis complex (MTBC) and non-tuberculous mycobacteria (NTM) may or may not have same clinical presentations, but the treatment regimens are always different. Laboratory differentiation between MTBC and NTM by routine methods are time consuming and cumbersome to perform. We have evaluated the role of GenoType® Mycobacterium common mycobacteria/additional species (CM/AS) assay for differentiation between MTBC and different species of NTM in clinical isolates from tuberculosis (TB) cases. Materials and Methods: A total of 1080 clinical specimens were collected from January 2010 to June 2012. Diagnosis was performed by Ziehl-Neelsen staining followed by culture in BacT/ALERT 3D system (bioMerieux, France). A total of 219 culture positive clinical isolates (BacT/ALERT® MP cultures) were selected for differentiation by p-nitrobenzoic acid (PNB) sensitivity test as and BIO-LINE SD Ag MPT64 TB test considering as the gold standard test. Final identification and differentiation between MTBC and different species of NTM were further confirmed by GenoType® Mycobacterium CM/AS assay (Hain Lifescience, Nehren, Germany). Results: Out of 219 BacT/ALERT® MP culture positive isolates tested by PNB as 153 MTBC (69.9%) and by GenoType® Mycobacterium CM/AS assay as 159 (72.6%) MTBC and remaining 60 (27.4%) were considered as NTM species. The GenoType® Mycobacterium CM/AS assay was proved 99.3% sensitive and 98.3% specific for rapid differentiation of MTBC and NTM. The most common NTM species were; Mycobacterium fortuitum 20 (33.3%) among rapid growing mycobacteria and Mycobacterium intracellulare 11 (18.3%) among slow growing mycobacteria. Conclusion: The GenoType® Mycobacterium assay makes rapid and accurate identification of NTM species as compared with different phenotypic and molecular diagnostic tool and helps in management of infections caused by different mycobacteria.

scholarly journals Paradoxical conservation of a set of three cellulose-targeting genes in Mycobacterium tuberculosis complex organisms

Microbiology ◽  
2010 ◽  
Vol 156 (5) ◽  
pp. 1468-1475 ◽  
Author(s):  
Felix Mba Medie ◽  
Iskandar Ben Salah ◽  
Michel Drancourt ◽  
Bernard Henrissat
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Cell Walls ◽  
Pcr Amplification ◽  
Plant Cell Walls ◽  
Rt Pcr ◽  
Bacterial Genomes ◽  
Tuberculosis Complex ◽  
Transcription Analysis ◽  
Cellulose Binding

The genome of the tuberculosis agent Mycobacterium tuberculosis encodes a putative cellulose-binding protein (CBD2), one candidate cellulase (Cel12), and one fully active cellulase (Cel6). This observation is puzzling, because cellulose is a major component of plant cell walls, whereas M. tuberculosis is a human pathogen without known contact with plants. In order to investigate the biological role of such cellulose-targeting genes in M. tuberculosis we report here the search for and transcription analysis of this set of genes in the genus Mycobacterium. An in silico search for cellulose-targeting orthologues found that only 2.5 % of the sequenced bacterial genomes encode the Cel6, Cel12 and CBD2 gene set simultaneously, including those of the M. tuberculosis complex (MTC) members. PCR amplification and sequencing further demonstrated the presence of these three genes in five non-sequenced MTC bacteria. Among mycobacteria, the combination of Cel6, Cel12 and CBD2 was unique to MTC members, with the exception of Mycobacterium bovis BCG Pasteur, which lacked CBD2. RT-PCR in M. tuberculosis H37Rv indicated that the three cellulose-targeting genes were transcribed into mRNA. The present work shows that MTC organisms are the sole mycobacteria among very few organisms to encode the three cellulose-targeting genes CBD2, Cel6 and Cel12. Our data point toward a unique, yet unknown, relationship with non-plant cellulose-producing hosts such as amoebae.

scholarly journals A study on mutation in genes associated with rifampicin and isoniazid resistance in Mycobacterium tuberculosis complex using line probe assay

2018 ◽  
Vol 6 (12) ◽  
pp. 3997
Author(s):  
Rohit Kumar ◽  
Shivendra Kumar Shahi ◽  
Rakesh Kumar ◽  
Balkrishna Mishra ◽  
Shailesh Kumar ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Early Diagnosis ◽  
Pulmonary Tuberculosis ◽  
Mycobacterium Tuberculosis Complex ◽  
Smear Microscopy ◽  
Common Mutation ◽  
Line Probe Assay ◽  
Tuberculosis Complex ◽  
Sensitivity Pattern ◽  
Probe Assay

Background: The term tuberculosis describe a clinical illness, which is predominantly caused by Mycobacterium tuberculosis and less common by other species. Infection is transmitted by infected droplets through respiratory route. Early diagnosis and appropriate management is the only way to control the spread of infection. The available diagnostic tools include, smear microscopy, culture and molecular methods. Culture is the gold standard, but it takes around 2-8 weeks to get the result and smear microscopy having less sensitivity. Molecular technique especially Line probe assay can be better option because of high sensitivity and specificity, and directly clinical sample can be used, and result will be made available within same day with sensitivity pattern. Present study was designed to use of LPA for early diagnosis.Methods: Laboratory based observational study conducted in department of microbiology, IGIMS Patna and TBDC, Patna. Sputum specimens were collected from clinically suspected cases of pulmonary tuberculosis, and subjected to smear microscopy, culture and LPA.Results: During the study period, 2841 patients were diagnosed as pulmonary tuberculosis. Strain of Mycobacterium tuberculosis complex in, 12% (347) patients were rifampicin and isoniazid resistant, 4% (117) and 3% (86) patients were rifampicin and isoniazid mono-resistant respectively. We found that rpoB MUT3 was the most common mutation in gene associated with rifampicin resistant and katG MUT1gene associated with isoniazid resistant.Conclusions: Present study support the use of LPA for early diagnosis of smear positive as well as smear negative pulmonary tuberculosis cases. Resulting early diagnosis and appropriate management of patients.

scholarly journals Identification and Discrimination of Mycobacterium tuberculosis Complex with Traditional and Real-Time PCR in Different Specimens in Iraq

2020 ◽  
Vol 62 (3) ◽  
Author(s):  
Shaymaa Mahmood Ali ◽  
Mohammad A. Al-Faham ◽  
Ahmed A. Mankhi
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Real Time ◽  
Real Time Pcr ◽  
Mycobacterium Tuberculosis Complex ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Tuberculosis Complex ◽  
Direct Smear ◽  
Smear Examination ◽  
Lowenstein Jensen

Background: Tuberculosis (TB) is a major public health issue and a main cause of global morbidity and mortality. TB is the world's ninth leading cause of death despite the numerous treatment strategies for managing the disease.Objective: To assess the traditional method (direct smear examination and culture) against real-time PCR as pulmonary and extrapulmonary tuberculosis laboratory diagnostic techniques.Cases and methods: Samples were collected from (612) TB cases, (409) of whom were pulmonary tuberculosis (PTB) and (203) were extrapolmonary tuberculosis (EPTB). The cases were seeking care at the Specialized Chest and Respiratory Disease Center/ National Reference Laboratory for Tuberculosis (NRL) in Baghdad, during the period 1st of May -1st of October 2019. Direct smear examination, Lowenstein-Jensen culture and Real Time PCR were used to diagnose TB.Results: Out of 612 samples received, 82(13.4%) were positive by smear microscopy, while 90(14.7%) were able to grow on Lowenstein-Jensen (LJ) media. Ninety DNA extracts from the samples which were positive on LJ media and 25 control specimens, were diagnosed with molecular analysis by using real time PCR to determine the species of Mycobacterium tuberculosis complex. The results revealed that the 71 samples (78.9%) were M. tuberculosis, three specimens (3.3%) were combined M. bovis and M. tuberculosis, and one M. tuberculosis, M. bovis, and M. bovis BCG, while15 (16.7%) were negative and subsequently excluded from study.Conclusion: The comparison between molecular diagnostic methods by using Real time PCR with conventional diagnostic methods, provides a new promising technique and is potentially a practical and rapid alternative to the slower traditional pulmonary and EPTB diagnostic culture. The results show M. bovis overall contribution on human TB in comparison to M. tuberculosis is minor among PTB and EPTB cases in the sample studied.

scholarly journals Evaluation of Three Nucleic Acid Amplification Methods for Direct Detection of Mycobacterium tuberculosis Complex in Respiratory Specimens

1999 ◽  
Vol 37 (6) ◽  
pp. 1932-1934 ◽  
Author(s):  
S. X. Wang ◽  
L. Tay
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Nucleic Acid ◽  
Mycobacterium Tuberculosis Complex ◽  
Direct Detection ◽  
Nucleic Acid Amplification ◽  
Smear Microscopy ◽  
Direct Test ◽  
Tuberculosis Complex ◽  
Chain Reaction ◽  
Respiratory Specimens

Two hundred thirty respiratory specimens from 230 patients were analyzed by using COBAS AMPLICOR PCR, Amplified Mycobacterium tuberculosis Direct Test, and ligase chain reaction methods. Results were compared with those of smear microscopy and radiometric culture (Bactec) methods. No significant differences were observed among the results of the three methods, which are acceptable for direct detection of M. tuberculosis complex in respiratory specimens.

scholarly journals Detection of Mycobacterium tuberculosis Complex Bacilli and Nucleic Acids From Tongue Swabs in Young, Hospitalized Children

2021 ◽  
Vol 11 ◽  
Author(s):  
Christopher Ealand ◽  
Julian Peters ◽  
Olivia Jacobs ◽  
Astika Sewcharran ◽  
Azra Ghoor ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Genomic Dna ◽  
Genetic Material ◽  
Smear Microscopy ◽  
Hospitalized Children ◽  
Tuberculosis Complex ◽  
Tuberculosis In Children ◽  
Positive Results ◽  
Culture Negative ◽  
Oral Epithelia

Diagnosis of tuberculosis in pediatric patients remains challenging due to inherent difficulties associated with obtaining respiratory samples for molecular and culture-based testing. To address this, recent studies have highlighted the utility of tongue swabs to detect Mycobacterium tuberculosis genomic DNA in the oral epithelia of tuberculosis infected adults. It is unknown whether tongue swabs have similar utility for diagnosis of childhood tuberculosis and if the presence of DNA in these swabs was associated with whole bacilli. We therefore sought to conduct a preliminary assessment of the utility of tongue swabs to detect tubercle bacilli and their associated genetic material in young children. For this, we recruited hospitalized children with clinically diagnosed tuberculosis (n = 26) or lower respiratory tract infection (LRTI, n = 9). These categories were blinded for downstream laboratory tests, which included PCR, spoligotyping, smear microscopy, and culture. Mtb genomic DNA was detected by PCR only in clinically diagnosed TB cases [11/26 (31.4%)] and not in cases with LRTI. Of these, 5/11 [45.5%] were associated with a spoligotype. Spoligotyping also detected an additional six specimens that were negative by PCR. Using smear microscopy, 19/26 [73.1%] and 4/9 [44.4] were Mtb positive in the tuberculosis or LRTI categories respectively. We noted positive results on all three tests in 5/26 [19.2%] in the tuberculosis category and 0/9 in the LRTI category. All specimens were culture negative. Collectively, these preliminary data present a compelling case for broader testing of tongue swabs to diagnose tuberculosis in children where obtaining standard sputum specimens is not easy.

1376: Molecular Diagnostic Detection of Haploid Germ Cells in Testicular Biopsy Specimens by Online RT-PCR

2004 ◽  
Vol 171 (4S) ◽  
pp. 362-363
Author(s):  
Mark G. Schrader ◽  
Markus Muller ◽  
Wolfgang Schulze ◽  
Steffen Weikert ◽  
Kurt Miller
Keyword(s):  
Germ Cells ◽  
Testicular Biopsy ◽  
Molecular Diagnostic ◽  
Rt Pcr ◽  
Biopsy Specimens ◽  
Diagnostic Detection

DETECTING MYCOBACTERIUM TUBERCULOSIS RAPIDLY AND MYCOBACTERIUM TUBERCULOSIS STRAINS WITHOUT IS6110 SEQUENCE BY PCR ASSAY

2012 ◽  
pp. 15-19
Author(s):  
Thi Chau Anh Nguyen ◽  
Hoang Bach Nguyen ◽  
Hai Duong Huynh ◽  
Nu Xuan Thanh Le ◽  
Xuan Cuong Le ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
16S Rdna ◽  
False Negative ◽  
Clinical Samples ◽  
Tuberculosis Complex ◽  
Pcr Assay ◽  
Complex Objects ◽  
Method Performance ◽  
Gene Coding ◽  
16S Rdna Pcr

Background: The Nested IS6110 PCR is used for detecting tuberculosis, however IS6110 sequence is not present in the genome of all strains of M.tuberculosis, the result may be false negative. The gene coding 16S ribosome always contains a short sequence specific to M. tuberculosis complex. Objects: Performance of the 16S Real-time PCR to detect M. tuberculosis and combining to the nested IS6110 PCR to determine the rate of Mtb strains without IS6110 from clinical samples. Materials and method: Performance of 16S rDNA PCR by commercial kit of Viet A Inc. for all 480 samples, the samples which were positive with the 16S rDNA PCR were retested in IS6110 PCR assay by in-house kit. Results: The Realtime 16S rDNA PCR detected 258 cases (53.8%) of tuberculosis. There were 3 (1.2 %) M. tuberculosis strains which do not harbor IS6110 sequence in genome. Conclusion: The IS6110 nested PCR can be applied more widely than the 16S rDNA realtime PCR. In case of using IS6110 PCR assay, results may show a low proportion of false negative. Combining 16S rDNA PCR with the IS6110 based PCR allowed detection of deletion of IS6110 sequence in M. tuberculosis isolates.

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