Development and evaluation of multiplex PCR in rapid diagnosis of abdominal tuberculosis

Hallur, Vinaykumar; Sharma, Mukut; Sethi, Sunil; Sharma, Kusum; Mewara, Abhishek; Dhatwalia, Sunil Kumar; Yadav, Rakesh; Bhasin, Deepak K.; Sinha, Saroj Kant; Rana, Satya Vati; Joshi, Kusum

doi:10.1016/j.diagmicrobio.2013.02.022

Cited by 41 publications

(35 citation statements)

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“…Previously reported PCR methods for genus detection seem to have different drawbacks apparently avoided by this new PCR. As alignment between published primers/probes and genomic targets used by other researchers (16S rRNA, ITS, and heat shock protein 65) suggested, some might not detect DNA from various mycobacteria (37,38), others would detect nonmycobacterial DNA (20,21,23,25,32), and others might combine both sensitivity and specificity limitations (22,24,26,39). For M. avium subspecies detection, we used the multicopy element IS1311, a target previously used in multiplex real-time PCR assays (25) that, as far as we know, is M. avium subspecies specific (5).…”

Section: Discussionmentioning

confidence: 99%

“…Multiplexing strategies allow for further improvement of diagnostics in terms of rapidity and efficiency, and multiplex real-time PCR represents a reliable alternative. Among the various multiplex PCR assays reported thus far (20)(21)(22)(23)(24)(25)(26), many are conventional PCRs or use melting curve analysis and need additional interpretation; some have not been used directly on clinical samples, and others seem to have some specificity or sensitivity issues. A recently published duplex real-time PCR assay for the detection of M. tuberculosis and M. avium complexes on human respiratory specimens has been shown to be reliable and cost-effective (27), but it could have an added value by including an additional target for genus detection.…”

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confidence: 99%

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Detection of Mycobacteria, Mycobacterium avium Subspecies, and Mycobacterium tuberculosis Complex by a Novel Tetraplex Real-Time PCR Assay

et al. 2015

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bMycobacterium tuberculosis complex, Mycobacterium avium, and many other nontuberculous mycobacteria are worldwide distributed microorganisms of major medical and veterinary importance. Considering the growing epidemiologic significance of wildlife-livestock-human interrelation, developing rapid detection tools of high specificity and sensitivity is vital to assess their presence and accelerate the process of diagnosing mycobacteriosis. Here we describe the development and evaluation of a novel tetraplex real-time PCR for simultaneous detection of Mycobacterium genus, M. avium subspecies, and M. tuberculosis complex in an internally monitored single assay. The method was evaluated using DNA from mycobacterial (n ‫؍‬ 38) and nonmycobacterial (n ‫؍‬ 28) strains, tissues spiked with different CFU amounts of three mycobacterial species (n ‫؍‬ 57), archival clinical samples (n ‫؍‬ 233), and strains isolated from various hosts (n ‫؍‬ 147). The minimum detectable DNA amount per reaction was 50 fg for M. bovis BCG and M. kansasii and 5 fg for M. avium subsp. hominissuis. When spiked samples were analyzed, the method consistently detected as few as 100 to 1,000 mycobacterial CFU per gram. The sensitivity and specificity values for the panel of clinical samples were 97.5 and 100% using a verified culture-based method as the reference method. The assays performed on clinical isolates confirmed these results. This PCR was able to identify M. avium and M. tuberculosis complex in the same sample in one reaction. In conclusion, the tetraplex real-time PCR we designed represents a highly specific and sensitive tool for the detection and identification of mycobacteria in routine laboratory diagnosis with potential additional uses. T he genusMycobacterium encompasses a large number of worldwide distributed pathogenic and opportunistic bacteria responsible for a broad range of infectious diseases. Mycobacterium tuberculosis complex, Mycobacterium avium, and many other nontuberculous mycobacteria (NTM) are of major medical and veterinary significance. Human tuberculosis (TB) due to infection with members of M. tuberculosis complex, especially with M. tuberculosis, remains an important health problem according to annual reports of the World Health Organization (WHO). Animal or bovine TB (bTB), caused primarily by Mycobacterium bovis but also by Mycobacterium caprae, poses serious socioeconomic and health threats (1) and is a notifiable disease listed under the World Organization for Animal Health (OIE) Terrestrial Animal Health Code (TAHC). Although bTB is controlled in most developed countries, its eradication has not been fully accomplished due to a lack of sensitivity of diagnostic tools and to the persistence of infection in extensive farming and wild populations (2). The disease remains a significant problem in many countries with less developed laboratory capacity (1). Accordingly, human TB caused by M. bovis or M. caprae is supposed to be infrequent in developed regions but is relatively common and possibly underestimated in ...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Detection of Mycobacteria, Mycobacterium avium Subspecies, and Mycobacterium tuberculosis Complex by a Novel Tetraplex Real-Time PCR Assay

et al. 2015

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“…Similarly, in the present study there was only one positive result out of seven (28.5%) patients given the tuberculin skin test. Common diagnostic methods for microbiological confirmation of abdominal TB were reported to have very low sensitivity (3,6,18,29). In this study, positive results were obtained only in two patients with bacterial sputum culture, and there were only one positive growth on culture media.…”

Section: Discussionmentioning

confidence: 73%

“…Diagnosis of abdominal TB in children is challenging due to nonspecific clinical features depending on host factors such as age and immunological status; and in most cases, pulmonary TB is absent making it difficult to establish the diagnosis (6,(14)(15)(16)(17)(18). Less than half of the patients can be diagnosed with abdominal TB when the only clinical presentation is abdominal complaint such as pain (3).…”

Section: Introductionmentioning

confidence: 99%

“…There are several reasons for high morbidity rate of this preventable disease such as poor treatment adherence, acquired drug resistance, treatment failure, and development of Mycobacterium tuberculosis infection without any clinical symptoms, which increases the risk of progression and extrapulmanory manifestations (2)(3)(4)(5). Abdominal TB is the sixth commonest extrapulmanory TB after lymphatic, genitourinary, bone and joint, miliary, and meningeal forms, constituting 5% of all TB cases (6,7). It occurs via the infection of gastrointestinal tract through hematogenous spread from a primary lung focus, or via lymphatics from infected nodes, ingestion of bacilli either from the sputum or from contaminated milk products, or by direct spread from adjacent organs (3,8,9).…”

Section: Introductionmentioning

confidence: 99%

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Clinical Presentation in a Series of Eight Children with Abdominal Tuberculosis: Experience of a Single-Center in Turkey

et al. 2017

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Background: Abdominal tuberculosis, the sixth commonest extrapulmanory tuberculosis, is easily misdiagnosed due to nonspecific gastrointestinal symptoms. The diagnosis is highly dependent on clinician's suspicion of the disease; therefore to increase awareness we retrospectively reviewed clinical features of eight children in a four year-period. Methods: The medical records of children with abdominal TB were retrospectively reviewed. The diagnosis of abdominal TB was defined as Mycobacterium tuberculosis infection of gastrointestinal tract along with peritoneal or solid organ involvement.

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Screening and identification of serum biomarkers of osteoarticular tuberculosis based on mass spectrometry

Chen

Jia

Lei

et al. 2020

Clinical Laboratory Analysis

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Development and evaluation of multiplex PCR in rapid diagnosis of abdominal tuberculosis

Cited by 41 publications

References 16 publications

Detection of Mycobacteria, Mycobacterium avium Subspecies, and Mycobacterium tuberculosis Complex by a Novel Tetraplex Real-Time PCR Assay

Detection of Mycobacteria, Mycobacterium avium Subspecies, and Mycobacterium tuberculosis Complex by a Novel Tetraplex Real-Time PCR Assay

Clinical Presentation in a Series of Eight Children with Abdominal Tuberculosis: Experience of a Single-Center in Turkey

Screening and identification of serum biomarkers of osteoarticular tuberculosis based on mass spectrometry

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