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Rapid diagnosis and simultaneous identification of tuberculous and bacterial meningitis by a newly developed duplex polymerase chain reaction.

ABSTRACT: The present study describes the development and evaluation of a duplex polymerase chain reaction (D-PCR) for diagnosis and simultaneous identification of tuberculous meningitis (TBM) and bacterial meningitis (BM) in a single reaction. A D-PCR with primers amplifying portions of the Mycobacterium tuberculosis IS6110 and the eubacteria 16SrDNA sequence in a same reaction mix was developed and tested on DNA extracted from 150 clinical CSF samples from different categories (TBM = 39, BM = 26, control infectious and non-infectious category = 85). The results indicate a clear differentiation between bands for eubacteria and M. tuberculosis with an analytical sensitivity of 10(3) cfu/ml for eubacteria and 10(2) cfu/ml for M. tuberculosis. When evaluated in clinical samples, D-PCR overall diagnosed 100 % confirmed TBM and 100 % confirmed BM cases with overall specificity of 96.5 %. D-PCR can be an effective tool for diagnosis and simultaneous identification of TBM or BM in a single PCR reaction. It saves time, cost, labour and sample amount and help in administration of appropriate antimicrobial therapy. The proposed diagnostic assay would be helpful in correct and rapid management of TBM and BM patients.

SUBMITTER: Nagdev KJ

PROVIDER: S-EPMC4363260 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Rapid diagnosis and simultaneous identification of tuberculous and bacterial meningitis by a newly developed duplex polymerase chain reaction.

Nagdev Khushboo J KJ Bhagchandani Sharda P SP Bhullar Shradha S SS Kapgate Rajkumar C RC Kashyap Rajpal S RS Chandak Nitin H NH Daginawala Hatim F HF Purohit Hemant J HJ Taori Girdhar M GM

Indian journal of microbiology 20150203 2

The present study describes the development and evaluation of a duplex polymerase chain reaction (D-PCR) for diagnosis and simultaneous identification of tuberculous meningitis (TBM) and bacterial meningitis (BM) in a single reaction. A D-PCR with primers amplifying portions of the Mycobacterium tuberculosis IS6110 and the eubacteria 16SrDNA sequence in a same reaction mix was developed and tested on DNA extracted from 150 clinical CSF samples from different categories (TBM = 39, BM = 26, contro ...[more]

PMID: 25805909

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Project description:Background and aimDifferent polymerase chain reaction (PCR) techniques have and are still being used for the direct detection of Brucella DNA in serum samples of different animal species and humans without being validated or properly validated, resulting in discrepancies. Thus, this study aimed to evaluate the diagnostic performance of the TaqMan Real-Time-PCR (RT-PCR) targeting the bcsp31 gene versus conventional PCR for the accurate diagnosis of brucellosis at the genus level in cattle sera.Materials and methodsOne hundred and eighty-four serum samples were collected from bacteriologically positive and negative cows with ages ranging from 1 to 5 years old at some infected private farms in the Nile Delta under quarantine measures as well as brucellosis free farms. These samples were classified into four groups after serological diagnosis and investigated by TaqMan RT-PCR and conventional PCR targeting the IS711 gene for Brucella DNA detection. The diagnostic performance characteristics of both PCR techniques were estimated considering the bacteriological results as a gold standard.ResultsTaqMan RT-PCR revealed superiority over conventional PCR; it was able to detect Brucella DNA in 95% (67/70) and 89% (25/28) of the cattle sera samples belonging to Group 1 (serologically and bacteriologically positive) and Group 2 (serologically negative but bacteriologically positive), respectively. On evaluating the diagnostic performance, TaqMan RT-PCR showed superior diagnostic sensitivity (93.9%), diagnostic specificity (88.4%), performance index (182.3), almost perfect kappa agreement (0.825±0.042), strong positive correlation (r=0.826), high accuracy based on the receiver operating characteristic (ROC) curve, and area under the ROC curve (0.911) at p<0.05 and CI of 95%.ConclusionA cattle serum sample is not the metric of choice for targeting Brucella genomic DNA by conventional PCR. The time-saving and rapid TaqMan RT-PCR method revealed a better diagnostic performance in the detection of Brucella DNA in cattle sera. Such performance offered by TaqMan RT-PCR may be considered a step toward the possibility of using such technology in the direct differentiation between Brucella-infected and -vaccinated cattle immunized by smooth vaccines from cattle sera using primers specific for such vaccines.

Dataset Information

Rapid diagnosis and simultaneous identification of tuberculous and bacterial meningitis by a newly developed duplex polymerase chain reaction.

Publications

Rapid diagnosis and simultaneous identification of tuberculous and bacterial meningitis by a newly developed duplex polymerase chain reaction.

Similar Datasets

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