sodC-Based Real-Time PCR for Detection of Neisseria meningitidis
Thomas, Jennifer Dolan, Hatcher, Cynthia P, Satterfield, Dara A, Theodore, M Jordan, Bach, Michelle C, Linscott, Kristin B, Zhao, Xin, Wang, Zin, Mair, Raydel, Schmink, Susanna, Arnold, Kathryn E, Stephens, David S, Harrison, Lee H, Hollick, Rosemary A, Andrade, Ana Lucia, Lamaro-Cardosa, Juliana, de Lemos, Ana Paula S, Gritzfeld, Jenna, Gordon, Stephen, Soysal, Ahmet, Bakir, Mustafa, Sharma, Dolly, Jain, Shabnam, Satola, Sarah W, Messonnier, Nancy and Mayer, Leonard W (2011) 'sodC-Based Real-Time PCR for Detection of Neisseria meningitidis'. PLoS ONE, Vol 6, Issue 5, e19361.
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Real-time PCR (rt-PCR) is a widely used molecular method for detection of Neisseria meningitidis (Nm). Several rt-PCR assays for Nm target the capsule transport gene, ctrA. However, over 16% of meningococcal carriage isolates lack ctrA, rendering this target gene ineffective at identification of this sub-population of meningococcal isolates. The Cu-Zn superoxide dismutase gene, sodC, is found in Nm but not in other Neisseria species. To better identify Nm, regardless of capsule genotype or expression status, a sodC-based TaqMan rt-PCR assay was developed and validated. Standard curves revealed an average lower limit of detection of 73 genomes per reaction at cycle threshold (Ct) value of 35, with 100% average reaction efficiency and an average R2 of 0.9925. 99.7% (624/626) of Nm isolates tested were sodC-positive, with a range of average Ct values from 13.0 to 29.5. The mean sodC Ct value of these Nm isolates was 17.6±2.2 (±SD). Of the 626 Nm tested, 178 were nongroupable (NG) ctrA-negative Nm isolates, and 98.9% (176/178) of these were detected by sodC rt-PCR. The assay was 100% specific, with all 244 non-Nm isolates testing negative. Of 157 clinical specimens tested, sodC detected 25/157 Nm or 4 additional specimens compared to ctrA and 24 more than culture. Among 582 carriage specimens, sodC detected Nm in 1 more than ctrA and in 4 more than culture. This sodC rt-PCR assay is a highly sensitive and specific method for detection of Nm, especially in carriage studies where many meningococcal isolates lack capsule genes.
|Subjects:||QW Microbiology and Immunology > QW 50 Bacteria (General). Bacteriology. Archaea|
WL Nervous System > WL 200 Meninges. Blood-brain barrier
|Groups:||Clinical Group (2002-2012)|
|Digital Object Identifer (DOI):||10.1371/journal.pone.0019361|
|Depositing User:||Users 43 not found.|
|Date Deposited:||16 May 2011 15:50|
|Last Modified:||28 May 2015 14:57|
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