A New Highly Sensitive and Specific Real-Time PCR Assay Targeting the Malate Dehydrogenase Gene of Kingella kingae and Application to 201 Pediatric Clinical Specimens.

is a significant pediatric pathogen responsible for bone and joint infections, occult bacteremia, and endocarditis in early childhood. Past efforts to detect this bacterium using culture and broad-range 16S rRNA gene PCR assays from clinical specimens have proven unsatisfactory; therefore, by the late 2000s, these were gradually phased out to explore the benefits of specific real-time PCR tests targeting the gene and the RTX locus of However, recent studies showed that real-time PCR (RT-PCR) assays targeting the sp. RTX locus that are currently available for the diagnosis of infection lack specificity because they could not distinguish between and the recently described species. Furthermore, analysis of the gene from a large collection of 45 strains showed that primers and probes from -based RT-PCR assays display a few mismatches with variations that may result in decreased detection sensitivity, especially in paucibacillary clinical specimens. In order to provide an alternative to - and RTX-targeting RT-PCR assays that may suffer from suboptimal specificity and sensitivity, a -specific RT-PCR assay targeting the malate dehydrogenase () gene was developed for predicting no mismatch between primers and probe and 18 variants of the gene from 20 distinct sequence types of This novel -specific RT-PCR assay demonstrated high specificity and sensitivity and was successfully used to diagnose infections and carriage in 104 clinical specimens from children between 7 months and 7 years old.