#97: Rapid, Noninvasive Detection of Kingella kingae Pediatric Vertebral Infections Using a Microbial Cell-free DNA Sequencing Test for Pathogen Identification
Authors: Sarmiento Adriana , Penella Ana Del Valle , Laufer P Marcelo , Sanchez-Vegas Carolina , Cotilla Manuel R , Trieu Connie , Arnold John C , Villegas Alejandro Jordan , Lindner Martin S , Ahmed Asim A 
Published: March 2021

Abstract

Background
Kingella kingae is a recognized cause of bone and joint infections (BJI) in infants. The diagnosis of Kingella kingae BJI can be challenging due to its fastidious growth with conventional culturing methods even when infected tissue is obtained. Kingella kingae spinal infections are likely an underdiagnosed entity given the limitations of culture-based methods and the reluctance to biopsy spinal locations of infection (in favor of empiric treatment). This approach often necessitates MRSA coverage. A sensitive, rapid, noninvasive diagnostic approach to pediatric vertebral infections would enable targeted therapy. Detection of circulating microbial cell-free DNA (mcfDNA) in the plasma originating from areas of sequestered infection through next-generation sequencing (NGS) has shown utility in pediatric pneumonia (Farnaes et al. DMID 2019) and a wide variety of infections in the immunocompromised host (Rossoff et al. OFID 2019) and potentially offers promise in resolving the etiology of pediatric vertebral infections.

Methods
The Karius test is a CLIA-certified/CAP-accredited NGS plasma test that detects circulating mcfDNA in the blood. After mcfDNA is extracted and NGS performed, human sequences are removed and remaining sequences are aligned to a curated pathogen database of &gt;1400 organisms. Organisms present above a statistical threshold are reported and quantified. The time to result reporting is on average 24 hours from sample receipt. Karius Test results over the prior 2 years were reviewed for detections of Kingella kingae in the context of spinal infections. Clinical chart review was performed by the treating pediatric infectious diseases physicians at each participating institution after IRB notification and approval.

Results
Six cases of Kingella kingae pediatric vertebral infections were identified across five institutions; clinical data were available for five cases across four institutions (see Table). Four cases were male; the average age was 15.3 months. Four of five cases had an antecedent URI. The clinical presentations were characterized by decreased mobility and relatively bland inflammatory response (lack of fever, bland inflammatory markers). The lumbar region was the most commonly affected vertebral location (80%). Blood cultures were negative in all cases; empiric anti-MRSA therapy was initiated in all cases. The time to result of Kingella kingae mcfDNA detection in the plasma was one day from sample receipt in all cases. McfDNA from co-pathogens were detected in 66.7% of cases (Haemophilus influenzae was the most common). The detection of Kingella kingae by the Karius test influenced a decision to narrow coverage in 80% of cases and a decision to forego biopsy in 60% of cases.

Conclusion
Plasma NGS for circulating mcfDNA offers a rapid, noninvasive means of detecting Kingella kingae pediatric vertebral infection. This culture-independent approach may enable specific diagnosis despite antibiotic pretreatment and obviate the need for an invasive procedure. Accurate identification of Kingella kingae has important implications on antibiotic stewardship enabling targeted therapy without the reliance on empiric MRSA coverage. Given the capacity to detect over 1400 organisms from a single sample NGS for mcfDNA offers a means to detect a broad variety of pathogens known to have predilection to cause pediatric spine infection.