# P-534. Comparison of Concurrent (Same or Next-Day) Conventional Microbial Testing to Next Generation Sequencing (NGS) Results: Review of Pediatric Samples over 4-Years (2020-2024)

## TL;DR

This study compares next-generation sequencing (NGS) with conventional microbial testing in pediatric patients, finding that NGS detects viruses well but has limitations in detecting bacteria and fungi.

## Contribution

The study provides insights into the reliability of NGS in pediatric infectious disease diagnosis compared to traditional methods.

## Key findings

- NGS reliably detected viruses when viral load was above quantification thresholds.
- NGS missed most bacterial infections outside the bloodstream and failed to detect certain fungal infections like dimorphic fungi and Cryptococcus.
- NGS identified invasive fungal infections that could not be cultured in two cases.

## Abstract

The role NGS has in diagnosis of infections remains unclear as high costs currently limit use. Strengths and limitations of this testing modality are still being studied.

271 plasma NGS (Karius Test ®) samples over 4 years were evaluated for the presence of conventional microbial testing occurring within one day. There were 68 instances for comparison (average 20.6 hours apart). Wound, non-BAL respiratory, stool and bacterial urine cultures were excluded. Positive NGS paired to negative blood culture were also excluded.

Viral: There was high concordance between NGS and viral plasma PCR (Table 1, Figure 1). When PCR viral load was above the threshold of quantification NGS successfully detected all cases of adenovirus, CMV, EBV, HSV1, and HHV6 (n=22). NGS detected CMV when PCR did not once. Viral PCR detection below the level of quantification (n=10) always corresponded to negative NGS. BK virus detection was variable. Twice PCR detected BK above quantification (139 and 9,530 copies/ml) but NGS did not. NGS detected BK once when it was undetected by PCR. Viral infections in other body compartments were inconsistently detected by NGS (Table 2).

Bacterial: All positive blood cultures corresponded to positive NGS except two cases of staphylococcus epidermidis, likely contaminants. NGS identified Staphylococcus epidermidis when Staphylococcus haemolyticus grew from culture. Most (5/6) cases of bacteria cultured from other body sites were missed by NGS although only two cases had greater than rare growth (Table 3).

Fungal: NGS did not detect dimorphic fungi in five instances of Blastomyces or Histoplasma antigen positivity. NGS failed to detect Cryptococcus neoformans fungemia and Rhizopus with dermal angioinvasion. NGS identified invasive fungal infections unable to be cultured twice (Rhizopus, Aspergillus).

Plasma NGS detected viruses when above the threshold for quantification but not when below the limit of quantification. BK virus had variable detection. NGS identified bacteremia well. A positive blood culture with negative NGS suggests contamination and favors antibiotic de-escalation. Detection of bacterial infections outside the bloodstream is unreliable. Dimorphic fungi and Cryptococcus were not detected by NGS, while Candida and molds were.

All Authors: No reported disclosures