Original ArtIcle

Application of 16S rRNA Gene-Targeted Next-Generation Sequencing for Bacterial Pathogen Detection in Continuous Ambulatory Peritoneal Dialysis Peritonitis

Young Ah Kim¹ Ea Wha Kang² Hye Su Moon⁴ Daewon Kim^3,4 Dongeun Yong^{3,4 *}

¹Department of Laboratory Medicine
²Department of Internal Medicine, National Health Insurance Service Ilsan Hospital, Goyang
³Department of Laboratory Medicine
⁴Research Institute of Bacterial Resistance, Yonsei University College of Medicine, Seoul, Korea

Abstract

16S rRNA gene-targeted next-generation sequencing (NGS) can detect microorganisms in a comprehensive reference database. To date, NGS has been successfully applied to samples such as urine, blood, and synovial fluid. However, there is no data for continuous ambulatory peritoneal dialysis (CAPD) fluid. The purpose of this study was to evaluate the clinical usefulness of microbiome analysis of CAPD fluids for the diagnosis of CAPD peritonitis.

We included 21 patients with high suspicion of CAPD peritonitis. Routine CAPD fluid culture was performed using a pellet of 50 mL CAPD fluid onto the chocolate and blood agar for two days, and thioglycollate broth for one week. 16S rRNA gene-targeted NGS of pellets, stored at -70°C was performed with MiSeq (Illumina, USA).

Many colonized or pathogenic bacteria were detected from CAPD fluids using NGS and the microbiomes were composed of 1 to 29 genera with a cut-off 1.0. Compared to the culture results, NGS detected the same pathogens in 6 of 18 valid results (three samples failed with low read count). Additionally, using NGS, anaerobes such as Bacteroides spp. and Prevotella spp. were detected in six patients. In two of five samples in which no bacterial growth was detected, possible pathogens were detected by NGS.

To our knowledge, this is the first report about the application of 16S rRNA gene-targeted NGS for diagnosis of CAPD peritonitis. Etiology of culture-negative CAPD peritonitis can be better defined in NGS. Furthermore, it also helped the detection of anaerobic bacteria.

Figures & Tables

Fig. 1. 1. Composition of gDNA Mock V34 (1707_C) and MiSeq QC (1903_C) (A), the principal coordinates analysis with CAPD fluid samples with QC controls in each run (B).Abbreviation: CAPD, continuous ambulatory peritoneal dialysis; QC, quality control; OTU, operational taxonomic unit