K MULRONEY1, J HALL2, T INGLIS2, A CHAKERA1, 3,
1 Translational Renal Research Group, Harry Perkins Institute of Medical Research, Perth, Western Australia; 2Department of Microbiology, PathWest Laboratory Medicine, Perth, Australia; 3Renal Unit, Sir Charles Gairdner Hospital, Perth, Western Australia
Aim: To assess the sensitivity and specificity of flow cytometry to directly identify bacteria in peritoneal dialysate from patients with suspected peritoneal dialysis associated peritonitis.
Background: Current diagnostic testing of dialysate from patients with suspected peritonitis is based on traditional culture-dependent microbiology. These assays take 1-3 days to become positive and culture-negative rates of over 20% are reported. There has been limited use of culture-independent microbiology techniques for detection of bacteria in dialysate, and none have been routinely adopted in a clinical setting. Flow cytometry based assessment of dialysate may provide a more rapid and accurate tool to detect bacteria in dialysate.
Methods: Dialysate samples from 10 patients with peritonitis were analysed by two PCR-based methods (16S PCR, and BioFire FilmArray®) and flow cytometry using an Attune NxT flow cytometer with 10 μM SYTO® 9, 2× Fixable Near-IR amine-reactive viability dye for identification of bacterial cell populations. Results were compared to traditional culture using BacTec bottles with bacterial identification confirmed by MALDI-TOF Biotyper analysis. Results: 16S PCR was accurate in 30% of samples, whereas the BioFire FilmArray® was accurate in 20% of cases. Flow cytometry was accurate in 90% of cases, with no false negative results. Direct detection of bacteria in dialysate substantially reduces the time for confirm infection compared to traditional culture.
Conclusions: Flow cytometry is an accurate technique for the detection of bacteria in dialysate samples that provides significant advantages over nucleic-acid based detection methods in terms of accuracy and traditional culture with regards to the time to obtain a result.