49 Propidium monoazide (PMA) sample pretreatment impacts the abundance of rare populations in high-throughput sequencing analysis of CF lung mycobiome and bacteriome

Abstract

Objective Propidium monoazide (PMA) pretreatment has increasingly been used in molecular biology for detecting viable pathogens. Our study aims to determine whether a PMA pretreatment in high-throughput sequencing (HTS) could provide a more realistic picture of lung mycobiome and bacteriome from cystic fibrosis (CF) patients, and consequently reflect more closely the clinical status of patients. Methods We compared HTS data of bacteriome and mycobiome of 15 sputum samples from 5 CF patients that were characterized using the Ion Torrent technique with and without prior PMA treatment of the DNA-extracts. Results PMA pretreatment had no significant effect on the entire and abundant bacterial community (genera expressed as operational taxonomic unit with a relative abundance of ≥1%) but caused a significant difference in the rare biosphere community ( Conclusion To conclude, PMA pretreatment seems to change the relative abundance of bacteria, especially in the rare populations, but not fungi. Given such a cumbersome protocol (PMA pretreatment coupled with HTS), we discuss its potential interest within the follow-up of CF patients. As only few studies suggested the use of this protocol in the characterization of the bacteriome may be clinically relevant, further studies using PMA pretreatment are warranted to improve our omic knowledge.