Quick and wide-range taxonomical repertoire establishment of the cystic fibrosis lung microbiota by tandem mass spectrometry on sputum samples

Abstract

Microorganisms proteotyping by tandem mass spectrometry has been recently shown as a powerful methodology to identify the wide-range taxonomy and biomass of microbiota. Sputum is the recommended specimen for routine microbiological monitoring of Cystic Fibrosis (CF) patients but has been rarely submitted to tandem mass spectrometry-based proteotyping. In this study, we compared the microbial components of spontaneous and induced sputum samples from three cystic fibrosis patients. Although the presence of microbial proteins is much lower than host proteins, we report that the microbiota’s components present in the samples can be identified, as well as host biomarkers and functional insights into the microbiota. No significant difference was found in microorganism abundance between paired spontaneous and induced sputum samples. Microbial proteins linked to resistance, iron uptake, and biofilm-forming ability were observed in sputa independently of the sampling method. This unbiased and enlarged view of the CF microbiome could be highly complementary to culture and relevant for the clinical management of CF patients by improving knowledge about the host-pathogen dynamics and CF pathophysiology.

Metaproteomics to understand how microbiota function: The crystal ball predicts a promising future

“… Metaproteomics can identify and quantify uncharted branches of the tree of life. Tandem mass spectrometry‐based proteotyping of organisms present in the sample can be based on taxa‐spectrum matches (TSMs) and taxon‐specific peptides (Hardouin et al, 2022 ; Lozano et al, 2022 ). In this figure focused on a theoretical archaeal enriched microbial community, the presence of an uncharacterized organism belonging to the Candidatus Lokyarchaeota phylum and another belonging to the Halobacteriales order are indicated, while other organisms characterized at the genus taxonomical rank are also confirmed.…”

“…Sample preparation can also be reduced to less than an hour (Hayoun et al, 2020 ), except for challenging samples such as soils (Herruzo‐Ruiz et al, 2021 ; Keiblinger et al, 2012 ) or for more specific sample preparations (Salvato et al, 2022 ). An automated, refined interpretation of metaproteomics results based on cascade searches applicable to any sample can be completed within 2 h (Hardouin et al, 2022 ). Thus, currently, the time between sampling and result could be less than 5 h (Figure 4 ) compared to 24 h workflow proposed 3 years ago (Heyer et al, 2019 ), leaving plenty of leeways to optimize the various stages of the procedure and obtain results even faster, thanks to improved protocols and greater computing power.…”

“…This information could also serve to obtain a quick modelled F I G U R E 4 Clinical and environmental diagnostics by metaproteomics, timelines and applications. A sample-to-result timeline is proposed based on previously published results (Hardouin et al, 2022), along with likely optimizations over the next decade. The fields of application of metaproteomics for diagnosis or routine analysis are schematized.…”

A Simplified Label-Free Method for Proteotyping Sets of Six Isolates in a Single Liquid Chromatography-High-Resolution Tandem Mass Spectrometry Analysis

“…It is even compatible with mixtures of microorganisms which would represent a deadlock for whole-cell MALDI-TOF. , High-throughput application of this recent technology is feasible in 96-well plate format and is robust in terms of bacterial loads . As a result, tandem mass spectrometry proteotyping has been applied in clinical diagnostics to discriminate Pneumococcus species within the Mitis group, to identify Francisella directly from hare carcasses without the need to perform cultures, to detect SARS-CoV-2 coronaviruses, to screen marine isolates, to identify the microbial flora in samples from cystic fibrosis patients, to authenticate historical remains, and to diagnose infectious disease from 15-century-old bones . Moreover, thanks to the capacity to analyze multiplex samples without the need for additional reagents, the methodology can be cost-effective, as recently shown .…”

An integrated metaproteomics workflow for studying host-microbe dynamics in bronchoalveolar lavage samples applied to cystic fibrosis disease

“…The upshot of successful metaproteomic analysis is the generation of unique information on proteins expressed by translationally active bacteria, along with a profile of human host proteins, within clinically important samples. Not surprisingly, metaproteomics has shown value in studying the role of microorganisms in CF airways in recent years ( 3 , 39 ).…”