Detection of Fusobacterium nucleatum subspecies in the saliva of pre-colorectal cancer patients, using tandem mass spectrometry

Abstract

Objective: Rising evidence links Fusobacterium nucleatum (F. nucleatum) with its four subspecies; nucleatum, polymorphum, animalis, and vincentii, with the development of colorectal cancer (CRC) and its precursor colorectal adenoma (CRA). This study aims to optimize a technique for and explore the capability of matrix-assisted laser-desorption ionization-tandem time-of-flight mass spectrometry (MALDI-TOF/TOF MS) to detect F. nucleatum subspecies directly from the saliva samples of CRA patients and controls without preculturing.

Design: Saliva samples were collected from four CRA patients and eight controls. Proteins were extracted and subjected to solid-phase extraction fractionation, enzymatically digested, and analyzed by MALDI-TOF/TOF MS. F. nucleatum subspecies strains were cultured and used as a positive control.

Results: A proteomics approach was developed to identify F. nucleatum subspecies directly from saliva samples. With this approach, the bacterial culturing step, which could take up to seven days, was bypassed. Overall, 157 F. nucleatum subspecies proteins were detected in the saliva samples. F. nucleatum subsp. nucleatum was absent in the patients while detected in half of the controls.

Conclusion: This study presents a novel technique for detecting F. nucleatum subspecies from saliva specimens that could later be employed to better understand a potential role of those subspecies in CRC development.

Keywords: Adenoma; Anaerobic bacteria; Colorectal cancer; Fusobacterium nucleatum; Saliva; Tandem mass spectrometry.