Pathogen proteotyping: A rapidly developing application of mass spectrometry to address clinical concerns.

For the rapid and reliable differentiation of clinically-relevant bacterial species, mass spectrometry-based methods have emerged in recent years as valid alternatives to existing techniques. Mass profiles generated by whole-cell Matrix-Assisted Laser Desorption Ionization-Time of Flight mass spectrometry have revolutionized microorganism identification and proven their potential for proteotyping at the species level. Indeed, the methodology has been widely deployed in clinical settings. However, the low resolution and dynamic range of the methodology has limited its capacity to distinguish between subspecies. This discrimination capacity is pivotal in cases where certain strains display virulence or antibiotic resistance, and for epidemiologic analyses. Moreover, sensitivity and specificity are both key parameters when attempting to discriminate between microorganisms present in complex multi-pathogenic samples. These two parameters are also essential to meet the growing interest in the characterization of microorganisms contained within even more complex samples, such as the human microbiome. Tandem mass spectrometry, with its high resolution, holds great potential for use in the real-time direct analysis of pathogens at the most relevant taxonomic rank in routine clinical practice. This review explores the numerous benefits and challenges of implementing advanced proteotyping methods, based on tandem mass spectrometry, in clinical laboratories. We provide an overview of the current applications and methodologies, while also discussing recent improvements and potential new approaches for typing, as well as their future applications.