Extracellular vesicle-based diagnosis of Mucormycosis: Candidate biomarkers, analytical challenges, and clinical translation

Currently, no standard serological test is available for the detection of Mucorales. Mucormycosis is a rapidly progressing angioinvasive infection with a high mortality rate. However, since a diagnostically useful target (1 → 3)-β-D-glucan or Aspergillus galactomannan is not present in Mucorales, a negative result for such targets is a true negative and does not rule out the diagnosis of mucormycosis. PCR detection using PCR for Mucorales can be performed in the early stages, and there is a commercially available quantitative PCR kit in some places, although it is not uniform. Extracellular vesicles (EVs) are membrane-bound particles present in biofluids that play a protective role against the degradation of proteins and nucleic acids and allow for their enrichment prior to measurement. Two studies provided evidence of Mucorales direct culture using extracellular RNA profiles from Rhizopus delemar and proteomic characterization of Mucor lusitanicus EVs, respectively; however, neither resulted in a patient-derived diagnostic test. This structured narrative review separates Mucorales EV direct evidence from extrapolations from other fungi and clinical EV assays. We recommend using a patient-derived discovery workflow involving EV-enriched RNA sequencing, EV-associated DNA analysis, and proteomics, followed by a locked multiplex assay of two or three independent fungal targets. Therefore, molecules from the CotH family, mucoricin, glucoamylase, iron-acquisition factors, and host-response signals should be considered as candidates and should be directly demonstrated within EVs from patients, rather than being considered as known EV markers.