Establishment of a matrix-assisted laser desorption ionization time-of-flight mass spectrometry database for rapid identification of infectious achlorophyllous green micro-algae of the genus Prototheca

Abstract

The possibility of using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for rapid identification of pathogenic and non-pathogenic species of the genus Prototheca has been recently demonstrated. A unique reference database of MALDI-TOF MS profiles for type and reference strains of the six generally accepted Prototheca species was established. The database quality was reinforced after the acquisition of 27 spectra for selected Prototheca strains, with three biological and technical replicates for each of 18 type and reference strains of Prototheca and four strains of Chlorella. This provides reproducible and unique spectra covering a wide m/z range (2000-20 000 Da) for each of the strains used in the present study. The reproducibility of the spectra was further confirmed by employing composite correlation index calculation and main spectra library (MSP) dendrogram creation, available with MALDI Biotyper software. The MSP dendrograms obtained were comparable with the 18S rDNA sequence-based dendrograms. These reference spectra were successfully added to the Bruker database, and the efficiency of identification was evaluated by cross-reference-based and unknown Prototheca identification. It is proposed that the addition of further strains would reinforce the reference spectra library for rapid identification of Prototheca strains to the genus and species/genotype level.

The Construction and Evaluation of Reference Spectra for the Identification of Human Pathogenic Microorganisms by MALDI-TOF MS

“…Previous studies reported MBS identification accuracies of 79.7–100% at the species level [11],[15]–[18],[22]–[26]. However, in this study, the accurate identification rates for 1019 strains in searches of the ORD (with 3995 PMRS) were 70.7% at the genus level and only 50.7% at the species level.…”

“…A complete database of microorganism PMRS is a prerequisite for ensuring that the MBS will exhibit strong identification capabilities [11], [14], [19]–[21]. The pathogens examined in this study included 94 different pathogen species (bacterial species, Mycoplasma spp., and Spirochaeta spp.)…”

Phylogenetic analysis of microalgae based on highly abundant proteins using mass spectrometry

“…In this study we described the MALDI-TOF MS-based method for monitoring the occurrence of HABs by rapid identification of harmful microalgae, as shown in previous studies for infectious bacteria [20] and achlorophyllous microalgae [18]. This rapid identification method, in combination with comprehensive profile databases of the highly abundant proteins of diverse toxic and non-toxic strains, could be effectively applied to identify eukaryotic microalgae or cyanobacteria during HAB events in fresh water and ocean environments.…”

“…MALDI-TOF MS can rapidly detect cyanobacteria and microalgae during HAB events in fresh water and ocean environments more rapidly than morphological and genetic identification. A previous study reported using MALDI-TOF MS to distinguish between pathogenic and non-pathogenic genotypes of the achlorophyllous microalgae species [18]. Thus the MALDI-TOF MS approach can be used to discriminate between toxic and non-toxic strains of the same species, or species of the same genus if MS databases including diverse toxic and non-toxic strains are first established.…”

Prototheca miyajii sp. nov., isolated from a patient with systemic protothecosis

“…Our previous studies have shown that comprehensive analysis by PCR of the SSU rDNA, internal transcribed spacer (ITS) and the LSU D1/D2 region is also useful for identification of species of the genus Prototheca (Hirose et al, 2013). In addition, an increasing number of studies demonstrate that matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) could be applied to the identification of species and genotypes of Prototheca (von Bergen et al, 2009;Ahrholdt et al, 2012;Murugaiyan et al, 2012;Wirth et al, 2012).…”