Intact spore MALDI-TOF mass spectrometry and proteomic analysis of Puccinia pathogenic fungi

Beinhauer, Jana; Raus, Martin; Hanzalová, A.; Horčička, P.; Šebela, Marek

doi:10.1016/j.bbapap.2016.06.002

Cited by 9 publications

(8 citation statements)

References 52 publications

(85 reference statements)

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“…Li et al (2000) and Welham et al (2000) were the first to transfer the application from bacteria to spores of clinically relevant filamentous fungi. Increasing numbers of protocols have been published to analyze fungal spores by MALDI-TOF MS for a broad variety of fungi and oomycetes, including many phytopathogens (Böhmer et al 2007;Kemptner et al 2009;Sulc et al 2009;Bhadauria et al 2010;Brun et al 2013;Beinhauer et al 2016). The technique was also applied to distinguish closely related species (Gruenwald et al 2015;Wigmann et al 2019) and could therefore also be a useful tool for the discrimination of the wheat bunt fungi T. caries, T. laevis, and T. controversa.…”

Section: Introductionmentioning

confidence: 99%

Discrimination of Tilletia controversa from the T. caries/T. laevis complex by MALDI-TOF MS analysis of teliospores

Förster

Sedaghatjoo

Maier

et al. 2022

Appl Microbiol Biotechnol

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The fungal genus Tilletia includes a large number of plant pathogens of Poaceae. Only a few of those cause bunt of wheat, but these species can lead to significant yield losses in crop production worldwide. Due to quarantine regulations and specific disease control using appropriate seed treatments for the different disease agents, it is of high importance to distinguish Tilletia caries and Tilletia laevis as causal agents of common bunt accurately from Tilletia controversa, the causal agent of the dwarf bunt. Several studies have shown that matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) is a useful tool to differentiate closely related fungal species. The aim of this study was to assess whether MALDI-TOF MS analysis is able to distinguish specimens of the three closely related pathogens T. caries, T. laevis, and T. controversa and whether it may constitute an alternative method to the morphology-based identification or germination tests. Spectral data are available via ProteomeXchange with identifier PXD030401. Spectra-based hierarchical cluster analysis (HCA) and discriminant analysis of principal components (DAPC) of the obtained mass spectra showed two main clusters. One cluster included specimens of T. controversa, whereas the second cluster comprised T. laevis and T. caries specimens. Even though main spectral profiles (MSPs) for species identification are missing, MALDI-TOF MS has proven to be a useful method for distinguishing between T. controversa and the two causal agents of common bunt, using direct analysis of teliospores, but was unable to separate T. caries and T. laevis species. Key points • MALDI-TOF MS was developed to classify Tilletia species causing bunt of wheat. • Best results were achieved when combining HCA and DAPC analysis. • The method resulted in an accuracy of 98.51% testing 67 Tilletia specimens. Graphical abstract

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Section: Introductionmentioning

confidence: 99%

Discrimination of Tilletia controversa from the T. caries/T. laevis complex by MALDI-TOF MS analysis of teliospores

Förster

Sedaghatjoo

Maier

et al. 2022

Appl Microbiol Biotechnol

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“…A binary FA:SA matrix was chosen to prepare MALDI sample spots on the target. This combination of matrix compounds has been shown a good choice to measure MALDI profile spectra of fungal intact cells or spores [11,31]. Evaluation tests with CHCA, saturated solution in 50% ACN with 2.5% (v/v) TFA (recommended for bacteria by the manufacturer of the MALDI Biotyper system), were not satisfactory enough as regards to the number and intensity of peaks.…”

Section: Resultsmentioning

confidence: 99%

“…But there is the possibility to extract peptides and proteins from a large initial amount of microbial cells (30–100 mg) under conditions mimicking the on-target extraction process followed by their digestion and identification via reversed-phase liquid chromatography of peptides coupled to MS/MS analysis. Then the resulting sequence-based molecular mass data can be assigned to the m/z values acquired in IC MALDI-MS. We have recently applied this approach to plant pathogenic fungi [11].…”

Section: Discussionmentioning

confidence: 99%

“…without any destructive pretreatment) or peptide/protein extracts. Then peptide/protein profile mass spectra are acquired, which is facilitated by choosing a proper matrix compound as a part of the sample preparation step [11]. The mass range for the acquisition is typically between m/z 1000 and 20,000 and the profile spectrum pattern is finally compared with a reference spectral database for identification [12].…”

Section: Introductionmentioning

confidence: 99%

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Intact cell MALDI-TOF mass spectrometric analysis of Chroococcidiopsis cyanobacteria for classification purposes and identification of possible marker proteins

et al. 2018

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Cyanobacteria represent a bacterial phyllum characteristic by the ability to photosynthesize. They are potentially applicable for the production of useful compounds but may also cause poisoning or at least health problems as they can produce cyanotoxins. The introduction of a fast methodology is important not only for fundamental taxonomic purposes, but also for reliable identifications in biological studies. In this work, we have used matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of intact cells to study Chroococcidiopsis strains. A library of the obtained reference mass spectra containing characteristic peptide/protein profiles was examined by software tools to characterize similarities and differences applicable for diagnostics and taxonomy. Both a similarity tree and heat map constructed from the mass spectrometric data proved consistent with 16S rRNA sequencing results. We show as novelty that a binary matrix combining ferulic and sinapinic acids performs well in acquiring reproducible mass spectra of cyanobacteria. Using the matrix solvent, a protein extraction from cells was done. After polyacrylamide gel electrophoresis, the separated protein fractions were in-gel digested and the resulting peptides analyzed by liquid chromatography coupled with tandem mass spectrometry. For the first time, photosystem protein components, phycobilisome proteins, electron transport proteins, nitrogen-metabolism and nucleic acids binding-proteins, cytochromes plus other enzymes and various uncharacterized proteins could be assigned to characteristic peaks in the mass spectrometric profiles and some of them suggested as markers in addition to 30S and 50S ribosomal proteins known from previous studies employing intact cell mass spectrometry of microorganisms.

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“…Spores from only a few plant fungal pathogens have been subjected to compositional analyses, including Uromyces appendiculatus , a rust fungus infecting wheat and beans, Blumeria graminis f. sp. hordei , an ascomycete causing barley powdery mildew, Magnaporthe oryzae , the causative agent of rice blast disease, several Puccinia rust basidiomycetes, and Colletotrichum acutatum that infects several commercially valuable fruit crops (Cooper et al 2006; Noir et al 2009; Gokce et al 2012; El-Akhal et al 2013; Quecine et al 2016; Zhang et al 2015; Beinhauer et al 2016). Proteomic studies of these spores showed that proteins involved in common processes such as protein homeostasis and metabolism are readily detectable in those spores, which is consistent with stress resistance and rapid growth during germination; however, one would expect proteins involved in those processes to also be important for vegetative growth.…”

Section: Pathogenic Fungal Sporesmentioning

confidence: 99%

Sporulation: how to survive on planet Earth (and beyond)

Huang

Hull

2017

Curr Genet

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Sporulation is a strategy widely utilized by a wide variety of organisms to adapt to changes in their individual environmental niches and survive in time and/or space until they encounter conditions acceptable for vegetative growth. The spores produced by bacteria have been the subjects of extensive studies, and several systems such as Bacillus subtilis have provided ample opportunities to understand the molecular basis of spore biogenesis and germination. In contrast, the spores of other microbes, such as fungi, are relatively poorly understood. Studies of sporulation in model systems such as Saccharomyces cerevisiae and Aspergillus nidulans have established a basis for investigating eukaryotic spores, but very little is known at the molecular level about how spores function. This is especially true among the spores of human fungal pathogens such as the most common cause of fatal fungal disease, Cryptococcus neoformans. Recent proteomic studies are helping to determine the molecular mechanisms by which pathogenic fungal spores are formed, persist, and germinate into actively growing agents of human disease.

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Intact spore MALDI-TOF mass spectrometry and proteomic analysis of Puccinia pathogenic fungi

Cited by 9 publications

References 52 publications

Discrimination of Tilletia controversa from the T. caries/T. laevis complex by MALDI-TOF MS analysis of teliospores

Discrimination of Tilletia controversa from the T. caries/T. laevis complex by MALDI-TOF MS analysis of teliospores

Intact cell MALDI-TOF mass spectrometric analysis of Chroococcidiopsis cyanobacteria for classification purposes and identification of possible marker proteins

Sporulation: how to survive on planet Earth (and beyond)

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