MALDI-TOF-mass -spectrometry of protein extracts for ascertainment of phylogenetic relationships between clinical isolates of Aspergillus fumigatus and A. niger

Ref ID: 19521

Author:

IA Riabinin1,2*, NV Vasilyeva1,2, TS Bogomolova1,2, GA Chilina1, JV Michaylova1, ON Pinegina1,2,
SS Belanov3

Author address:

1Kashkin Research Institute of Medical Mycology, Mechnikov North-Western State Medical University,
Saint-Petersburg, Russia
2Chair of Medical Microbiology, Mechnikov North-Western State Medical University, Saint-Petersburg,
Russia
3Department of

Full conference title:

6th Advances Against Aspergillosis 2014

Abstract:

Purpose:
Aspergillus fumigatus and A. niger are common causative agents of aspergillosis in humans.
The possibility of nosocomial infection and antifungal drug resistance demand the creation
of universal methods for typing and “œparentage” Aspergillus clinical strains. There was a lot of
successful attempts to study Aspergillus molecular epidemiology using genetic methods such as
microsatellite typing, rep-PCR, RAPD-PCR, etc. Novel perspectives in this field appeared since
the beginning of MALDI-TOF-based identification of filamentous fungi. This paper analyzes the
possibility of using different processing methods of protein mass spectra obtained by MALDI-TOFMS
for the Aspergillii identification and molecular phylogeny.
Methods:
21 strains of A. fumigatus and 19 strains of A. niger isolated from clinical specimens during
2010 – 2013 years are preserved in Russian Collection of Pathogenic Fungi were studied. Isolates
were previously identified by morphology and sequencing of ITS1, ITS2 and β -tubulin regions
(Genetic Analyzer 3500, Applied Biosystems). Strains were sub-cultivated on Sabouraud dextrose
broth without agitation at 37° C overnight. Protein extraction from colonies was performed according
to Bruker standard protocol with formic acid and acetonitrile. MALDI-TOF-mass spectrometry was
made with using Autoflex speed TOF/TOF (Bruker Daltonics). Collected spectra were identified
with using “œFungi Library” database, then converted into MSP and compared by MSP-dendrogram,
hierarchical (5thdegree) principal component analysis (PCA) and matrix of composite correlation
index (CCI).
Results:
Rate of identification varied in wide range: for A. fumigatus strains – 1,833 – 2,234, for A. niger
strains 1,726 – 2,071. Spectra of several strains were identified with low rate, but MSP-dendrograms
show these spectra were closely related with reference-spectra of A. fumigatus and A. niger from
database respectively. PCA-clustering allowed to create the most detailed classifications, which
coincided in generally with the results of PCA-dendrograms (fig. 1), except some (sub)groups
containing single isolates. CCI-matrices showed these single strains (mass-spectra) were the most
deviated from studied isolates. In summary, both strains of A. fumigatus and strains of A. niger
were subdivided into 5 groups and subgroups. MSP-dendrograms, in contrast, were unsuitable for
separation of strains (spectra) groups.
Conclusion:
The ascertainment of phylogenetic relationships between Aspergillus spp. strains by MALDI-massspectra
analyzing tools (e.g. MSP-dendrogram) is helpful not only for epidemiological studies,
but although for primary species identification. CCI-matrix is suitable for demonstration a few
atypical isolates. Hierarchical PCA-clustering and dendrogram are probable methods of choice for
construction of subspecies classification of Aspergillii strains.

Fig 1. PCA-dendrograms and intraspecific classification of studied strains.

Abstract Number: 48

Conference Year: 2014

Link to conference website: http://www.AAA2014.org

New link: NULL

Conference abstracts, posters & presentations

Showing 10 posts of 17325 posts found.

  • Title

    Author

    Year

    Number

    Poster