Abstract
BackgroundThe Cryptococcus neoformans/C. gattii species complex comprises two sibling species that are divided into eight major molecular types, C. neoformans VNI to VNIV and C. gattii VGI to VGIV. These genotypes differ in host range, epidemiology, virulence, antifungal susceptibility and geographic distribution. The currently used phenotypic and molecular identification methods for the species/molecular types are time consuming and expensive. As Matrix-Assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS) offers an effective alternative for the rapid identification of microorganisms, the objective of this study was to examine its potential for the identification of C. neoformans and C. gattii strains at the intra- and inter-species level.MethodologyProtein extracts obtained via the formic acid extraction method of 164 C. neoformans/C. gattii isolates, including four inter-species hybrids, were studied.ResultsThe obtained mass spectra correctly identified 100% of all studied isolates, grouped each isolate according to the currently recognized species, C. neoformans and C. gattii, and detected potential hybrids. In addition, all isolates were clearly separated according to their major molecular type, generating greater spectral differences among the C. neoformans molecular types than the C. gattii molecular types, most likely reflecting a closer phylogenetic relationship between the latter. The number of colonies used and the incubation length did not affect the results. No spectra were obtained from intact yeast cells. An extended validated spectral library containing spectra of all eight major molecular types was established.ConclusionsMALDI-TOF MS is a rapid identification tool for the correct recognition of the two currently recognized human pathogenic Cryptococcus species and offers a simple method for the separation of the eight major molecular types and the detection of hybrid strains within this species complex in the clinical laboratory. The obtained mass spectra provide further evidence that the major molecular types warrant variety or even species status.
Highlights
Cryptococcosis is a life-threatening infection caused for the inhalation of infectious propagules of the encapsulated yeasts Cryptococcus neoformans or C. gattii [1]
All isolates were clearly separated according to their major molecular type, generating greater spectral differences among the C. neoformans molecular types than the C. gattii molecular types, most likely reflecting a closer phylogenetic relationship between the latter
The analysis of 20 isolates for each of the eight major molecular types of the C. neoformans/C. gattii species complex by MALDITOF MS, resulted in a library of 960 individual mass spectra and 160 combined mass spectra, which allowed for the correct identification of all isolates according to the two currently recognized species C. neoformans and C. gattii, with identification scores ranging between 1.882 and 2.711 (Table 1)
Summary
Cryptococcosis is a life-threatening infection caused for the inhalation of infectious propagules of the encapsulated yeasts Cryptococcus neoformans or C. gattii [1]. Neoformans (serotype D) and the well-recognized hybrid AD is mainly an opportunistic infection of HIV patients [2,3,4], while C. gattii (serotypes B and C) is mainly affecting immunocompetent hosts, involving humans, domestic and wild animals [2,5,6] Both species can be differentiated from each other by phenotypic methods [7], and at intra-specific level, by a range of genotyping methods, including PCR fingerprinting [8,9], restriction fragment length polymorphism (RFLP) analysis [10], amplified fragment length polymorphism (AFLP) analysis [11], multilocus microsatellite typing (MLMT) [12] and multilocus sequence typing (MLST) [13,14]. The Cryptococcus neoformans/C. gattii species complex comprises two sibling species that are divided into eight major molecular types, C. neoformans VNI to VNIV and C. gattii VGI to VGIV These genotypes differ in host range, epidemiology, virulence, antifungal susceptibility and geographic distribution. As Matrix-Assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS) offers an effective alternative for the rapid identification of microorganisms, the objective of this study was to examine its potential for the identification of C. neoformans and C. gattii strains at the intra- and inter-species level
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
