ABSTRACT
Fungi belonging to the Cryptococcus neoformans/C. gattii species complex (CNGSC) are pathogens causing severe infections in humans and animals, that for humans may result in a mortality rate ranging up to 70%. The CNGSC is divided into eight major molecular types, that may differ in their virulence and susceptibility. In order to fully understand the epidemiology of cryptococcosis, it is important to study the world distribution and population structure of these pathogens. The present study is the first presenting a population of strains isolated in Poland and one of the few using a multi-species animal group as a source of the specimen. The pathogen was present in 2.375% of the tested animals. The URA5-RFLP and MALDI-TOF MS analyses have revealed that the population consisted exclusively of C. neoformans strains, with a predominance of major molecular type VNIV (C. neoformans var. neoformans). The MALDI-TOF MS was used to perform the CNGSC strains identification on both the species and sub-species level. Despite the fact that the animals providing the specimens were not treated with 5-fluorocytosine, around 10% of the tested population presented MIC values exceeding 64 mg/L, indicating the existence of the 5-fluorocytosine-resistant strains in the environment.
Subject(s)
Animal Diseases/epidemiology , Animal Diseases/microbiology , Cryptococcosis/veterinary , Cryptococcus neoformans/classification , Animal Diseases/history , Animals , Antifungal Agents/pharmacology , Cryptococcus neoformans/drug effects , Cryptococcus neoformans/genetics , Cryptococcus neoformans/isolation & purification , History, 21st Century , Microbial Sensitivity Tests , Molecular Typing , Mycological Typing Techniques , Poland/epidemiology , Polymorphism, Restriction Fragment Length , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
An innovative type of monolayer system was synthesized as a support for a bilayer lipid membrane on a gold electrode surface. The coupling of the phospholipid bilayer to the support layer is accomplished using zirconium-phosphate-carboxylate (ZPC) chemistry that provides coordinative linkage between the self-assembled, functionalized alkanethiol support monolayer and phosphatidylserine (PS). Binding the PS to the interface in this manner prevents the lipids from coming into direct contact with the metal electrode and leaves sufficient space for an aqueous supporting electrolyte to reside between the lipid and tether layers. The presence of an aqueous environment was confirmed by neutron reflectivity measurements of this interfacial system. We expect that the bilayer lipid membrane coupled to the solid support by a ZPC linkage will be almost as flexible as a natural membrane because of the labile nature of the interactions between PS and the alkanethiol layer. We anticipate that this interfacial system will provide nearly physiological conditions for membrane proteins, allowing them to retain their activity following reconstitution into the bilayer structure. In this work, the organization of the tethering monolayer was assessed using cyclic voltammetry, and corrections for dielectric permittivity values for the alkanethiol layer were calculated based on the neutron reflectometry results. Neutron reflectometry was also used to assess the organization of the tether layer and the degree of its hydration. Finally, this layer was used for the formation of a PS lipid bilayer structure by vesicle fusion and spreading, where the liposome deposition was monitored by atomic force microscopy (AFM).
