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Molekulární diagnostika fungálních patogenů z klinických vzorků
CIMICKÁ, Jana
This study is focused on optimization and use of molecular methods in mycological diagnostics. The patient's life often depends on early diagnosis, which current culture methods cannot provide. The solution is based on pan-fungal primers of ITS region and next generation sequencing (NGS). These methods were used for fungal detection from FFPE tissue samples from patients with ulcerative colitis (UC), Crohn disease (CD) and 10 controls without chronic inflammatory changes in the intestine. Results of this study have three levels. Firstly, the process of optimization highlights the importance of sterile DNA isolation in a laminar box which can prevent contamination by Malassezia spp. Secondly, raw data showed balanced rate of Ascomycota and Basidiomycota in CD samples similar to negative controls, while UC samples indicated higher representation of Ascomycota than other groups. Thirdly, detailed data showed fungal genera Malassezia, Cladosporium and Toninia in all groups, while genus Candida was found only in UC samples and genera Engyodontium and Ramularia were found in CD samples. In conclusion, mycobiome plays a role in inflammatory bowel diseases; however, its compartments are still questionable. The optimization of pan-fungal nested PCR and NGS helped with introduction of mycological diagnostics from FFPE samples to Bioptická laboratoř s.r.o. and is used for differential diagnostics.
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The mycobiomes of vegetative organs of the dominant Mediterranean seagrass Posidonia oceanica (Posidoniaceae, Alismatales)
Soperová, Beáta ; Vohník, Martin (advisor) ; Kohout, Petr (referee)
Posidonia oceanica is a dominant seagrass of the Mediterranean Sea and its mycobiome has been increasingly studied, especially in recent years. Dozens of fungi associated with roots, rhizomes or leaves have been already described, but studies on the mycobiome of leaf sheaths are completely absent. One of the root-associated mycobionts is Posidoniomyces atricolor, which has not yet been isolated from any other organ. This thesis attempted to describe the mycobiome of vegetative organs of P. oceanica using two different methodological approaches - microscopic observation and molecular analyses. Microscopic observation confirmed the presence of fungal colonization in all vegetative organs except leaves. The conclusions from the microscopic part were subsequently verified by isolating and identifying several different endomycobionts, including P. atricolor, which has been also isolated for the first time from rhizomes and leaf sheaths. Leaf sheath mycobiome was then described for the first time ever.
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