Rosa, C. P. | Search bioRxiv

bioRxiv2020

Faster Cryptococcus melanization increases virulence in experimental and human cryptococcosis

Cryptococcus spp. are important human pathogens responsible for about 180,000 deaths per year. Studying their virulence attributes can lead to better cryptococcosis prevention and treatment. In this work, we systematically investigated virulence attributes of Cryptococcus spp. clinical isolates and correlated them with patient data. We collected 66 C. neoformans and 19 C. gattii isolates from Brazilian patients and analyzed multiple phenotypes related with their capsule, production of laccase, melanin and extracellular vesicles. We also tested their virulence in Galleria mellonella and ability to evade macrophage LC3-associated phagocytosis (LAP). All phenotypes analyzed varied widely among the isolates, but C. neoformans isolates tended to melanize faster and more intensely and produce thinner capsules in comparison with C. gattii. We also observed correlations that match previous studies, such as that between secreted laccase - but not total melanin production - and disease outcome in patients. The most striking results, though, came from our measurements of Cryptococcus colony melanization kinetics, which followed a sigmoidal curve for most isolates. Faster melanization correlated positively with LAP evasion, virulence in G. mellonella and worse prognosis in humans. These results suggest that the speed of melanization, more than the total amount of melanin Cryptococcus spp. produces, is crucial for virulence. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=162 SRC="FIGDIR/small/222794v2_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@41c48borg.highwire.dtl.DTLVardef@1ce8e1eorg.highwire.dtl.DTLVardef@1d4568corg.highwire.dtl.DTLVardef@17e2003_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2018

Opsonin-free, real-time imaging of Cryptococcus neoformans capsule during budding

Cryptococcus neoformans is a unicellular fungal pathogen that causes meningoencephalitis, killing hundreds of thousands of patients each year. Its most distinctive characteristic is a polysaccharide capsule that envelops the whole cell. It is the major virulence attribute and the antigen for serologic diagnosis. We have developed a method for easy observation of the capsule and its growth dynamics using the cell-separation reagent Percoll and differential interference contrast (DIC) microscopy. Percoll suspension is far less disruptive of cell physiology than methods relying on antibody binding to the capsule, and measurements made with it are equivalent with India ink. Time-lapse microscopy observations using this method suggest that during budding, a dividing cell can regulate whether the capsule polysaccharide it produces is deposited on the capsule of the bud or on its own. This observation has important implications for our understanding of the C. neoformans capsule induction process during budding.\n\nList of abbreviations and acronyms

microbiology↗