Cell-wall dyes interfere with Cryptococcus neoformans melanin deposition.

Melanization is an intrinsic characteristic of many fungal species, but details of this process are poorly understood because melanins are notoriously difficult pigments to study. While studying the binding of cell-wall dyes, Eosin Y or Uvitex, to melanized and non-melanized Cryptococcus neoformans cells we noted that melanization leads to reduced fluorescence intensity, suggesting that melanin interfered with dye binding to the cell wall. The growth of C. neoformans in melanizing conditions with either of the cell-wall dyes resulted in an increase in supernatant-associated melanin, consistent with blockage of melanin attachment to the cell wall. This effect provided the opportunity to characterize melanin released into culture supernatants. Released melanin particles appeared mostly as networked structures having dimensions consistent with previously described extracellular vesicles. Hence, dye binding to the cell wall created conditions that resembled the 'leaky melanin' phenotype described for certain cell-wall mutants. In agreement with earlier studies on fungal melanins biosynthesis, our observations are supportive of a model whereby C. neoformans melanization proceeds by the attachment of melanin nanoparticles to the cell wall through chitin, chitosan, and various glucans.

Wolbachia Endosymbionts Modify Drosophila Ovary Protein Levels in a Context-Dependent Manner.

UNLABELLED: Endosymbiosis is a unique form of interaction between organisms, with one organism dwelling inside the other. One of the most widespread endosymbionts is Wolbachia pipientis, a maternally transmitted bacterium carried by insects, crustaceans, mites, and filarial nematodes. Although candidate proteins that contribute to maternal transmission have been identified, the molecular basis for maternal Wolbachia transmission remains largely unknown. To investigate transmission-related processes in response to Wolbachia infection, ovarian proteomes were analyzed from Wolbachia-infected Drosophila melanogaster and D. simulans. Endogenous and variant host-strain combinations were investigated. Significant and differentially abundant ovarian proteins were detected, indicating substantial regulatory changes in response to Wolbachia Variant Wolbachia strains were associated with a broader impact on the ovary proteome than endogenous Wolbachia strains. The D. melanogaster ovarian environment also exhibited a higher level of diversity of proteomic responses to Wolbachia than D. simulans. Overall, many Wolbachia-responsive ovarian proteins detected in this study were consistent with expectations from the experimental literature. This suggests that context-specific changes in protein abundance contribute to Wolbachia manipulation of transmission-related mechanisms in oogenesis. IMPORTANCE: Millions of insect species naturally carry bacterial endosymbionts called Wolbachia. Wolbachia bacteria are transmitted by females to their offspring through a robust egg-loading mechanism. The molecular basis for Wolbachia transmission remains poorly understood at this time, however. This proteomic study identified specific fruit fly ovarian proteins as being upregulated or downregulated in response to Wolbachia infection. The majority of these protein responses correlated specifically with the type of host and Wolbachia strain involved. This work corroborates previously identified factors and mechanisms while also framing the broader context of ovarian manipulation by Wolbachia.