Investigating the Potential for Meiosis in Shell-Building Amoebae (Arcellinida: Amoebozoa) During Encystment Using Fluorescence Microscopy and Single-Cell Transcriptomics

Although Amoebozoa are considered to be an asexual major clade, here I investigate the potential of a sexual life stage within testate amoeba (Arcellinida:Amoebozoa) due to evidence within the literature of a cryptic sexual lifestage (Mignot & Raikov, 1992). Microbial eukaryotes are often regarded as primitive with simple life cycles, and due to this misconception the diversity of microbial life cycles is widely understudied. In a single study of its kind, Raikov and Mignot (1992) found evidence of meiosis in an encysted life stage of the arcellinid Arcella vulgaris. Based on these insights, I investigate the hypothesis that Arcellinida have a sexual life stage during encystment using modern molecular and microscopy methods. I use confocal microscopy to visualize chromatin structure of vegetative and encysted cells; as well as image analysis to estimate DNA content in order to learn more about ploidy levels within these two stages. I also examine molecular data (i.e. transcriptomes) to look at the differential expression of gene families associated with or specific to meiosis in encysted cells. Here I develop a protocol for linking patterns in gene expression in Arcellinida to nuclear architecture. I find a difference in nuclear architecture between encysted and non-encysted cells, and characterize a set of candidate genes involved in regulating encystment of Hyalosphenia papilio cells.