UBC Theses and Dissertations
New ichthyosporean protists and their biology as inferred from molecular genetic and light and electron microscopic study Marshall, Wyth
Precipitated by unexpected discoveries, this thesis is dedicated to the study of the biology of ichthyosporeans. While searching for undiscovered opisthokonts living osmotrophically in marine invertebrate digestive tracts, I established 177 cultures of ichthyosporeans. Ichthyosporeans are one of six understudied unicellular lineages related to the multicellular animals and fungi. Ichthyosporeans are comprised of approximately 30 genera but, prior to this thesis, only four genera had been cultivated. I identified and described three new genera and six species using microscopy and molecular phylogenetic techniques. Two, which I named Abeoforma whisleri and Pirum gemmata, were most closely related to divergent clone sequences and had no known relatives. My other four species, each isolated between eight and 126 times, were related to single isolates Sphaeroforma arctica and Pseudoperkisus tapetis, also found in marine invertebrates. I described one as the new genus and species Creolimax fragrantissima because of its amoeboid reproductive and dispersal stage and fragrance. The other three species were closely related and morphologically indistinguishable. To delimit species, I sequenced three loci from multiple isolates and applied a genealogical concordance species concept. Once delimited, I was able to describe variations in life cycle, morphology and a possible difference in host preference. Rather than adapt cytological techniques to describe the life cycle of S. tapetis, the most abundant species, I used population genetics to work in reverse. Absence of heterozygotes provided evidence for haploidy. Phylogenetic incongruence and a lack of support for linkage between two loci signified a history of recombination consistent with a sexual cycle. I described the ultrastructure of five species using high-pressure frozen cells from healthy, luxuriantly growing cultures. The quality of preservation allowed me to describe features that were new to ichthyosporeans, such as, spindle pole bodies and tubular extensions of the cell that penetrated the cell wall. These features were found in both clades and may have been present in the ancestral ichthyosporean. By combining genetic evidence for sex, observations of asexual reproduction in culture and collection frequencies I proposed a life cycle involving infrequent recombination within a predominantly asexual organism that infected invertebrate hosts indiscriminately via asexual endospores.
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