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All Subjects: Protistology
Creators: Gile, Gillian

Defining the Limitations of Illumina Amplicon Sequencing as it Relates to Community Diversity Analysis

Description

The termite Coptotermes gestroi is a small subterranean termite originating from Southeast Asia. The hindgut of C. gestroi contains five distinct species of parabasalid: Pseudotrichonympha leei, two undescribed species from the genus Holomastigotoides, and two undescribed species from the genus Cononympha. This study investigates the protist symbionts in C. gestroi and the relationship between their relative abundance as inferred by Illumina sequence reads and the directly observed abundances for each protist genus. Illumina amplicon sequencing as a means of DNA analysis is a proven method for identification and diversity analysis, although the specific ratios of sequence reads to cell abundance in protists is not well known. In this study, protist communities were observed under light microscopy; cells were counted under hemocytometer and characterized at the molecular level using Illumina amplicon sequencing. When comparing sequence read abundances to cell abundances, some general trends were found in both analysis methods. Cononmypha repeatedly formed the majority of the community, while Holomastigotoides and Pseudotrichonympha were responsible for a smaller yet similar portion of the population. Cell counts and sequence reads were also compared using an assumed linear model, with R2 values generated to quantify the relationship between both. The results suggest that Illumina sequencing can be used to obtain rough estimates of community diversity, but the high variability within the data suggest that the read abundances should be treated with caution.

ContributorsAvilucea, Erin L. (Author) / Gile, Gillian (Thesis director) / DeMartini, Francesca (Committee member) / Taerum, Stephen Joshua (Committee member) / School of Mathematical and Natural Sciences (Contributor) / Barrett, The Honors College (Contributor)

Created2018-05

Distribution and relative abundance of five protist genera within the Zootermopsis nevadensis nuttingi hindgut

Description

The termite Zootermopsis nevadensis nuttingi, which is located in coastal forests of the western United States, plays an important ecological role in the breakdown and digestion of wood. Vital to this role are symbiotic protists residing in the termite's hindgut. Five protist genera of varying size and morphology make up this gut community. Despite years of study on this termite species, little was known about the spatial organization of the protist community within Zootermopsis nevadensis nuttingi. To resolve this issue, a study was conducted in which the distribution of protist genera among gut segments was observed and elucidated. This was done by separating hindgut segments, then counting the protists using a hemocytometer at a magnification of 200x. 60 segments from 20 termites were examined, and the total number of protists counted was 69,560. Images were also taken using a scanning electron microscope. Statistically significant, distinct distribution patterns were found for Trichonympha, Trichomitopsis and Streblomastix, while the small genera of Hexamastix and Tricercomitus appeared to have no special distribution. Trichomitopsis was more abundant in the posterior hindgut, Streblomastix was more abundant anteriorly, while the distribution of Trichonympha varied by colony. Hexamastix and Tricercomitus make up a large majority of the protists observed in any segment, followed by Streblomastix, Trichomitopsis and Trichonympha. Understanding the distribution of different protists within the hindgut may improve our understanding of the ecological relationships among protists as well as their individual roles in lignocellulose digestion, contributing to a better understanding of the hindgut system as a whole.

ContributorsPiarowski, Christina Marie (Author) / Gile, Gillian (Thesis director) / DeMartini, Francesca (Committee member) / Taerum, Stephen (Committee member) / School of International Letters and Cultures (Contributor) / School of Human Evolution and Social Change (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)

Created2018-05

Plasmodium Cost of Resistance and Life Stage Development within the Mosquito Vector

Description

Hundreds of thousands of people die annually from malaria; a protozoan of the genus Plasmodium is responsible for this mortality. The Plasmodium parasite undergoes several life stages within the mosquito vector, the transition between which require passage across the lumen of the mosquito midgut. It has been observed that in about 15% of parasites that develop ookinetes in the mosquito abdomen, sporozoites never develop in the salivary glands, indicating that passage across the midgut lumen is a significant barrier in parasite development (Gamage-Mendis et al., 1993). We aim to investigate a possible correlation between passage through the midgut lumen and drug-resistance trends in Plasmodium falciparum parasites. This study contains a total of 1024 Anopheles mosquitoes: 187 Anopheles gambiae and 837 Anopheles funestus samples collected in high malaria transmission areas of Mozambique between March and June of 2016. Sanger sequencing will be used to determine the prevalence of known resistance alleles for anti-malarial drugs: chloroquine resistance transporter (pfcrt), multidrug resistance (pfmdr1) gene, dihydropteroate synthase (pfdhps) and dihydrofolate reductase (pfdhfr). We compare prevalence of resistance between abdomen and head/thorax in order to determine whether drug resistant parasites are disproportionately hindered during their passage through the midgut lumen. A statistically significant difference between resistance alleles in the two studied body sections supports the efficacy of new anti-malarial gene surveillance strategies in areas of high malaria transmission.

ContributorsPhillips, Keeley Isabella (Author) / Huijben, Silvie (Thesis director) / Gile, Gillian (Committee member) / Young, Steven (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)

Created2021-05