Date of Award

2024

Document Type

Thesis

Degree Name

Bachelors

Department

Natural Sciences

First Advisor

Walstrom, Katherine

Area of Concentration

Biology, Chemistry

Abstract

The term microbial dysbiosis describes the interaction of microorganisms with cancerous cells, facilitating tumor progression and metastasis. Streptococcus mutans (S. mutans), a cariogenic bacterium, employs its adherence and acid-tolerant capabilities to collaborate with other bacteria in invading host cells, thereby promoting oral squamous cell carcinoma and other extra-oral diseases. In contrast, Lactobacillus casei (L. casei) exhibits probiotic properties known to assist in the treatment and prevention of intestinal cancers, fostering microbial homeostasis. When grown together, S. mutans and L. casei engage in a competitive relationship, with L. casei hindering the dominance of S. mutans. However, the intricacies of this relationship are not fully understood, leaving a knowledge gap regarding their impact on cell cycle disruptions and the underlying molecular pathways. This study investigates the effects of S. mutans and L. casei on the nematode Caenorhabditis elegans (C. elegans) through two life stage assays quantifying larval populations. Assay 1 compared worm development on S. mutans versus L. casei, using OP50 bacteria as a control, while assay 2 evaluated whether L. casei provided a protective effect. Statistical tests including an ANOVA and t-tests were used to analyze the differences between the means of the treatments. Differential interference contrast (DIC) microscopy was used to capture images of C. elegans L4 hermaphrodites and their eggs. The ANOVA results indicated that the number of eggs and L4 worms were very different between treatments in assay 1 (p-value = 0.0004 < 0.05). Worms cultivated in S. mutans exhibited a decrease in the number of offspring produced, along with delays in the development of both the eggs and offspring. A closer examination of DIC images revealed morphological defects for the eggs and the small and twisted pharynx phenotypes for the L4s. Notably, worm growth on L. casei prior to S. mutans exposure did not confer protective effects, as indicated by the comparable phenotypic and developmental delays and the consistent reductions in egg and offspring counts. The assay 2 t-test results (p-values = 0.8825 and 0.6647 > 0.05) support this point, indicating that the means of the S. mutans (OP50) and S. mutans (L. casei) treatments are very similar. The literature suggests that egg defects may be due to egg retention, a defective eggshell, or mutations having to do with the sperm or oocyte. The twisted pharynx phenotype in adults is due to a defect in actin attachment to the cuticle during growth, but it does not affect the worm’s feeding ability. The small phenotype can be explained by mutations in collagen genes and their related biosynthetic pathway products that form the cuticle. Overall, cellular pathway mutations or cuticle defects seem to be the common factor. To explain what bacterial component is causing these effects, a hypothesis was formed that an S. mutans virulence factor is affecting C. elegans genetic expression, which could alter its metabolism and/or immune response to result in the observed phenotypes. Some proposed ways to test this include a genetic screen or chemical purification of the S. mutans virulence factors. This project successfully positions C. elegans as a model organism for microbial dysbiosis research by employing the oral pathogen S. mutans. It is the first examination of this bacteria in C. elegans, laying the groundwork for identifying molecules causing cellular aberrations that may be associated with human diseases.

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