Document Type

Honors Project

First Advisor

Stephen Baron

Degree Award Date

Spring 5-2025

Keywords

Tetracycline, antibiotic resistance, Escherichia coli, tetB, tetA, tetM, resistance gene, tetracycline resistance

Disciplines

Molecular Biology

Abstract

The purpose of this study was to determine the prevalence rates of the tetracycline resistance genes tetA, tetB, and tetM in environmental Escherichia coli isolates. Samples were from both an urban and an agricultural waterway were filtered and incubated with Coliscan™ medium to differentiate for fecal coliforms. The resulting colonies were streaked onto EMB and then TSA plates to ensure isolation and presence of E. coli. All media used were supplemented with 10 µg/mL tetracycline in order to maintain selective pressure on the isolates. DNA from the isolates was then extracted and PCR was run using tetA, tetB, and tetM primers to see what genes were present in each isolate, using conditions from a published study. Because multiple non-specific products were observed, we modified the procedure by increasing the annealing temperature from 54°C to 56°C, shortening cycling times, and lowering the concentration of Mg+2. These changes greatly improved specificity with the tetA and tetB, but not the tetM primers. Overall, results tentatively suggested that tetA occurred in isolates from urban and agricultural areas at prevalence rates of 100% and 33.3%, respectively and possibly in a lab strain of E. coli, K12. Results with tetM were too difficult to interpret due to non-specific product formation, but tetB was possibly present in at least one of the environmental strains and in E. coli K12.

Force Open Access

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