Civil Engineering

 

Date of this Version

Spring 5-2016

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A THESIS Presented to the Faculty of The Graduate College at the University of Nebraska In Partial Fulfillment of Requirements For the Degree of Master of Science, Major: Civil Engineering, Under the Supervision of Professor Xu Li. Lincoln, Nebraska: May, 2016

Copyright © 2016 Rachel E. Levine

Abstract

Certain microbes can transform antibiotics in the environment. However, little is known about the identity of these microbes and their antibiotic biotransformation processes. The objectives of this study were to (1) isolate bacterial strains capable of transforming antibiotics, (2) determine the biotransformation kinetics of antibiotics, (3) characterize the effects of background carbons on the biotransformation kinetics, and (4) identify biotransformation products under various environmental conditions. Sulfadiazine (SDZ) was used as the model antibiotic in this study due to its frequent occurrence in livestock wastes. Surface soil from a cattle feedlot was collected to enrich potential SDZ degrading bacteria. A mixed culture was obtained after several cycles of enrichment in a mineral solution containing 10 mg/L SDZ as the sole carbon and energy source. Despite repeated efforts, no single SDZ degrading strain could be isolated from the mixed culture. 16S rRNA gene sequence analysis showed that the culture consisted primarily of two major bacterial species, Brevibacterium epidermidis and Castellaniella denitrificans. The degradation kinetics of SDZ by the mixed culture could be described using a mirrored logistic function, with a biotransformation rate measured to be at 4.86 mg∙L-1∙d-1. Three types of background carbons were tested: diluted R2A medium, glucose, and humic acid. The mixed culture had the fastest and slowest SDZ biotransformation rates when diluted R2A and humic aicd were used as the background carbon, respectively, at concentrations equivalent to SDZ on a carbon basis. The mixed culture could also degrade other sulfonamide compounds such as sulfamethazine and sulfamerazine, at transformation rates slower than that of SDZ, but could not degrade sulfathiazole. Using liquid chromatography tandem mass spectrometry, we identified 2-aminopyrimidine (2-AP) as a major biotransformation product of SDZ in the absence and presence of the background carbons tested. Another biotransformation product detected was confirmed to not be 4-aminobenzenesulfonate, the remaining structure after the cleavage of 2-AP from SDZ. This work presents a comprehensive study of microbial biotransformation of SDZ under various environmental conditions.

Adviser: Xu Li

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