Degree Type

Dissertation

Date of Award

2001

Degree Name

Doctor of Philosophy

Department

Animal Science

First Advisor

Tim S. Stahly

Abstract

An initial study was designed validate an in vitro antimicrobial assay and evaluate the activity of natural antimicrobial animal proteins/peptides. A radial diffusion assay was utilized. By regressing known concentrations of tested compound against their respective clearance zones, an equation was developed to determine the minimum inhibitory concentrations (MIC) for polymyxin B (control antibiotic) which were 0.76, 0.76, and 0.90 mug/mL for Escherichia coli, Escherichia coli (nalidixic acid-resistant), and Staphylococcus aureus, respectively. The intra- and inter-assay variations were 0.18 and 0.2%, respectively. Lactoferrin, lactoferricin B, hen egg lysozyme, and alpha-lactalbumin LDT2 were determined in vitro to kill bacteria. Each of the tested proteins/peptides was active against a nalidixic acid-resistant strain of E. coli. The antimicrobial activity of each protein/peptide in animal digesta fluid was 130 to 300% greater than that in the acetic acid media. Lactoferrin activity was decreased (P < .07) when exposed to zinc, iron, magnesium, calcium, sodium, and potassium. Lysozyme had an increase (P < .09) in antimicrobial activity when exposed to zinc, magnesium, and calcium. A decrease in pH from 6.5 to 2 resulted in a loss of antimicrobial activity of 65% for lactoferrin, lactoferricin, and lysozyme. Therefore, the enhanced antimicrobial activity of the proteins/peptides in the presence of the digesta fluid does not seem to be affected by pH 44 or mineral concentration.;A second study was performed to evaluate the efficacy of orally ingested natural proteins/peptides contained in digesta contents to kill antibiotic-susceptible and -resistant bacteria, in vitro. The seven treatments consisted of a control (no antimicrobial agent supplemented), lactoferrin, lactoferricin, lysozyme, neutrophil peptides, purothionin, or polymyxin B (antibiotic). Weanling Sprague-Dawley rats were self-fed their respective experimental diets for 3 days. Following oral ingestion, each protein/peptide exhibited antimicrobial activity in the small and large intestine digesta (P < .06) and against Gramnegative and Gram-positive bacteria (P < .11) (both antibiotic-susceptible and -resistant strains, P < .10). The antibiotic polymyxin B also effectively killed in vivo each bacterium except antibiotic-resistant Staphylococcus aureus, while the control diet exhibited no antimicrobial activity. Based on these results, ingested natural proteins/peptides retain antimicrobial activity in monogastric animals throughout the gastrointestinal tract and effectively kill both antibiotic-susceptible and -resistant bacteria.

DOI

https://doi.org/10.31274/rtd-180813-12510

Publisher

Digital Repository @ Iowa State University, http://lib.dr.iastate.edu

Copyright Owner

Laura Lee Greiner

Language

en

Proquest ID

AAI3048704

File Format

application/pdf

File Size

110 pages

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