The Effects of Co-culturing on In Vitro Soil Bacterial Cultivability and the Discovery of Novel Antibiotics

Date of Award

11-2019

Degree Type

Thesis

Degree Name

Master of Science in Biomedical Sciences

First Advisor

Valerie Cadet, PhD

Second Advisor

Kimberly Baker, PhD

Third Advisor

Shafik Habal, MD

Fourth Advisor

Zhiqian Wu, PhD

Fifth Advisor

Richard White, PhD, FAHA

Abstract

Since the 1928 discovery of penicillin by Alexander Fleming, antibiotics have been an integral component of healthcare. However, within a few years of use, resistant bacterial strains were found against all utilized antibiotics which limited their therapeutic efficacy. Increased prevalence of infections caused by resistant species led the Centers for Disease Control and Prevention (CDC) to release an expanded list of antibioticresistant priority pathogens (AARPs) in 2013 and in 2017 the World Health Organization (WHO) reorganized this list yet again. Since environmental microbes remain an optimal source of antibiotics, new microbial culturing methods have been developed to increase microbial yield and biodiversity. However, these novel methods required protracted experimentation and lack experimental control due to use of in situ incubation. The purpose of this study was to develop a microbial co-culturing system that allowed diffusion of metabolites between environmental microbes in a laboratory setting with the goal of increasing microbial yield and biodiversity in comparison to traditional in vitro culturing methods. Soil samples from Radium Springs, Georgia, USA, a sparsely studied site, were co-cultured utilizing semi-solid agar inoculation, a novel culturing technique. Isolated soil microbes were then co-cultured with both multi-drug-resistant Acinetobacter baumannii (MDR-AB), methicillin-resistant Staphylococcus aureus (MRSA) to determine whether the environmental microbes possessed any antibiotic properties against these AARPs. Plate analysis of co-cultured environmental samples revealed that the novel technique yielded more microbes with diverse colony morphologies at a higher density than similar samples cultured using serial dilution, filter, and filtrate culturing methods. Plate analysis of co-cultured pathogen samples revealed several microbes with inhibitive properties. This technique could prove especially powerful in increasing bacterial yield from environmental samples and aid in antibiotic research and discovery for antibiotic-resistant priority pathogens (ARPPs)

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