UC San Diego

Surfaces in healthcare settings are colonized by pathogens, where they persist and are transmitted to other locations. Cleaning solutions that contain bacterial spores are an alternative to current methods for cleaning healthcare surfaces. My dissertation investigates how effectively these spores germinate on surfaces used in healthcare settings, whether they inhibit the survival of methicillin-resistant Staphylococcus aureus (MRSA), and if incorporated into a 3D printed structure, how effectively they germinate and inhibit MRSA. The first chapter investigates the germination efficiency of bacterial spores on surfaces from healthcare settings. Droplets of media containing a mixture of 5 Bacillus spp. and Priestia spp. were placed on laminate or vinyl, at either ambient or elevated relative humidity, then collected over a three-day timeseries via swabbing, followed by heat shocking and plating to determine the ratio of germinated spores to the total number of spores and cells. I found that there was a low, baseline level of germination, and that only very nutrient rich media would achieve high germination rates. The type of surface and the humidity also had a significant influence, but only at later timepoints in the experiments. The second chapter focuses on the inhibition of MRSA using the spore mix from chapter 1. Droplets containing MRSA by itself or in co-culture with the spore mix were placed on laminate or vinyl and incubated at over a three-day timeseries. CFU enumeration was used to determine whether the survival of MRSA was influenced by the spore mix. Surface, humidity, and competition medium significantly influenced the survival of MRSA, but the presence of the spore mix did not inhibit MRSA survival. The third chapter uses 3D printed structures that contain a strain of B. subtilis shown to have potent anti-MRSA inhibitory activity. Using optical density readings and CFU enumeration, the germination and growth of B. subtilis out of the structure and its inhibition of MRSA was investigated. Approximately 20% of the spores in the structures germinate and move out of the structure in liquid culture to inhibit MRSA, but the structures do not completely eliminate MRSA in either liquid culture or on solid agar surfaces.