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System for Monitoring of Mutation-in-Progress i ABSTRACT In nature, evolution progresses to assure the survival of the strongest of a species. With each mutation, those characteristics that best assure the survival of the species are propagated through successive, on-going mutations. As there is no known method to monitor such mutation-in-progress, the objective of this research was to devise a technique to monitor the progression of mutation. Pathogens are known to survive (mutate to gain resistance) in an antibioticcontaining environment. One such pathogen, staphylococcus aureus is known to cause diseases ranging from superficial dermatological lesions to systemic and life threatening infections such as endocarditis. One particular strain of staphylococcus aureus methicillin resistant Staphylococcus aureus (MRSA) is known to be resistant to almost all antibiotics. A pilot study established that it is feasible to monitor the growth of staphylococcus aureus using overnight cultures as well as direct culture of nasal swab through visible spectroscopy. Using such spectroscopic technique, it was possible to discriminate between methicillin susceptible and resistant strains of Staphylococcus aureus. Based on these results, a novel system coined the System for Monitoring Mutation-in-Progress (SMMIP) was devised. SMMIP allows nutrients to be replenished and extracted to allow for long term tracking. In this study, the use of SMMIP to monitor the mutation-in-progress is demonstrated through the adaptation of ciprofloxacin resistance, a fluoroquinolone antibiotic, in Staphylococcus aureus. SMMIP was successful in automatically monitoring the growth of methicillin susceptible Staphylococcus aureus (MSSA) ATCC 29213 in Mannitol Salt Broth (MSB) containing ciprofloxacin. Resistant mutants in MSB containing 0.5 μg/ml of ciprofloxacin emerged on Days 6, 4, and 7 from samples containing initial inocula of 6.0 x 10 CFU/ml, 1.3 x 10 CFU/ml, and 1.3 x 10 CFU/ml respectively. The ciprofloxacin mutants were detected by monitoring wavelength shifts of the acidogenic profiles of the growth medium. The SMMIP measures the growth of

A system for monitoring mutation-in-progress : a study of adaptive mutation to Ciprofloxacin resistance in Staphylococcus aureus