Sylvia Kwakye- skwakye1@swarthmore.edu
Swarthmore College
500 College Avenue
Swarthmore PA 19081-1397
Popular version of paper 3aED2
Presented Wednesday morning, June 18, 1997
133rd ASA Meeting, State College, PA
Embargoed until June 18, 1997
Regular and effective sterilization of equipment and medical implants is extremely important to hospitals and to manufacturers of medical implants, who spend enormous amounts of money on sterilization procedures. In spite of these precautions, an estimated two million people become infected while in hospitals in the United States alone every year. This is not because the hospitals are not doing a good job of sterilizing equipment but rather because of the evolution of new and hardier strains of bacteria.
Most sterilization technologies today have disadvantages associated with their use. Ethylene oxide, for instance, which is the current standard for gas sterilization, is hazardous to human health and harmful to the environment. Peracetic acid, vapor phase hydrogen peroxide, hydrogen peroxide plasma, and formaldehyde liquids are costly and difficult to use.
In fact, the use of chemicals in sterilization is not a good idea in general, since they contribute directly to the evolution of resistant bacterial strains. The chemicals kill off most of the bacteria but those that survive, because they are less susceptible to the chemical, multiply in the absence of competition. Thus, the effectiveness of chemical sterilization gradually reduces with time.
Non-chemical techniques such as steam and radiation sterilization present issues of material compatibility, and cannot be used in vivo. Also, steam requires a relatively high input of energy to generate the high temperatures and pressures needed for effective sterilization.
With these problems in mind, our research group is involved in a project to assess the feasibility of applying acoustic cavitation to intracorporeal sterilization of medical implants and equipment.
Inertial acoustic cavitation is the formal term for the phenomenon of rapid bubble growth and violent collapse induced by ultrasound. It is known to be responsible for damage to biological cells, especially blood cells, in vivo and in vitro. The temperatures and pressures generated during bubble collapse can produce direct and chemically-mediated effects on nearby cells. Furthermore, fluid microjets driven by asymmetrical bubble collapse may have enough force to puncture cellular membranes or bacterial walls. Clearly, there is some damage to cells during cavitation but the mechanism of interaction is not clearly understood. This area of interaction is the focus of our research.
Bioluminescence is a very sensitive and convenient tool that provides a means to quantify repair mechanisms employed by bacteria in response to specific kinds of damage. In naturally luminous bacteria, light is produced by proteins encoded by lux genes. In our research, we used genetic cloning techniques to place these lux genes into normally non-luminous (non-pathogenic) strains of E. coli bacteria. These strains were constructed to produce light in response to various types of stresses such as heat shock, (or protein damage) membrane damage, oxidation and DNA damage. The addition of a small volume of micron-sized bubbles to our bacterial assays before exposure to ultrasound provides nucleation sites required for cavitation. We control the extent and intensity of cavitation by varying acoustic parameters such as the pulse width or the amplitude of the 1 MHz ultrasonic wave used in our experiments. Measurement of luminescence produced per viable cell enables us to describe the mechanism, and degree of damage done by the timed ultrasound exposures.
The exposures kill about 25% of bacterial cells on average under present laboratory conditions. Too much killing might compromise our academic interest in producing a qualitative and quantitative assessment of sound-cell interactions. Dead cells do not emit light. With a greater understanding of the killing process, we hope to achieve close to 100% killing with each exposure.
There are two clear advantages associated with this method. The energy input for ultrasound production is much less than for methods requiring heat. Also, since cavitation is a random process, killing depends on the proximity of a bacterium to a bubble. So, if for example, 5% of the bacteria survive an exposure, they will be a random 5% of the bacterial population, not the hardiest or most resistant 5%.
The success of this project has implications not only for the medical field but also, for other applications in which sterilization is needed. In a water purification system for instance, naturally occurring microbubbles in the water can be excited by ultrasound to cavitate and reduce bacterial populations without the use of any externally applied chemicals whatsoever.
With more research and a better understanding of the interaction between cavitating bubbles and bacteria, we hope to propose an acoustic solution to the problem of selection in sterilization.
Acknowledgement I would like to acknowledge Professors Carr Everbach and Amy Vollmer, supervisors of the project and Matt Halpern, student research partner.