Methicillin-Resistant Staphylococcus aureus on Public Transportation Vehicles: Sampler Performance, Prevalence, and Epidemiology DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Jonathan Kyle Lutz, M.P.H. Graduate Program in Public Health The Ohio State University 2011 Dissertation Committee: J. Mac Crawford, Advisor Armando E.S. Hoet Jiyoung Lee John R. Wilkins III 1 Copyright by Jonathan Kyle Lutz 2011 i Abstract Methicillin-resistant Staphylococcus aureus (MRSA) is an important public health threat, with significant disease burden. Of particular concern are the growing number of cases that have origins in the community (CA-MRSA); these infections often affect individuals without underlying risk factors for disease. Because MRSA colonizes a large number of people, can be easily spread by fomites, and survives on surfaces for months, the environment is an important element in MRSA transmission. Public transportation vehicles, which have high levels of crowding and significant hand-to-fomite contact, may represent an important site for MRSA transmission. This study determined the background prevalence of MRSA on buses from a large transportation agency. Prior to vehicle sampling, a laboratory-based study was conducted to evaluate the performance of four sampling methods for recovery of S. aureus from a stainless steel surface. Results indicate that overall, the electrostatic wipe resulted in the greatest performance across all inoculating concentrations ([relative sampling efficiency=SE; 2 analytical sensitivity=Sn] SE48h=17.8%; Sn48h=7 CFU/100cm ). However, the swab sampler performed well when corrected for area actually sampled (SE48h=24.3%; 2 Sn48h=76 CFU/100cm ), but is greatly limited by the surface area which can be effectively sampled. Among the contact-based methods, the roller sampler outperformed ii 2 the contact plate (roller: SE48h=9.9%; Sn48h=17 CFU/100cm ; contact plate: 2 SE48h=0.04%; Sn48h=1,412 CFU/100cm ), and both contact samplers performed better at higher inoculating concentrations. Statistically significant differences were observed between the samplers only at the lowest two and highest inoculation concentrations (pconc 1= 0.009; pconc 2= 0.007; pconc 5= 0.029). Overall, the wipe resulted in the greatest number of replications that resulted in positive growth, with 73.8% at 24 hours, and 90.5% at 48 hours. Using this laboratory data, a field-based surveillance study was conducted to determine whether S. aureus (both methicillin-susceptible Staphylococcus aureus [MSSA] and MRSA) could be isolated from bus surfaces. A total of 237 samples were collected from 40 buses from a major urban transportation agency. Sixty-eight percent of buses were contaminated with S. aureus (both MSSA and MRSA), and 63% with MRSA. In total, 17% of the 237 samples were positive for S. aureus (both MSSA and MRSA) and 15% for MRSA. Categorical analysis indicated that at the sample level, sampling date (pS. aureus, July= 0.075) and sample location (pMRSA= 0.000; pS. aureus= 0.000) on a vehicle were associated with vehicle contamination, At the bus level, no factors were significantly associated with vehicle contamination, but ridership (pS. aureus= 0.088) and sampling date (pMRSA, July= 0.074; pS. aureus, July= 0.074) indicated potentially important associations. Finally, the antibiotic resistance profiles and molecular characteristics of positive bus isolates were evaluated. Multiple resistance patterns were observed, with high levels of multi-class resistance. Sixty-five percent (65%) of isolates were of the community- iii associated staphylococcal cassette chromosome mec (SCCmec) type IV, and 20% were healthcare-associated SCCmec type II. Multiple pulsed-field gel electrophoresis patterns were noted, with the majority being CA-types USA300/400; the HA-type USA100 was also found. The hospital-associated strain types were resistant to more classes of antibiotics, although multi-class resistance was also observed among the community- associated strains. High levels of clindamycin resistance were noted across all study isolates, for both MSSA and MRSA. Clonality results demonstrated that buses may be effective mixing vessels for MRSA strains, as multiple unique strains were noted in some study vehicles. iv Dedication This document is dedicated to my family; especially my parents, who never discouraged me from unceasingly asking ―but why?‖ to all life’s mysteries. And to my beloved Darlene, who is my anchor, my calming force, and who reminded me to laugh every day. Finally, this process would not have been possible without my brother, my friend, and my mentor – Eric. v Acknowledgments First and foremost, this project would not have been possible without the tireless efforts of my advisor, Dr. Mac Crawford. Mac provided a rich graduate experience, acting as mentor, counsel, and friend; the value of his academic guidance is only surpassed by the kindness of his heart. Also, I am extremely grateful to the entire esteemed committee: Dr. Armando Hoet, who oversaw all MRSA field activities and first ―introduced‖ me to MRSA; Dr. Jiyoung Lee, who provided expertise for the sampler evaluation, and who always pushed me to examine the practical relevance any research; and Dr. Wilkins, for instilling in me a sense of epidemiological inquisitiveness. I would also like to extend gratitude to the transportation agency for their participation in the study. Special thanks to the employees directly involved in coordinating this work. I am also greatly indebted to the numerous volunteers that helped collect and analyze samples, including Dr. Rocio Hoet, Dr. Joany Van Balen, Jade Braman, Melissa Mikolaj, ―View‖ Wannasawat Ratphitagsanti, Chongtao Ge, Sophia Dailey, Colleen Shockling-Dent, and Debbie Lutz; and the review of Dr. Eric Lutz. Last – but certainly not least – I would like to thank Shasha Bai, whose statistical guidance prevented me from too many gray hairs during the process. vi Vita June 1997 .......................................................St. Charles Preparatory School, Columbus, Ohio 1999 – 2000....................................................Project Manager/Scientist, BioGard Environmental Services, Phoenix, Arizona 2000 – 2002....................................................Project Scientist, Environmental Management Group, Phoenix, Arizona June 2001 ......................................................B.S. Zoology, The Ohio State University 2003 – 2005....................................................Project Manager/Staff Industrial Hygienist, PGD Environmental Group, Cleveland, Ohio 2005 – 2006....................................................Project Manager, Anderson Group International, Bakersfield, California 2006 – 2009....................................................Operations Manager/Industrial Hygienist, Auburn Environmental, Cleveland, Ohio May 2007 ......................................................M.P.H., Cleveland State University vii 2008 to present ..............................................Graduate Teaching Associate, Department of Environmental Health Sciences, College of Public Health, The Ohio State University 2011................................................................President/Industrial Hygienist, Auburn Environmental, Cleveland, Ohio Publications Lutz, J. K., & Lee, J. (2011). Prevalence and antimicrobial-resistance of Pseudomonas aeruginosa in swimming pools and hot tubs. International Journal of Environmental Research and Public Health, 8(2), 554-564. Fields of Study Major Field: Public Health viii Table of Contents Abstract ............................................................................................................................... ii Dedication ........................................................................................................................... v Acknowledgments.............................................................................................................. vi Vita .................................................................................................................................... vii List of Tables ..................................................................................................................... xi List of Figures .................................................................................................................. xiii Chapter 1: Introduction ....................................................................................................... 1 Chapter 2: Background ..................................................................................................... 15 Chapter 3: Performance of Surface Sampling Methods for the Recovery of S. aureus ... 48 Chapter 4: Methicillin-Resistant Staphylococcus aureus Contamination on Public Transportation Vehicle Surfaces ....................................................................................... 85 Chapter 5: Molecular Epidemiology of Methicillin-Resistant Staphylococcus aureus Contamination on Public Transportation Vehicle Surfaces ............................................ 120 Chapter 6: Synthesis and Discussion .............................................................................. 147 References ....................................................................................................................... 167 ix Appendix A: Efficacy of Modified Cleaning Techniques for the Reduction of