Research Article Effects of Chlorophyll-Derived Efflux Pump Inhibitor Pheophorbide a and Pyropheophorbide a on Growth and Macrolide Antibiotic Resistance of Indicator and Anaerobic Swine Manure Bacteria Mareike Kraatz, 1 Terence R. Whitehead, 1 Michael A. Cotta, 1 Mark A. Berhow, 1 and Mark A. Rasmussen 2 1 National Center for Agricultural Utilization Research, ARS, USDA, 1815 N. University Street, Peoria, IL 61604, USA 2 Leopold Center for Sustainable Agriculture, 209 Curtiss Hall, Iowa State University, Ames, IA 50011, USA Correspondence should be addressed to Terence R. Whitehead; terry.whitehead@ars.usda.gov Received 21 October 2013; Revised 12 December 2013; Accepted 19 December 2013; Published 11 February 2014 Academic Editor: Federico Pea Copyright © 2014 Mareike Kraatz et al. his is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Natural plant compounds, such as the chlorophyll a catabolites pheophorbide a (php) and pyropheophorbide a (pyp), are potentially active in the gastrointestinal tracts and manure of livestock as antimicrobial resistance-modifying agents through inhibition of bacterial elux pumps. To investigate whether php, a known elux pump inhibitor, and pyp inluence bacterial resistance, we determined their long-term efects on the MICs of erythromycin for reference strains of clinically relevant indicator bacteria with macrolide or multidrug resistance elux pumps. Pyp reduced the inal MIC endpoint for Staphylococcus (S.) aureus and Escherichia (E.) coli by up to 1536 and 1024 g erythromycin mL −1 or 1.4- and 1.2-fold, respectively. Estimation of growth parameters of S. aureus revealed that pyp exerted an intrinsic inhibitory efect under anaerobic conditions and was synergistically active, thereby potentiating the efect of erythromycin and partially reversing high-level erythromycin resistance. Anaerobe colony counts of total and erythromycin-resistant bacteria from stored swine manure samples tended to be lower in the presence of pyp. Tylosin, php, and pyp were not detectable by HPLC in the manure or medium. his is the irst study showing that pyp afects growth and the level of sensitivity to erythromycin of S. aureus, E. coli, and anaerobic manure bacteria. 1. Introduction Agricultural antimicrobial drug use is regarded a major driver of one of today’s foremost global public health challenges: more frequent clinical antimicrobial treatment failures due to resistant microorganisms [1–4]. In the U.S. swine and other livestock production, much of the use of antimicrobials is nontherapeutic and/or occurs in the form of free-choice medicated feeds and water [1, 2, 5]. his results in exposures of the animals’ gastrointestinal tract and waste microbiota to inconsistent, oten sublethal or subinhibitory concentrations [1, 6]. As even ultralow (≪MIC) antimicrobial concentrations can confer a selective pressure towards the persistence of resistance in microbial communities [7–12], induction of gut and waste microbial resistance is an inevitable collateral efect of oral antimicrobials in animal agriculture [13–16]. More than 335 million tons (dry weight) of manure, a valuable fertilizer, are produced by U.S. agriculture per year [17]. Soil amendment with manure presents a sig- niicant route of transmission of antimicrobial resistance from livestock bacteria to human clinical pathogens [4, 18– 24]. Of greatest concern in this context is the increasing prevalence of multidrug resistance (MDR), especially in Gram-positive pathogens, such as Staphylococcus (S.) aureus, Streptococcus pneumonia, and enterococci [25, 26]. MDR is frequently caused by bacterial elux pumps that primarily confer broader, compound nonspeciic functions unrelated to antimicrobials and are ubiquitous among bacteria [22, 25, 27–29]. MDR is a baseline resistance for the emergence of further resistance mechanisms, and, due to its physiological determination, it naturally persists [29–32]. Hindawi Publishing Corporation International Journal of Antibiotics Volume 2014, Article ID 185068, 14 pages http://dx.doi.org/10.1155/2014/185068