Please cite this article in press as: Falcón R, et al. High vancomycin MICs within the susceptible range in Staphylococcus aureus bacteraemia isolates are associated with increased cell wall thickness and reduced intracellular killing by human phagocytes. Int J Antimicrob Agents (2016), http://dx.doi.org/10.1016/j.ijantimicag.2016.01.014 ARTICLE IN PRESS G Model ANTAGE-4756; No. of Pages 8 International Journal of Antimicrobial Agents xxx (2016) xxx–xxx Contents lists available at ScienceDirect International Journal of Antimicrobial Agents j o ur nal ho me pag e: http://www.elsevier.com/locate/ijantimicag High vancomycin MICs within the susceptible range in Staphylococcus aureus bacteraemia isolates are associated with increased cell wall thickness and reduced intracellular killing by human phagocytes Rocío Falcón a,1 , Alba Martínez b,1 , Eliseo Albert a , Silvia Madrid a , Rosa Oltra c , Estela Giménez a , Mario Soriano d , Víctor Vinuesa a , Daniel Gozalbo b , María Luisa Gil e , David Navarro a,f,∗ a Microbiology Service, Hospital Clínico Universitario de Valencia, Fundación INCLIVA, Valencia, Spain b Department of Microbiology, Faculty of Pharmacy, University of Valencia, Valencia, Spain c Unit of Infectious Diseases, Hospital Clínico Universitario de Valencia, Fundación INCLIVA, Valencia, Spain d Centro de Investigación Príncipe Felipe, Valencia, Spain e Department of Microbiology, Faculty of Biological Sciences, University of Valencia, Valencia, Spain f Department of Microbiology, School of Medicine, University of Valencia, Valencia, Spain a r t i c l e i n f o Article history: Received 17 November 2015 Accepted 29 January 2016 Keywords: Vancomycin Minimum inhibitory concentration (MIC) Staphylococcus aureus Intracellular killing Cell wall thickness Human phagocytes a b s t r a c t Vancomycin minimum inhibitory concentrations (MICs) at the upper end of the susceptible range for Staphylococcus aureus have been associated with poor clinical outcomes of bloodstream infections. We tested the hypothesis that high vancomycin MICs in S. aureus bacteraemia isolates are associated with increased cell wall thickness and suboptimal bacterial internalisation or lysis by human phagocytes. In total, 95 isolates were evaluated. Original vancomycin MICs were determined by Etest. The susceptibility of S. aureus isolates to killing by phagocytes was assessed in a human whole blood assay. Internalisation of bacterial cells by phagocytes was investigated by flow cytometry. Cell wall thickness was evaluated by transmission electron microscopy. Genotypic analysis of S. aureus isolates was performed using a DNA microarray system. Vancomycin MICs were significantly higher (P = 0.006) in isolates that were killed suboptimally (killing index <60%) compared with those killed efficiently (killing index >70%) and tended to correlate inversely (P = 0.08) with the killing indices. Isolates in both killing groups were internalised by human neutrophils and monocytes with comparable efficiency. The cell wall was significantly thicker (P = 0.03) in isolates in the low killing group. No genotypic differences were found between the isolates in both killing groups. In summary, high vancomycin MICs in S. aureus bacteraemia isolates were associated with increased cell wall thickness and reduced intracellular killing by phagocytes. © 2016 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved. 1. Introduction Staphylococcus aureus bacteraemia is a common nosocomial and community-acquired infection associated with high morbidity and mortality [1]. Vancomycin minimum inhibitory concentrations (MICs) at the upper end of the susceptible range (MIC ≥ 1.5 mg/L) as measured by Etest both for meticillin-resistant S. aureus (MRSA) ∗ Corresponding author. Present address: Microbiology Service, Hospital Clínico Universitario de Valencia and Department of Microbiology, School of Medicine, Av. Blasco Ibá ˜ nez 17, 46010 Valencia, Spain. Tel.: +34 96 386 4657; fax: +34 96 386 4173. E-mail address: david.navarro@uv.es (D. Navarro). 1 These two authors contributed equally to this work. and meticillin-susceptible S. aureus (MSSA) isolates have been associated with poor clinical outcomes of bloodstream infections treated with vancomycin and even with -lactam antibiotics [2–9]. Based on these observations, a debate has emerged over whether the use of alternative agents, such as linezolid or daptomycin, as first-line agents or in combined therapies in this particular set- ting is clinically justified [10–13]. Other studies, however, failed to reproduce these findings [14,15]. Vancomycin treatment failures of bacteraemia episodes caused by MRSA isolates with high MICs have been postulated to occur due to the impossibility of achiev- ing drug concentrations within the therapeutic range [3]. However, this could not be demonstrated in a recent study [14]. Moreover, bacteraemia episodes caused by MSSA isolates displaying high van- comycin MICs, even those not treated with this drug, have been http://dx.doi.org/10.1016/j.ijantimicag.2016.01.014 0924-8579/© 2016 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.