Molecular biology, genetics and biotechnology Proteomic changes in Bacteroides fragilis exposed to subinhibitory concentration of piperacillin/tazobactam Luciano de Castro Veloso a , Kênia Valéria dos Santos c , Hélida Monteiro de Andrade b , Simone da Fonseca Pires b , Simone Gonçalves dos Santos a , Mariana Jesus Vaz Trindade a , Luiz de Macêdo de Farias a , Maria Auxiliadora Roque de Carvalho a, * a Universidade Federal de Minas Gerais, Instituto de Ciências Biológicas, C4-204 Departamento de Microbiologia, Laboratório de Microbiologia Oral e Anaeróbios, Av. Antônio Carlos, 6627 Pampulha, CEP-31270-901 Belo Horizonte, Minas Gerais, Brazil b Universidade Federal de Minas Gerais, Instituto de Ciências Biológicas, L4-161, Departamento de Parasitologia, Laboratório de Leishmanioses, Av. Antônio Carlos, 6627 Pampulha, CEP-31270-901 Belo Horizonte, Minas Gerais, Brazil c Universidade Federal do Espírito Santo, Centro de Ciências da Saúde, Departamento de Patologia, Av. Marechal Campos,1468 Maruípe, CEP-29040-090 Vitória, Espírito Santo, Brazil article info Article history: Received 14 November 2012 Accepted 12 April 2013 Available online 23 April 2013 Keywords: Bacteroides fragilis Piperacillin/tazobactam Subinhibitory concentration Proteome Two-dimensional electrophoresis abstract Bacteroides fragilis is the anaerobe most frequently isolated from clinical specimens and piperacillin/ tazobactam is among the drugs that can be used to treat polymicrobial infections in which this bacteria is often involved. During antibiotic therapy, inhibitory concentrations of antibiotics are always followed by subinhibitory concentrations which can generate phenotypic changes in bacteria. So, in this study we aimed to evaluate changes in the proteomic profile of B. fragilis grown in a sub-MIC of PTZ, using 2-D electrophoresis followed by matrix-assisted laser desorption/ionization time-of-flight/time of-flight. Analysis of the 2-DE gels showed 18 spots with significantly different volume percentages between experimental conditions and 12 were successfully identified by MS/MS. Two proteins with decreased abundance in sub-MIC condition were involved in the glycolysis (glyceraldehyde-3-phosphate dehy- drogenase and triose phosphate isomerase), others two involved in amino acid metabolism (Oxoacyl- (acyl-carrier protein) synthase II and dihydrodipicolinate reductase), and finally, one protein involved in fatty acid metabolism (UDP-N-acetylglucosamine acyltransferase). Among the proteins with increased abundance, we founded three ATP synthase (alpha, beta, and alpha type V), which could be involved in antibiotic bacterial resistance by efflux pump, one protein involved in glycolysis (enolase), and one involved in protein degradation (aminoacyl-histidine dipeptidase). In conclusion, our data show overall changes in the proteome of B. fragilis conducted by sub-MIC of PTZ, whose consequences on bacterial physiology deserve further investigation. Ó 2013 Elsevier Ltd. All rights reserved. 1. Introduction The capacity of an antibiotic (ATB) to achieve and surpass the minimum inhibitory concentration (MIC) in a tissue depends on its pharmacokinetic and pharmacodynamic properties. There are various situations in which pathogens are subjected to ATBs at subinhibitory concentrations for a variable period of time. Among other consequences, this may result in treatment failure and the selection of resistant cells [1]. Since the 1940s, subinhibitory concentrations of ATBs have been the subject of numerous studies. Their effects on the physiology of a variety of bacterial species manifest mainly as morphological changes, structural changes in the cell wall, and changes in growth kinetics, the inhibition of enzymes and/or toxins, and the loss of adhesive properties [4,2,3,6,5]. In this context, some researchers have used transcriptome and proteome analysis to obtain a global and integrated view of bacterial metabolism in the presence of ATBs at subinhibitory concentrations [7]. Despite the undeniable clinical relevance of the effects of sub- inhibitory concentrations of ATBs on bacterial physiology, few studies have investigated these phenomena in obligate anaerobes. The most recent study on anaerobes was conducted by Gerber et al. * Corresponding author. Tel.: þ55 31 3409 2743. E-mail addresses: lucianocvbio@gmail.com (L.deC. Veloso), keniavaleria@ gmail.com (K.V. dos Santos), helidandrade@gmail.com (H.M. de Andrade), simonefpires@gmail.com (S.daF. Pires), simonegsantos@gmail.com (S.G. dos Santos), marianavazbio@yahoo.com.br (M.J. Vaz Trindade), macedo@icb.ufmg.br (L.deM. de Farias), marc@icb.ufmg.br (M.A.R. de Carvalho). Contents lists available at SciVerse ScienceDirect Anaerobe journal homepage: www.elsevier.com/locate/anaerobe 1075-9964/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.anaerobe.2013.04.007 Anaerobe 22 (2013) 69e76