Downloaded from www.microbiologyresearch.org by IP: 93.91.26.109 On: Wed, 27 Apr 2016 22:51:53 Identification of HLA-A*01- and HLA-A*02- restricted CD8 + T-cell epitopes shared among group B enteroviruses Andreas O. Weinzierl, 1,2 Despina Rudolf, 1 Dominik Maurer, 1 Dorothee Wernet, 3 Hans-Georg Rammensee, 1 Stefan Stevanovic ´ 1 and Karin Klingel 2 Correspondence Karin Klingel karin.klingel@med. uni-tuebingen.de 1 Department of Immunology, Institute for Cell Biology, University of Tu ¨ bingen, Auf der Morgenstelle 15, 72076 Tu ¨ bingen, Germany 2 Department of Molecular Pathology, University of Tu ¨ bingen, Liebermeisterstraße 8, 72076 Tu ¨ bingen, Germany 3 Institute of Clinical and Experimental Transfusion Medicine, University of Tu ¨ bingen, Otfried-Mu ¨ ller- Str. 4/1, 72076 Tu ¨ bingen, Germany Received 22 January 2008 Accepted 15 May 2008 Acute enteroviral infections ranging from meningitis, pancreatitis to myocarditis are common and normally well controlled by the host immune system comprising virus-specific CD8 + cytotoxic T lymphocytes (CTL). However, in some patients enteroviruses and especially coxsackieviruses of group B are capable of inducing severe chronic forms of diseases such as chronic myocarditis. Currently, it is not known whether divergences in the CTL-related immune response may contribute to the different outcome and course of enterovirus myocarditis. A pre-requisite for the study of CTL reactions in patients with acute and chronic myocarditis is the identification of CTL epitopes. In order to define dominant enterovirus CTL epitopes, we have screened, by using gamma interferon (IFN-c) ELISPOT, 62 HLA-A*01- and 59 HLA-A*02-positive healthy blood donors for pre-existing CTL reactions against 12 HLA-A*01 and 20 HLA-A*02 predicted CTL epitopes derived from coxsackieviruses of group B. Positive CTL reactions were verified by FACS analysis in a combined major histocompatibility complex-tetramer IFN-c staining. A total of 14.8 % of all donors reacted against one of the three identified epitopes MLDGHLIAFDY, YGDDVIASY or GIIYIIYKL. The HLA-A*02-restricted epitope ILMNDQEVGV was recognized by 25 % of all tested blood donors. For this peptide, we could demonstrate specific granzyme B secretion, a strong cytolytic potential and endogenous processing. All four epitopes were homologous in 36– 92 % of group B enteroviruses, providing a strong basis for monitoring the divergence of T-cell- based immune responses in enterovirus-induced acute and chronic diseases. INTRODUCTION Enteroviruses are non-enveloped, positive-strand RNA viruses that belong to the family Picornaviridae. Currently, the enterovirus genus represents 68 human serotypes classified into five species [A–D and polioviruses (Stanway et al., 2005)] differing by about 40 % in their nucleotide sequence (Simmonds & Welch, 2006). Enteroviral evolution occurs through genetic drift (Chua et al., 2001), which is primarily prominent in group B enteroviruses (Simmonds & Welch, 2006), including coxsackieviruses of group B (CVB). In addition, the occurrence of interserotypic recombination events between different coxsackievirus strains has also been reported (Bouslama et al., 2007). It has been estimated that 5–10 million symptomatic infections with non-polio enteroviruses occur annually in the USA (Strikas et al., 1986). Although most of these infections are associated with a mild febrile illness or upper respiratory infection (Rotbart et al., 1999), enteroviruses can also cause severe, potentially fatal diseases such as acute flaccid paralysis [caused by CVB in 13 % of all cases (Saeed et al., 2007)], aseptic meningitis [caused by enteroviruses in up to 90 % of all cases (Rotbart, 1995)] or acute viral myocarditis with CVB3 as one of the primary causative agents (Baboonian et al., 1997; Kim et al., 2001; McManus et al., 1991). Whereas in adults acute CVB myocarditis is rarely lethal, children are particularly susceptible to diverse CVB infections with mortality rates due to myocarditis in the range of 30–50 % (Modlin & Rotbart, 1997). In most patients acute myocarditis is abrogated by the successful Supplementary tables are available with the online version of this paper. Journal of General Virology (2008), 89, 2090–2097 DOI 10.1099/vir.0.2008/000711-0 2090 2008/000711 G 2008 SGM Printed in Great Britain