Characterization of surface proteins of Cronobacter muytjensii using monoclonal antibodies and MALDI-TOF Mass spectrometry
1 Department of Biotechnology and Genetic Engineering, P. O Box 3030, Jordan University of Science and Technology, Irbid 22110, Jordan
2 Department of Biochemistry and Biophysics, Texas A & M University, College Station, Texas 77843-2128, USA
3 Princes Haya Biotechnology Center, Jordan University of Science and Technology, Irbid, 22110, Jordan
4 Department of Food Science, Purdue University, West Lafayette, Indiana, 47907, USA
BMC Microbiology 2011, 11:148 doi:10.1186/1471-2180-11-148Published: 25 June 2011
Cronobacter spp. is a newly emerging pathogen that causes meningitis in infants and other diseases in elderly and immunocompromised individuals. This study was undertaken to investigate surface antigenic determinants in Cronobacter spp. using monoclonal antibodies (MAbs) and MALDI-TOF Mass spectrometry.
Spleenocytes from mice that were immunized with heat-killed (20 min, 80°C) Cronobacter cells were fused with SP2 myeloma cells. Five desirable MAbs (A1, B5, 2C2, C5 and A4) were selected. MAbs A1, B5, 2C2 and C5 were of IgG2a isotype while A4 was an IgM. Specificity of the MAbs was determined by using immunoblotting with outer membrane protein preparations (OMPs) extracted from 12 Cronobacter and 6 non-Cronobacter bacteria. All MAbs recognized proteins with molecular weight ranging between 36 and 49 kDa except for one isolate (44) in which no OMPs were detected. In addition, MAbs recognized two bands (38-41 kDa) in four of the non-Cronobacter bacteria. Most of the proteins recognized by the MAbs were identified by MALDI-TOF peptide sequencing and appeared to be heterogeneous with the identities of some of them are still unknown. All MAbs recognized the same epitope as determined by an additive Index ELISA with their epitopes appeared to be conformational rather than sequential. Further, none of the MAbs recognized purified LPS from Cronobacter spp. Specificity of the MAbs toward OMPs was further confirmed by transmission electron microscopy.
Results obtained in this study highlight the immunological cross-reactivity among Cronobacter OMPs and their Enterobacteriaceae counterparts. Nevertheless, the identity of the identified proteins appeared to be different as inferred from the MALDI-TOF sequencing and identification.