High-level resistance to class IIa bacteriocins is associated with one general mechanism in Listeria monocytogenes

Department of Dairy and Food Science, Centre for Advanced Food Studies, LMC, The Royal Veterinary and Agricultural University, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark 1 Department of Biochemistry, University of Stellenbosch, Private Bag X1, 7602 Matieland, South Africa 2 German Research Centre for Biotechnology, Department of Cell Biology, Mascheroder Weg 1, Braunschweig D-38124, Germany 3 Laboratoire de Microbiologie Fondamentale et Appliquée, CNRS FRE 2224, IBMIG, UFR Sciences, 40 Avenue du Recteur Pineau, 86022 Poitiers Cedex, France 4 Author for correspondence: Anne Gravesen. Tel: +45 3528 3272. Fax: +45 3528 3231. e-mail: alg{at}kvl.dk Class IIa bacteriocins may be used as natural food preservatives, yet resistance development in the target organisms is still poorly understood. In this study, the understanding of class IIa resistance development in Listeria monocytogenes is extended, linking the seemingly diverging results previously reported. Eight resistant mutants having a high resistance level (at least a 10 3 -fold increase in MIC), originating from five wild-type listerial strains, were independently isolated following exposure to four different class IIa bacteriocin-producing lactic acid bacteria (including pediocin PA-1 and leucocin A producers). Two of the mutants were isolated from food model systems (a saveloy-type sausage at 10 °C, and salmon juice at 5 °C). Northern blot analysis showed that the eight mutants all had increased expression of EII Bgl and a phospho-ß-glucosidase homologue, both originating from putative ß-glucoside-specific phosphoenolpyruvate-dependent phosphotransferase systems (PTSs). However, disruption of these genes in a resistant mutant did not confer pediocin sensitivity. Comparative two-dimensional gel analysis of proteins isolated from mutant and wild-type strains showed that one spot was consistently missing in the gels from mutant strains. This spot corresponded to the MptA subunit of the mannose-specific PTS, , found only in the gels of wild-type strains. The mptACD operon was recently shown to be regulated by the 54 transcription factor in conjunction with the activator ManR. Class IIa bacteriocin-resistant mutants having defined mutations in mpt or manR also exhibited the two diverging PTS expression changes. It is suggested here that high-level class IIa resistance in L. monocytogenes and at least some other Gram-positive bacteria is developed by one prevalent mechanism, irrespective of wild-type st