Mutations in Penicillin-Binding Proteins (PBP) Genes and in non-PBP Genes During Selection of Penicillin-resistant Streptococcus gordonii

Department of Fundamental Microbiology, University of Lausanne, Switzerland

^* To whom correspondence should be addressed. Email: philippe.moreillon{at}unil.ch.

	Abstract

Penicillin-resistance in Streptococcus spp. involves multiple mutations in both penicillin-binding proteins (PBPs) and non-PBP genes. Here we studied the development of penicillin-resistance in the oral commensal S. gordonii. Cyclic exposure of bacteria to 2-folds increasing penicillin concentrations selected for a progressive 250-500-fold MIC increase (from 0.008 to 2-4 µg/ml). The major MIC increase ( 35-fold) was related to non-PBP mutations, whereas PBP mutations accounted only for a 4 to 8-fold additional increase. PBP mutations occurred in the class B PBP 2X and 2B, which carry a transpeptidase domain, but not in class A PBP 1A, 1B or 2A, which carry an additional transglycosylase domain. Therefore, we tested whether inactivation of class A PBPs affected resistance development in spite of the absence of mutations. Deletion of PBP 1A or 2A profoundly slowed down resistance development, but only moderately affected resistance in already highly-resistant mutants (MIC = 2-4 µg/ml). Thus, class A PBPs might facilitate early development of resistance by stabilizing penicillin-altered peptidoglycan via transglycosylation, whereas they might be less indispensable in highly-resistant mutants which have re-established a penicillin-insensitive cell-wall building machinery. The contribution of PBP and non-PBP mutations alone could be individualized in DNA transformation. Both PBP and non-PBP mutations conferred some level of intrinsic resistance, but combining them together synergized to ensure high-level resistance ( 2 µg/ml). The results underline the complexity of penicillin-resistance development and suggest that inhibition of transglycosylase might be an as yet underestimated way to interfere with early resistance development.