Major metalloprotease gene of Serratia marcescens is conserved and provides a molecular typing method to differentiate clinical isolates

The occurrence and heterogeneity of the 51-kDa major metalloprotease gene from Serratia marcescens which is thought to be involved in pathogenesis was examined using polymerase chain reaction (PCR) and restriction endonuclease analysis (REA). Ten clinical isolates of S. marcescens associated with a previously characterized outbreak and 80 epidemiologically unrelated isolates were investigated. In all isolates, a 1.5-kbp DNA fragment was amplified from the metalloprotease gene. REA of the amplified DNA showed 3 restriction fragment length polymorphism (RFLP) patterns using HincII and 5 patterns using RsaI. Molecular subtyping with the protease gene was performed on 20 epidemiologically unrelated isolates and 10 outbreak-associated isolates. Size-separated genomic DNA fragments from HincII or RsaI enzyme digestions were hybridized with the 1.5-kbp metalloprotease gene probe by the method of Southern. Analysis of combined hybridization profiles of all 30 isolates produced 23 different protease genotypes. All 20 epidemiologically unrelated isolates had different profiles, whereas 8 of 10 outbreak-associated isolates showed identical profiles. Discriminatory power of protease genotyping method was compared with the established molecular subtyping method of ribotyping using a gene probe for Escherichia coli 16S rRNA. Results showed agreement for 20 of the 23 genotypes by both methods. Two other epidemiologically unrelated isolates that displayed the same ribotype were differentiated by protease genotyping. Molecular subtyping using the major metalloprotease gene of S. marcescens reported here provided differentiation of clinically related and unrelated isolates and could be used in epidemiological investigations to increase the discriminatory power of ribotyping.