How to identify Raoultella spp. including R. ornithinolytica isolates negative for ornithine decarboxylase? The reliability of the chromosomal bla gene

Although Raoultella planticola and Raoultella ornithinolytica were described more than 20 years ago, identifying them remains difficult. The reliability of the chromosomal bla gene for this identification was evaluated in comparison with that of the 16S rDNA and rpoB genes in 35 Raoultella strains from different origins. Of the 26 strains previously identified as R. planticola by biochemical tests alone or in association with molecular methods, 21 harboured a bla gene with 99.8% identity with the bla gene of two reference R. ornithinolytica strains (blaORN gene) and 5 harboured a bla gene with 99.2% identity with the bla gene of two reference R. planticola strains (blaPLA gene). The 9 isolates previously identified as R. ornithinolytica harboured a blaORN gene. The bla gene-based identification was confirmed by 16S rDNA and rpoB sequencing. The 21 isolates newly identified as R. ornithinolytica had a test negative for ornithine decarboxylase (ODC). Molecular experiments suggested one copy of ODC-encoding gene in both ODC-negative R. ornithinolytica and R. planticola strains and two copies in ODC-positive R. orninthinolytica strains. Analysis of the 35 bla genes allowed us (i) to confirm an identity of only 94% between the bla genes of the two Raoultella species while this identity was > 98% for rpoB and > 99% for 16S rDNA genes and (ii) to develop and successfully apply a bla PCR RFLP assay for Raoultella spp. identification. Overall, this study allowed us to discover ODC-negative R. ornithinolytica and to provide a reliable Raoultella identification method widely available as not requiring sequencing equipment.