A re-evaluation of the taxonomic position of five strains, one assigned to Cronobacter sakazakii (strain 1330T, isolated from spiced meat purchased in Slovakia), two previously assigned to Cronobacter genomospecies 1 (strains NCTC 9529T and 731, isolated from water and a leg infection, respectively) and two previously assigned to Cronobacter turicensis (strains 96 and 1435, isolated from onion powder and rye flour, respectively) was carried out. The analysis included phenotypic characterization, 16S rRNA gene sequencing and multilocus sequence analysis (MLSA) of seven housekeeping genes (atpD, fusA, glnS, gltB, gyrB, infB, ppsA; 3036 bp). 16S rRNA gene sequence analysis and MLSA showed that strain 1330T formed an independent phylogenetic lineage in the MLSA, with Cronobacter dublinensis LMG 23823T as the closest neighbour. DNA–DNA reassociation and phenotypic analysis revealed that strain 1330T represented a novel species, for which the name Cronobacter condimenti sp. nov. is proposed (type strain 1330T = CECT 7863T = LMG 26250T). Strains NCTC 9529T, 731, 96 and 1435 clustered together within an independent phylogenetic lineage, with C. turicensis LMG 23827T as the closest neighbour in the MLSA. DNA–DNA reassociation and phenotypic analysis confirmed that these strains represent a novel species, for which the name Cronobacter universalis sp. nov. is proposed (type strain NCTC 9529T = CECT 7864T = LMG 26249T).

Cronobacter spp. (formerly Enterobacter sakazakii) can be isolated from a wide range of foods and environments, and its association with neonatal infections has drawn considerable attention from regulatory authorities. The principle route of neonatal infection has been identified as the ingestion of contaminated infant formula. A number of methods have been developed to identify Cronobacter spp., however these were before the most recent (2012) taxonomic revision of the genus into seven species. In this study, phenotyping, protein profiling and molecular methods were used to identify Cronobacter strains which had been recently isolated from ingredients used in the preparation of infant formula.Pulsed field gel electrophoresis revealed that different Cronobacter strains had been recovered from the same food products. All isolates were identified as C sakazakii according to four genus specific PCR probes and protein profiling using MALDI-TOF analysis.However, 16S rDNA sequence analyses and fusA allele sequencing gave more accurate identification: four strains were C sakazakii, one strain was C malonaticus and the remaining strain was C universalis. Multilocus sequence typing showed the strains were different sequence types.These results demonstrate the presence of different Cronobacter species in food ingredients used in the preparation of infant formula, and also the need for molecular identification and profiling methods to be revised according to taxonomic revisions.

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