Genotypic and phenotypic characterisation of a collection of Cronobacter (Enterobacter sakazakii) isolates

Enterobacter sakazakii has been identified as the causative agent of serious neonatal infections, associated with high mortality rate. In many cases, powdered infant formula (PIF) has been identified as the source of infection. Recently, E. sakazakii was proposed to be classified in a new genus, Cronobacter. Since knowledge on this pathogen is still incomplete, there is a need for molecular characterization schemes in order to help with epidemiological investigation and evaluate strain variability. The objectives of this study were to combine genotypic (pulsed-field gel electrophoresis [PFGE], 16S rRNA gene sequencing, and automated ribotyping) methods with traditional phenotypic biochemical methods to characterize a collection of Cronobacter isolates from various origins. In addition, the relative growth dynamics were compared by estimating the growth rates for each isolate in non-selective broth (BHI) at 25 degrees C and 37 degrees C. According to biochemical test profiles the majority of isolates were identified as Cronobacter sakazakii, which seemed to be the most common species distributed in the environment of PIF production plants. Furthermore, the PFGE technique displayed very high discriminatory power as 61 distinct pulsotypes were revealed among the 150 Cronobacter isolates. Combining information on sample origin and pulse type, 64 isolates were deemed as unique strains. Although genetic typing data for the strains clearly delineated them into clusters closely corresponding to biochemical speciation results, it was not without discrepancies as some strains did not group as predicted. Important for quantitative risk assessment is the fact that despite the high genetic heterogeneity observed for this collection, most Cronobacter strains displayed similar growth rates irrespective of species designation.