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16S rRNA gene based analysis of Enterobacter sakazakii strains from different sources and development of a PCR assay
Angelika Lehner1, Taurai Tasara, Roger Stephan
1Institute for Food Safety and Hygiene, Vetsuisse Faculty, University of Zurich, CH-8057 Zurich, Switzerland. lehnera@fsafety.unizh.ch <lehnera@fsafety.unizh.ch>
Background:
E. sakazakii is considered to be an opportunistic pathogen, implicated in food borne diseases causing meningitis or enteritis especially in neonates and infants. Cultural standard identification procedures for E. sakazakii include the observation of yellow pigmentation of colonies and a positive glucosidase activity. Up to now, only one PCR system based on a single available 16S rRNA gene sequence has been published for E. sakazakii identification. However, in our hands a preliminary evaluation of this system to a number of target and non-target strains showed significant specificity problems of this system. In this study full-length 16S rRNA genes of thirteen E. sakazakii strains from food, environment and human origin as well as the type strain ATCC 51329 were sequenced. Based on this sequence data a new specific PCR system for E. sakazakii was developed and evaluated.
Results:
By phylogenetic analysis of the new full-length 16S rRNA gene sequence data obtained we could show the presence of a second phylogenetic distinct lineage within the E. sakazakii species. The newly developed 16S rRNA gene targeting PCR system allows identification of E. sakazakii strains from both lineages. The assay's ability to correctly identify different E. sakazakii isolates as well as to differentiate E. sakazakii from other closely related Enterobacteriaceae species and other microorganisms was shown on 75 target and non-target strains.
Conclusion:
By this study we are presenting a specific and reliable PCR identification system, which is able to correctly identify E. sakazakii isolates from both phylogenetic distinct lines within the E. sakazakii species. The impact of this second newly described phylogenetic line within the E. sakazakii species in view of clinical and food safety aspects need further investigation.
Insights
A new PCR method accurately identifies all E. sakazakii strains, including a newly discovered lineage. This reliable identification system enhances food safety and clinical diagnostics for this opportunistic pathogen.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Cronobacter sakazakii (E. sakazakii) is an opportunistic pathogen causing severe infections in neonates and infants.
- Current identification relies on phenotypic traits and a limited PCR system with specificity issues.
- Previous PCR methods for E. sakazakii identification have shown significant specificity problems.
Purpose of the Study:
- To develop and evaluate a new, specific PCR identification system for E. sakazakii.
- To address the limitations of existing PCR methods for E. sakazakii detection.
- To sequence and analyze full-length 16S rRNA genes for improved E. sakazakii characterization.
Main Methods:
- Sequencing of full-length 16S rRNA genes from thirteen E. sakazakii strains and the type strain.
- Phylogenetic analysis of the obtained 16S rRNA gene sequence data.
- Development and evaluation of a novel PCR system targeting the 16S rRNA gene.
Main Results:
- Discovery of a second, phylogenetically distinct lineage within the E. sakazakii species.
- The new PCR system successfully identified E. sakazakii strains from both lineages.
- The assay demonstrated high specificity, correctly identifying 75 target and non-target strains.
Conclusions:
- A specific and reliable PCR identification system for E. sakazakii has been developed.
- This system accurately identifies E. sakazakii isolates from both identified phylogenetic lineages.
- Further investigation is needed to understand the clinical and food safety implications of the newly described phylogenetic line.
