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Development and evaluation of rpoB based PCR systems to differentiate the six proposed species within the genus Cronobacter


Stoop, B; Lehner, A; Iversen, C; Fanning, S; Stephan, R (2009). Development and evaluation of rpoB based PCR systems to differentiate the six proposed species within the genus Cronobacter. International Journal of Food Microbiology, 136(2):165-168.

Abstract

Although there are various PCR based methods described in the literature to detect the genus Cronobacter, none of these methods is able to differentiate the six proposed species within the genus Cronobacter. Here we report for the first time different rpoB based PCR systems which enable the identification to species level of strains previously confirmed to belong to the genus Cronobacter.
Different primer pairs based on the rpoB sequences of the six Cronobacter species type strains were designed. Thereafter, 58 target and non-target strains, previously described in the Cronobacter taxonomy paper (Iversen et al. 2007a), were included for the specificity evaluation. C. turicensis, C. muytjensii, C. dublinensis and C. genomospecies 1 can be reliably identified by the proposed single primer pairs. Only target strains showed a correctly sized amplification product, whereas no amplification product was obtained for all non-target strains used in this study (100% specificity). However, as the rpoB gene sequences of C. sakazakii and C. malonaticus are closely related, a two step procedure is necessary in order to discern C. malonaticus from C. sakazakii. We therefore recommend a two-step procedure in which the primer pairs Cmalf/Cmalr are used in a follow up PCR on strains that are found to be positive in the amplification with the C. sakazakii specific primers Csakf/Csakr.

Abstract

Although there are various PCR based methods described in the literature to detect the genus Cronobacter, none of these methods is able to differentiate the six proposed species within the genus Cronobacter. Here we report for the first time different rpoB based PCR systems which enable the identification to species level of strains previously confirmed to belong to the genus Cronobacter.
Different primer pairs based on the rpoB sequences of the six Cronobacter species type strains were designed. Thereafter, 58 target and non-target strains, previously described in the Cronobacter taxonomy paper (Iversen et al. 2007a), were included for the specificity evaluation. C. turicensis, C. muytjensii, C. dublinensis and C. genomospecies 1 can be reliably identified by the proposed single primer pairs. Only target strains showed a correctly sized amplification product, whereas no amplification product was obtained for all non-target strains used in this study (100% specificity). However, as the rpoB gene sequences of C. sakazakii and C. malonaticus are closely related, a two step procedure is necessary in order to discern C. malonaticus from C. sakazakii. We therefore recommend a two-step procedure in which the primer pairs Cmalf/Cmalr are used in a follow up PCR on strains that are found to be positive in the amplification with the C. sakazakii specific primers Csakf/Csakr.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:05 Vetsuisse Faculty > Institute of Food Safety and Hygiene
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:2009
Deposited On:12 Jan 2010 16:24
Last Modified:06 Dec 2017 22:30
Publisher:Elsevier
ISSN:0168-1605
Publisher DOI:https://doi.org/10.1016/j.ijfoodmicro.2009.04.023
PubMed ID:19467725

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