Advanced sample preparation for the molecular quantification of Staphylococcus aureus in artificially and naturally contaminated milk.
(2011) In International Journal of Food Microbiology 145. p.61-65- Abstract
- Sample treatment is an essential element when using real-time PCR for quantification of pathogens directly on food samples. This study comparatively evaluated three different principles of sample treatment, i.e. immunomagnetic separation based on phage-derived cell wall binding molecules, matrix solubilization and flotation, in order to establish their suitability for quantifying low numbers of Staphylococcus aureus in milk. All three procedures succeeded to remove S. aureus from the milk matrix, either raw or pasteurized, and, as a result of the concentration of the target cells, minimized the effect of milk associated PCR inhibitors. Sample preparation based on immunomagnetic separation albeit of being user friendly, specific and rapid,... (More)
- Sample treatment is an essential element when using real-time PCR for quantification of pathogens directly on food samples. This study comparatively evaluated three different principles of sample treatment, i.e. immunomagnetic separation based on phage-derived cell wall binding molecules, matrix solubilization and flotation, in order to establish their suitability for quantifying low numbers of Staphylococcus aureus in milk. All three procedures succeeded to remove S. aureus from the milk matrix, either raw or pasteurized, and, as a result of the concentration of the target cells, minimized the effect of milk associated PCR inhibitors. Sample preparation based on immunomagnetic separation albeit of being user friendly, specific and rapid, failed to allow quantification of low and medium numbers (<10(4)CFU) of S. aureus. In a mastitic milk model cell wall binding domain (CBD)-based target cell extraction revealed results most closely matching those derived from culture-based quantification. Both matrix lysis and flotation allowed quantification of S. aureus at a level of 1-10 cells per ml. Both methods resulted in higher numbers of bacterial cell equivalents (bce) than plating could reveal. Since both methods harvest cells that have been subjected to either mechanical and chemical stresses before quantification, we concluded that the higher bce numbers resulted from a disaggregation of S. aureus clusters initially present in the inoculum. Conclusively, since likely each S. aureus cell of a toxigenic strain contributes to enterotoxin production, molecular quantification could provide an even more realistic impact assessment in outbreak investigations than plating does. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/1732556
- author
- Aprodu, Iuliana ; Walcher, Georg ; Schelin, Jenny LU ; Hein, Ingeborg ; Norling, Börje ; Rådström, Peter LU ; Nicolau, Anca and Wagner, Martin
- organization
- publishing date
- 2011
- type
- Contribution to journal
- publication status
- published
- subject
- in
- International Journal of Food Microbiology
- volume
- 145
- pages
- 61 - 65
- publisher
- Elsevier
- external identifiers
-
- wos:000289819900011
- pmid:21035221
- scopus:79953034873
- pmid:21035221
- ISSN
- 0168-1605
- DOI
- 10.1016/j.ijfoodmicro.2010.09.018
- language
- English
- LU publication?
- yes
- id
- 99250c4f-177c-4d02-8ce9-2b66cb29aef2 (old id 1732556)
- date added to LUP
- 2016-04-01 10:37:55
- date last changed
- 2022-01-26 01:02:06
@article{99250c4f-177c-4d02-8ce9-2b66cb29aef2,
abstract = {{Sample treatment is an essential element when using real-time PCR for quantification of pathogens directly on food samples. This study comparatively evaluated three different principles of sample treatment, i.e. immunomagnetic separation based on phage-derived cell wall binding molecules, matrix solubilization and flotation, in order to establish their suitability for quantifying low numbers of Staphylococcus aureus in milk. All three procedures succeeded to remove S. aureus from the milk matrix, either raw or pasteurized, and, as a result of the concentration of the target cells, minimized the effect of milk associated PCR inhibitors. Sample preparation based on immunomagnetic separation albeit of being user friendly, specific and rapid, failed to allow quantification of low and medium numbers (<10(4)CFU) of S. aureus. In a mastitic milk model cell wall binding domain (CBD)-based target cell extraction revealed results most closely matching those derived from culture-based quantification. Both matrix lysis and flotation allowed quantification of S. aureus at a level of 1-10 cells per ml. Both methods resulted in higher numbers of bacterial cell equivalents (bce) than plating could reveal. Since both methods harvest cells that have been subjected to either mechanical and chemical stresses before quantification, we concluded that the higher bce numbers resulted from a disaggregation of S. aureus clusters initially present in the inoculum. Conclusively, since likely each S. aureus cell of a toxigenic strain contributes to enterotoxin production, molecular quantification could provide an even more realistic impact assessment in outbreak investigations than plating does.}},
author = {{Aprodu, Iuliana and Walcher, Georg and Schelin, Jenny and Hein, Ingeborg and Norling, Börje and Rådström, Peter and Nicolau, Anca and Wagner, Martin}},
issn = {{0168-1605}},
language = {{eng}},
pages = {{61--65}},
publisher = {{Elsevier}},
series = {{International Journal of Food Microbiology}},
title = {{Advanced sample preparation for the molecular quantification of Staphylococcus aureus in artificially and naturally contaminated milk.}},
url = {{http://dx.doi.org/10.1016/j.ijfoodmicro.2010.09.018}},
doi = {{10.1016/j.ijfoodmicro.2010.09.018}},
volume = {{145}},
year = {{2011}},
}