Quantitative Analysis of Staphylococcus aureus in Skimmed Milk Powder by Real-Time PCR

Ikeda, Tetsuya; Tamate, Naoto; Yamaguchi, Keiji; Makino, Sou‐ichi

doi:10.1292/jvms.67.1037

Cited by 21 publications

(15 citation statements)

References 17 publications

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“…The same may be observed when the milk is not cooled properly after milking. Ikeda et al (2005) reported an outbreak in Japan in 2000, with more than 10 000 cases of staphylococcal enterotoxicosis, milk powder produced from pasteurised milk having been identified as the source of the infection.…”

Section: Resultsmentioning

confidence: 99%

Influence of soft cheese technology on the growth and enterotoxin production of Staphylococcus aureus

Necidová¹,

Šťástková²,

Pospíšilová³

et al. 2009

Czech J. Food Sci.

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Necidová L., Šťástková Z., Pospíšilová M., Janštová B., Strejček J., Dušková M., Karpíšková R. The aim of this study was to monitor S. aureus growth and toxin production in soft cheese during the technological processing. In model experiments, raw milk was inoculated separately with five S. aureus strains isolated from milk and milk products. All the strains were producers of staphylococcal enterotoxins (SEs) of types A, B, or C. SEs were detected by the enzyme-linked fluorescence assay (ELFA) performed in the MiniVIDAS device. This study has shown that the amount of SEs varied with the tested strains and stages of the technological process. SEs were detected in soft cheese made from pasteurised milk inoculated with 2.9 × 10 5 CFU/g of S. aureus. The prevention of S. aureus contamination and multiplication during the cheese making process is a prerequisite for the production of safe soft cheese. The most important enterotoxin dose build-up factor can be overcome by strict compliance with the cooling requirements during the manufacture, distribution and storage of the product.

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Section: Resultsmentioning

confidence: 99%

Influence of soft cheese technology on the growth and enterotoxin production of Staphylococcus aureus

Necidová¹,

Šťástková²,

Pospíšilová³

et al. 2009

Czech J. Food Sci.

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“…Several PCR-based methods for foodborne S. aureus identification have been published, targeting only a few different species-specific molecular markers. The predominant target was nuc gene used in either conventional PCR (Ercolini et al 2004;Pinto et al 2005;Ikeda et al 2005;Cremonesi et al 2005) or quantitative real-time PCR (Hein et al 2005;Alarcon et al 2006). Other popular targets were species-specific regions of the DNA coding for 16S or 23S rRNA or coa gene (Cremonesi et al 2005;Baron et al 2004;Sabet et al 2006) and putative transcriptional regulator genes (Liu et al 2005;Goto et al 2007).…”

Section: Introductionmentioning

confidence: 99%

Rapid and Sensitive Detection of Staphylococcus aureus in Food Using Selective Enrichment and Real-Time PCR Targeting a New Gene Marker

et al. 2008

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Staphylococcus aureus is a bacterial pathogen considered a principal etiological agent of food poisoning. The aim of this study was to develop and evaluate a rapid and sensitive method for the detection of S. aureus in food by using selective enrichment and a new species-specific real-time polymerase chain reaction (PCR). Specific primers and a TaqMan probe targeted to specific S. aureus gene encoding for acriflavine resistance protein were designed. The real-time PCR was highly specific for S. aureus with 100% inclusivity and 100% exclusivity determined using 83 S. aureus strains and 64 non-S.-aureus strains. PCR detection limit of 6.8×10 1 and 3.4×10 1 CFU ml −1 were obtained with 100% and 70% detection probability, respectively. The single selective enrichment based on the study of different enrichment conditions was selected and a lysis by boiling was used to obtain bacterial DNA. Out of 112 food samples analyzed, 61 were positive by the PCR-based method and 53 by the standard method. Out of ten food matrices artificially contaminated at a level of 10°CFU g −1 , ten and six were positive by the respective methods. Moreover, 10°CFU 10 g −1 was detected in all ten artificially contaminated samples after a large-scale enrichment using PCR-based detection, in contrast to seven false negative by standard detection. The developed method facilitated the detection of S. aureus on the next day after the sample reception. This method can be used for S. aureus detection as a faster, highly specific, and more sensitive alternative to microbiological method with the potential for providing of improved food-processing hygiene control.

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“…Nakayama et al (2006) konnten sea -sei in künstlich kontaminierter Milch mit Nachweisgrenzen von 10 2 -10 4 KbE/ml nachweisen. Ikeda et al (2005b) untersuchten in Zusammenhang mit einem Ausbruch durch SEA-bildende S. aureus Milchpulver quantitativ auf sea, um damit indirekt auf die Belastung mit S. aureus zu schließen. Eine allgemeine Verwendung quantitativer PCR-Nachweise für SE-Gene zur Abschätzung des Toxingehaltes von Lebensmitteln ist jedoch bei unbekannten Proben kaum möglich, da die Nachweise aufgrund unterschiedlicher Nachweisgrenzen für die einzelnen Toxine für jedes Lebensmittel und jedes Toxin kalibriert werden müßten (Letertre et al, 2003a, Ercolini et al, 2004Nakayama et al, 2006).…”

Section: Polymerasekettenreaktionunclassified

“…Allerdings wurden häufi g nur Reinkulturen oder klinisches Material untersucht. Alarcón et al (2006) und Ikeda et al (2005b) verwendeten den DNeasy Tissue Kit von Qiagen für die Aufbereitung von Rindfl eischproben bzw. Milchpulver.…”

Section: Dna-extraktionunclassified

Staphylokokken-Enterotoxine: Bildung, Eigenschaften und Nachweis

2007

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Zusammenfassung: Staphylococcus aureus kann sich unter geeigneten Bedingungen in Lebensmitteln vermehren und dabei Enterotoxine bilden, die beim Konsumenten Erbrechen und Durchfall auslösen. Im Gegensatz zu dem Erreger selbst sind diese Toxine sehr widerstandsfähig insbesondere gegenüber hohen Temperaturen und können durch eine küchenmäßige Bearbeitung des sie enthaltenden Lebensmittels im allgemeinen nicht mehr inaktiviert werden. Da demnach die Anwesenheit der Staphylokokken zur Auslösung der Erkrankung nicht mehr erforderlich ist, handelt es sich bei dieser um eine typische Lebensmittelintoxikation. Die Abschätzung einer potentiellen Gesundheitsgefährdung über den Erregernachweis ist daher nur bei rohen Lebensmitteln sinnvoll. Bei Produkten, die im Verlauf ihrer Herstellung einem für die Keime abträglichen Verfahren, z. B. einer Wärmebehandlung, unterzogen wurden, kann der Thermonukleasetest (indirekter Nachweis einer Kontamination mit hoher Anzahl an Staphylokokken, unabhängig davon, ob die Erreger zum Zeitpunkt der Untersuchung noch präsent sind) oder der direkte Enterotoxinnachweis durchgeführt werden. Der Nachweis der Toxingene mit molekularbiologischen Verfahren hat in diesem Zusammenhang nur unterstützende Funktion, da im Fall eines positiven Ergebnisses aus diesem nicht geschlossen werden kann, daß die Gene auch exprimiert wurden, also Toxine im Lebensmittel vorhanden sind. In der vorliegenden Übersichtsarbeit werden der Erreger und seine Enterotoxine sowie die verschiedenen Nachweisverfahren unter besonderer Berücksichtigung der molekularbiologischen Methoden besprochen.Abstract: Under appropriate conditions, Staphylococcus aureus can grow in food and produce enterotoxins which cause vomiting and diarrhoea when ingested. In contrast to the staphylococci these toxins are resistant especially to high temperatures and are not inactivated by usual kitchen practice. As the presence of staphylococci is not essential to cause illness, this condition is a typical food poisoning. The estimation of a potential health risk by targeting the organism is therefore only appropriate to raw food. For products which underwent a procedure detrimental to the bacteria, e. g. heat treatment, testing for thermonuclease (indirect detection of a contamination with high numbers of staphylococci, irrespective of the presence of living bacteria at the time of testing) or the detection of enterotoxins can be applied. In this context, the detection of enterotoxin genes using molecular biological tests provides only supplemental information, as positive results are not evidentiary for gene expression and thus for the presence of toxins in the food. In this review, the causative organism and its toxins as well as methods of detection are discussed with special emphasis on molecular biological methods.

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Quantitative Analysis of Staphylococcus aureus in Skimmed Milk Powder by Real-Time PCR

Cited by 21 publications

References 17 publications

Influence of soft cheese technology on the growth and enterotoxin production of Staphylococcus aureus

Influence of soft cheese technology on the growth and enterotoxin production of Staphylococcus aureus

Rapid and Sensitive Detection of Staphylococcus aureus in Food Using Selective Enrichment and Real-Time PCR Targeting a New Gene Marker

Staphylokokken-Enterotoxine: Bildung, Eigenschaften und Nachweis

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