Detection and quantification of the iap gene of Listeria monocytogenes and Listeria innocua by a new real-time quantitative PCR assay

Hein, Ingeborg; Klein, Dieter; Lehner, Angelika; Bubert, Andreas; Brandl, Ernst; Wagner, Martin

doi:10.1016/s0923-2508(00)01166-9

Cited by 121 publications

(82 citation statements)

References 15 publications

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“…The detection limit of 1-10 cell equivalents per PCR reaction was comparable with published real-time PCR assays for L. monocytogenes (Hein et al, 2001;Rodriguez-Lazaro et al, 2004a;Berrada et al, 2006). Also, based on inoculation of a variety of different foods, the assay was capable of detecting 1-5 CFU per 25 g/ml with no interference from the microflora present in these food samples.…”

Section: Article In Presssupporting

confidence: 67%

“…Recently conventional and real-time PCR assays have been developed for the detection of L. monocytogenes in foods (Nogva et al, 2000;Hein et al, 2001;Bhagwat, 2003;Koo and Jaykus, 2003;Somer and Kashi, 2003;D'Agostino et al, 2004;Rodriguez-Lazaro et al, 2004b;Wang et al, 2004;Rudi et al, 2005). Many are based on the detection of virulence genes in L. monocytogenes and have been tested in a variety of foods.…”

Section: Discussionmentioning

confidence: 99%

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Rapid real-time PCR detection of Listeria monocytogenes in enriched food samples based on the ssrA gene, a novel diagnostic target

et al. 2008

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A real-time PCR assay was designed to detect a 162-bp fragment of the ssrA gene in Listeria monocytogenes. The specificity of the assay for L. monocytogenes was confirmed against a panel of 6 Listeria species and 26 other bacterial species. A detection limit of 1-10 genome equivalents was determined for the assay. Application of the assay in natural and artificially contaminated culture enriched foods, including soft cheese, meat, milk, vegetables and fish, enabled detection of 1-5 CFU L. monocytogenes per 25 g/ml of food sample in 30 h. The performance of the assay was compared with the Roche Diagnostics 'LightCycler foodproof Listeria monocytogenes Detection Kit'. Both methods detected L. monocytogenes in all artificially contaminated retail samples (n ¼ 27) and L. monocytogenes was not detected by either system in 27 natural retail food samples. The method developed in this study has the potential to enable the specific detection of L. monocytogenes in a variety of food types in a time-frame considerably faster than current standard methods. The potential of the ssrA gene as a nucleic acid diagnostic (NAD) target has been demonstrated in L. monocytogenes. We are currently developing NAD tests based on the ssrA gene for a range of common foodborne and clinically relevant bacterial pathogens. r

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Section: Article In Presssupporting

confidence: 67%

Section: Discussionmentioning

confidence: 99%

Rapid real-time PCR detection of Listeria monocytogenes in enriched food samples based on the ssrA gene, a novel diagnostic target

et al. 2008

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“…Due to the unsatisfactory results obtained, we decided to perform further tests employing probebased assays in order to identify an adequate specificity. Primers and probes targeting iap gene designed by Hein et al (2001) were chosen for this assay. The assay was tested against the strains previously analyzed using the same protocols.…”

Section: Qpcr Assaysmentioning

confidence: 99%

Detection and Enumeration of Listeria Monocytogenes in Fresh Cut Vegetables Using MPN-Real-Time PCR

Russo¹,

Botticella²,

Amodio³

et al. 2015

Acta Hortic.

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Listeria to 10 g of food were incubated in Fraser broth at 30°C for 48 h for standard MPN analysis. After incubation, broth samples were taken from each tube and DNA was extracted. DNA from enrichment tubes was used as template in a qPCR assay targeting a 64 bp hlyA gene sequence of L. monocytogenes. Results of this assay were than compared with those of standard MPN analysis and a complete accordance was observed. Furthermore, we tested an enrichment free approach using the same qPCR assay. Samples were prepared as described for MPN-qPCR while DNA extraction was performed prior to enrichment of inoculated salads. This approach allowed us to identify L. monocytogenes in samples spiked with 10-10 5 CFU/g. The whole process, including DNA extraction, required less than four hours, thus providing a fast and reliable tool for detection of L. monocytogenes in fresh cut vegetables.

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“…Previous studies (Hein et al, 2001;Nogva et al, 2000;Norton & Batt, 1999) have indicated that the LLO gene (hly) and p60 gene (iap) are the most commonly used PCR amplification targets for the detection of L. monocytogenes. Thus, the detection of these two genes at the transcript level has important practical significance in food samples.…”

Section: Honokiol Represses the Transcription Of Genes Encoding Llo Amentioning

confidence: 99%

Effect of honokiol on exotoxin proteins listeriolysin O and p60 secreted by Listeria monocytogenes

Meng

Zhao

Guo

et al. 2015

Journal of Medical Microbiology

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Listeria monocytogenes is considered one of the most important foodborne pathogens. The virulence-related proteins listeriolysin O (LLO) and p60 are critical factors involved in Listeria pathogenesis. In the present study, we investigated the effect of honokiol on LLO and p60 secreted from L. monocytogenes. A listeriolysin assay was used to investigate the haemolytic activities of L. monocytogenes exposed to honokiol, and the secretion of LLO and p60 was detected by immunoblot analysis. Additionally, the influence of honokiol on the transcription of LLO and p60 genes (hly and iap, respectively) was analysed by real-time reverse transcription PCR. TNF-a release assays were performed to elucidate the biological relevance of changes in LLO and p60 secretion induced by honokiol. According to the data, honokiol showed good anti-L. monocytogenes activity, with MICs of 8-16 mg ml 21 , and the secretion of LLO and p60 was decreased by honokiol. In addition, the transcription of hly and iap was inhibited by honokiol. Our results indicate that TNF-a production by RAW264.7 cells stimulated with L. monocytogenes supernatants was inhibited by honokiol. Based on these data, we propose that honokiol could be used as a promising natural compound against L. monocytogenes and its virulence factors.

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Detection and quantification of the iap gene of Listeria monocytogenes and Listeria innocua by a new real-time quantitative PCR assay

Cited by 121 publications

References 15 publications

Rapid real-time PCR detection of Listeria monocytogenes in enriched food samples based on the ssrA gene, a novel diagnostic target

Rapid real-time PCR detection of Listeria monocytogenes in enriched food samples based on the ssrA gene, a novel diagnostic target

Detection and Enumeration of Listeria Monocytogenes in Fresh Cut Vegetables Using MPN-Real-Time PCR

Effect of honokiol on exotoxin proteins listeriolysin O and p60 secreted by Listeria monocytogenes

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