A new SYBR green real-time PCR assay for semi-quantitative detection of Echinococcus multilocularis and Echinococcus canadensis DNA on bilberries (Vaccinium myrtillus) : Food and Waterborne Parasitology
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Malkamäki , S , Näreaho , A , Lavikainen , A , Oksanen , A & Sukura , A 2019 , ' A new SYBR green real-time PCR assay for semi-quantitative detection of Echinococcus multilocularis and Echinococcus canadensis DNA on bilberries (Vaccinium myrtillus) : Food and Waterborne Parasitology ' , Food and Waterborne Parasitology , vol. 17 , e00068 . https://doi.org/10.1016/j.fawpar.2019.e00068
| Title: | A new SYBR green real-time PCR assay for semi-quantitative detection of Echinococcus multilocularis and Echinococcus canadensis DNA on bilberries (Vaccinium myrtillus) : Food and Waterborne Parasitology |
| Author: | Malkamäki, S.; Näreaho, A.; Lavikainen, A.; Oksanen, A.; Sukura, A. |
| Contributor organization: | Veterinary Biosciences Veterinary Pathology and Parasitology HUSLAB Seppo Meri / Principal Investigator Departments of Faculty of Veterinary Medicine Helsinki One Health (HOH) Antti Sukura / Principal Investigator |
| Date: | 2019-11-06 |
| Language: | eng |
| Number of pages: | 7 |
| Belongs to series: | Food and Waterborne Parasitology |
| ISSN: | 2405-6766 |
| DOI: | https://doi.org/10.1016/j.fawpar.2019.e00068 |
| URI: | http://hdl.handle.net/10138/312746 |
| Abstract: | Berries and vegetables are potential transmission vehicles for eggs of pathogenic parasites, such as Echinococcus spp. We developed a SYBR Green based semi-quantitative real-time PCR (qPCR) method for detection of Echinococcus multilocularis and Echinococcus canadensis DNA from berry samples. A set of primers based on the mitochondrial NADH dehydrogenase subunit 1 (nad1) gene was designed and evaluated. To assess the efficacy of the assay, we spiked bilberries (Vaccinium myrtillus) with a known amount of E. multilocularis eggs. The detection limit for the assay using the NAD1_88 primer set was 4.37 × 10−5 ng/μl of E. multilocularis DNA. Under artificial contamination of berries, 50 E. multilocularis eggs were reliably detected in 250 g of bilberries. Analytical sensitivity of the assay was determined to be 100% with three eggs. As an application of the assay, 21 bilberry samples from Finnish market places and 21 bilberry samples from Estonia were examined. Previously described sieving and DNA extraction methods were used, and the samples were analyzed for E. multilocularis and E. canadensis DNA using semi-quantitative real-time PCR and a melting curve analysis of the amplified products. Echinococcus DNA was not detected in any of the commercial berry samples. This easy and fast method can be used for an efficient detection of E. multilocularis and E. canadensis in bilberries or other berries, and it is applicable also for fruits and vegetables. © 2019 The Authors |
| Subject: |
Bilberry
Cestoda Echinococcus qPCR SYBR green DNA adult Article bilberry controlled study DNA determination DNA extraction Echinococcus canadensis Echinococcus multilocularis Estonia Finland fruit limit of detection mitochondrial gene nad1 gene nonhuman parasite egg count parasite identification parasite transmission priority journal quantitative analysis real time polymerase chain reaction sensitivity and specificity vegetable 416 Food Science |
| Peer reviewed: | Yes |
| Rights: | cc_by_nc_nd |
| Usage restriction: | openAccess |
| Self-archived version: | publishedVersion |
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