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    Detection of Trichinella spiralis, T. britovi and T. pseudospiralis in muscle tissue with real-time PCR (2008)

    Art
    Zeitschriftenartikel / wissenschaftlicher Beitrag
    Autoren
    Guenther, Sebastian
    Nöckler, Karsten
    von Nickisch-Rosenegk, Markus
    Landgraf, Maria
    Ewers, Christa
    Wieler, Lothar H
    Schierack, Peter
    Quelle
    Journal of Microbiological Methods; 75(2) — S. 287–292
    ISSN: 0167-7012
    Sprache
    Englisch
    Verweise
    DOI: 10.1016/j.mimet.2008.06.019
    Pubmed: 18639594
    Kontakt
    Institut für Mikrobiologie und Tierseuchen

    Robert-von-Ostertag-Str. 7-13
    Gebäude 35
    14163 Berlin
    Tel.+49 30 83 8-518 40/518 43 Fax.+49 30 838 45 18 51
    email:mikrobiologie@vetmed.fu-berlin.de

    Abstract / Zusammenfassung

    Infections caused by Trichinella species occur throughout the world in many wild and domestic animals resulting in trichinellosis in men. In Europe, domestic pigs are predominantly infected by three Trichinella species: T. spiralis, T. britovi and T. pseudospiralis. Present methods for detection of Trichinella spp. (compressorium method, artificial digestion) do not always sufficiently recognize Trichinella larvae and these techniques are labor-intensive, time consuming and do not differentiate isolates on the species level since there are no distinguishing morphological features. Additionally, conventional PCRs cannot quantify numbers of larvae in infectious material. In order to better meet these requirements, we developed a real-time PCR assay for the accurate, rapid and specific identification of the three common European species of the genus Trichinella. The assay targets the large subunit of the mitochondrial rRNA (rrnL) and enables sensitive determination and discrimination of larvae in muscle tissue samples. The real-time PCR assay was developed and validated using reference and field strains from T. spiralis, T. britovi and T. pseudospiralis. In the described real-time PCR assay, the melting points of specific amplificates were always discernable via the melting curve from melting points of unspecific amplificates. This is important for the methods workflow because only C(T) values connected with the additional melting curve analysis allow a distinction of the individual species with confidence. The sensitivity of the technique enabled detection down to 0.1 Trichinella larva per gram meat sample. High disruption levels of tissues by mincing generally resulted in higher sensitivities than protocols without mincing. With its short completion time as well as accurate and specific detection of selected species this assay could become a convenient tool for the fast detection of Trichinella larvae in meat.