Comparison of four methods of extracting DNA from D. gallinae (Acari: Dermanyssidae)Sophie Desloire, Claire Valiente Moro, Claude Chauve and Lionel Zenner
Unité Mixte de Recherche PEV ENVL/INRA 958, Service de Parasitologie, École Nationale Vétérinaire de Lyon, 1 avenue Bourgelat, BP 83, 69280 Marcy l'Étoile, France
(Received 14 October 2005; accepted 31 March 2006 ; published online 6 July 2006)
Abstract - Dermanyssus gallinae is one of the most serious ectoparasites of poultry and it has been implicated as a vector of several major pathogenic diseases. Molecular detection of such pathogens in mites is crucial and therefore, an important step is the extraction of their DNA from mites. So, we compared four DNA extraction protocols from engorged and unfed individual mites: a conventional method using a Cethyl Trimethyl Ammonium Bromide buffer (CTAB), a Chelex resin, a Qiamp DNA extraction kit and a more recent one filter-based technology (FTA). The DNA samples have been tested for their ability to be amplified by an amplification of a D. gallinae 16S rRNA gene region. The best results were obtained using CTAB and Qiagen methods at the same time with unfed and engorged mites (96% and 100% of amplified samples). FTA produced similar results when using unfed mites but not when processing engorged ones (96% and 70%). Finally, the Chelex method was the least efficient in terms of DNA amplification, especially when applied on engorged individuals (50%). The possible inhibitor role of these Chelex extracted DNA was demonstrated by the means of a PCR control on PUC plasmid. No difference was observed with CTAB, Qiamp DNA extraction kit or FTA methods using DNA extracted one year before.
Key words: D. gallinae / DNA extraction / engorged mites
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© INRA, EDP Sciences 2006