A new high-throughput analysis to screen Ehrlichia ruminantium in field ticks

Vachiéry Nathalie, Cangi Nidia, Pinarello Valérie, Bournez Laure, Lefrançois Thierry, Albina Emmanuel, Neves Luis. 2016. A new high-throughput analysis to screen Ehrlichia ruminantium in field ticks. In : Tropical animal diseases and veterinary public health: joining forces to meet future global challenges. AITVM, STVM, FAO. Berlin : AITVM, Résumé, 2 p. AITVM-STVM Joint International Conference, Berlin, Allemagne, 4 Septembre 2016/8 Septembre 2016.

Note générale : Mr Thierry Lefrançois est l'expert invité du Cirad

Résumé : In order to improve sample screening capacity of Ehrlichia ruminantium in ticks, an automatic DNA extraction method for Amblyomma ticks and a new QPCR targeting E. ruminantium pCS20 region, pCS20 Sol1 QPCR, were developed. A comparison between the new pCS20 Sol1 QPCR, a previously published pCS20 CowTM QPCR and the gold standard nested pCS20 PCR have been carried out. pCS20 Sol1TM QPCR was highly sensitive (up to 2.4 copies per sample) and specific (16 E. ruminantium strains detected and no cross-reaction with close-related species). We showed that Sol1 QPCR and pCS20 nested PCR had similar sensitivity and specificity however there was limited risk of contamination and timeless process for pCS20 Sol1 QPCR. In parallel, a tick 16SSyb rRNA QPCR was successfully developed for DNA quality control. DNA yield and quality from ticks extracted automatically, using 16SSyb rRNA QPCR were high. 16SSyb rRNA QPCR results (n=671 ticks) showed also a good reproducibility of automatic DNA extraction with a mean Ct=23+/-3. The whole method of screening, including automatic DNA extraction and pCS20 Sol1 QPCR, demonstrated a limit of detection between 6 and 0.6 copies per sample. It was also reproducible with restricted standard deviation of Ct (+/-1.3) for samples at the limit of detection. The development of a new automatic DNA extraction and QPCR allows for improving tick sample processing and E. ruminantium diagnostic capacities using high throughput methods. More widely, the automatic DNA extraction based on DNA/RNA virus kit and the new DNA quality control method, 16SSyb rRNA QPCR, can then be used for screening other bacterium and virus and for other tick species. (Texte intégral)

Mots-clés libres : Ehrlichia ruminantium, Amblyomma ticks, High throughput analysis

Classification Agris : L73 - Maladies des animaux
L72 - Organismes nuisibles des animaux

Auteurs et affiliations

• Vachiéry Nathalie, CIRAD-BIOS-UMR CMAEE (FRA)
• Cangi Nidia, CIRAD-BIOS-UMR CMAEE (GLP)
• Pinarello Valérie, CIRAD-BIOS-UMR CMAEE (GLP) ORCID: 0000-0002-9209-2111
• Bournez Laure, INRA (FRA)
• Lefrançois Thierry, CIRAD-BIOS-UMR CMAEE (FRA) ORCID: 0000-0001-8793-5228
• Albina Emmanuel, CIRAD-BIOS-UMR CMAEE (GLP)
• Neves Luis, Eduardo Mondlane University (MOZ)

Source : Cirad-Agritrop (https://agritrop.cirad.fr/581667/)

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