New method using quantitative PCR to follow the tick blood meal and to assess the anti-feeding effect of topical acaricide against Rhipicephalus sanguineus on dogs.

A 28-day study was conducted to assess the dynamic of blood feeding by Rhipicephalus sanguineus ticks on dogs treated or not with a novel topical combination of fipronil, amitraz and (S)-methoprene. Dogs were infested weekly through exposure to ticks in crates for 4h. Ticks were then counted in the crates at 2h and 4h post dog exposure. Ticks were also counted and removed from the dogs at 2h, 4h, 6h, 12h and 24h post tick exposure. The inhibition of blood feeding was assessed by both tick quantification and designing and performing a quantitative PCR (qPCR) to detect the canine hydroxymethylbilane synthase (HMBS) gene in ticks. The percentage of repellency sensu lato based on the ticks collected in crates at 2h varied from 4.7% at day 28 to 48.3% at day 7. The immediate mortality rate of the ticks expelled at 2h varied from 1.5% at day 21 to 31.7% at day 7. The efficacy calculation showed that the acaricidal combination started to kill ticks in as little as 2h. The average efficacy reached 90.0% at 12h post crate challenges and 100% at 24h post exposure in crates. The inclusion of an internal amplification control was used to ensure that no significant template-derived PCR inhibition (≤ 6.2%) affected the overall results. The reduction of blood feeding was significant at 4h (>80.0%) and >99.0% at 24h post tick exposure in the crate. The high repellency rate and the lethal efficacy of CERTIFECT(®) resulted in significantly fewer live attached ticks, consequently reducing blood intake and fluid exchanges.