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Frontline Tri-Act®) Against Haemaphysalis Longicornis Tick Infestation in Dogs Wilfried Lebon1* Julian Liebenberg2 Nadège Perier3 Doug Carithers4 Fréderic Beugnet3

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Frontline Tri-Act®) Against Haemaphysalis Longicornis Tick Infestation in Dogs Wilfried Lebon1* Julian Liebenberg2 Nadège Perier3 Doug Carithers4 Fréderic Beugnet3 Efficacy of a Single Spot-on Administration of Fipronil and Permethrin (Frontline Tri-act®) Against Haemaphysalis longicornis Tick Infestation in Dogs Wilfried Lebon1* Julian Liebenberg2 Nadège Perier3 Doug Carithers4 Fréderic Beugnet3 1Boehringer Ingelheim, Saint-Vulbas, France 2Clinvet, Universitas, Bloemfontein, South Africa 3Boehringer Ingelheim, Lyon, France 4Boehringer Ingelheim, Animal Health USA Inc., Duluth, USA *E-mail: [email protected] KEY WORDS: Haemaphysalis longicornis, formed at 24 hours post-infestations on Days fipronil, permethrin, Frontline Tri-Act, dog, 8, 15, 22, and 29, and ticks were removed efficacy. and counted at 48 hours post-treatment (on Day 2) and after each tick infestation on ABSTRACT Days 9, 16, 23, and 30. Acaricidal efficacies A blinded controlled laboratory study was for 24h thumb-counts were 100%, 98.8%, conducted to assess the acaricidal efficacy of 91.7%, and 84.4% on Days 8, 15, 22, and the fipronil/permethrin spot-on formulation 29, respectively. Efficacies of 98.1%, 100%, (Frontline Tri-Act®, Boehringer Ingelheim) 100%, 98.1%, and 94.7% were determined against Haemaphysalis longicornis ticks. based on removal counts performed on Days Fourteen healthy dogs were included in this 2, 9, 16, 23, and 30, respectively. There were study. Dogs in Group 1 served as untreated significant differences of the number of live controls while dogs in Group 2 were treated with Frontline Tri-Act on Day 0 at the ticks between treated and control group at minimum recommended dose of 0.1 mL/ each time-point (p <0.005). A single topical kg, corresponding to 6.76 mg/kg fipronil, administration of the combination of fipronil and 50.48 mg/kg permethrin. All dogs were plus permethrin was highly effective against experimentally infested with 50 viable unfed H. longicornis for four weeks. female H. longicornis ticks on days -2, 7, INTRODUCTION 14, 21, and 28. Thumb counts were per- Haemaphysalis longicornis, also called the Intern J Appl Res Vet Med • Vol. 17, No. 2, 2019. 93 ‘longhorn tick’ or ‘bush tick,’ is the most mosquitoes, stable flies, and sandflies under frequently encountered tick species on cattle laboratory conditions.23, 24 Moreover, its in Asia, New Zealand, Australia, and, the ability to reduce the risk of transmission of Pacific Islands,1, 2, 3 but is also commonly canine babesiosis and ehrlichiosis as well observed on dogs and cats in other countries as leishmaniosis has been demonstrated in like in Japan.4, 5 In 2017, it was first reported experimental studies.21,25 on a sheep in United States (New Jersey),6 Recently, the concomitant use of while retrospective tick collections revealed fipronil/permethrin (Frontline Tri-Act) with that populations of H. longicornis existed afoxolaner/milbemycin oxime (Nexgard as early as 2013.7 Now, the tick has been re- Spectra®) has shown to provide a safe and ported in several US states and is considered effective prevention strategy against both well established.8 endo- and ectoparasites, including vector- This tick species is characterized by borne diseases under field conditions.26 having both bisexual and parthenogenetic This study was conducted to assess populations.1, 3 Both males and females are the immediate and persistent efficacy of commonly observed in their native habitats Frontline Tri-Act against H. longicornis in all over Asia, whereas only parthenogenetic experimentally infested dogs. females are present in invaded countries like MATERIALS AND METHODS Australia, New Zealand, the Pacific Islands, This study was designed in accordance with and the US. Like other three-host ixodid the Committee for Medicinal Products for tick species, H.longicornis can infest a wide Veterinary use (CVMP) “Guideline for the range of wild and domestic animals, even testing and evaluation of the efficacy of though the adults have a tropism towards antiparasitic substances for the treatment wild and domestic ruminants. H. longicor- and prevention of tick and flea infesta- nis is known as a vector of several bacteria, tion in dogs and cats”, EMEA/CVMP/ protozoans, and viruses such as Anaplasma EWP/005/2000-Rev.3,27 as well as the spp.,9, 10 Ehrlichia spp.,11 Babesia gibsoni “World Association for the Advancement of (the agent of canine babesiosis in Asia),12 Veterinary Parasitology” (WAAVP) second Theileria orientalis (agent of cattle thei- edition guidelines for evaluating the efficacy leriosis),3 Flavivirus (causing tick-borne of parasiticides for the treatment, prevention encephalitis),13 Rickettsia japonica (causing and control of flea and tick infestation on Japanese spotted fever),14 and Phlebovirus dogs and cats.28 (causing severe fever with thrombocytope- nia syndrome/ SFTS).15 It was a parallel group, randomized, blinded, and controlled laboratory study Due to the medical and economic impor- which complied with the Good Clinical tance of H. longicornis, successful control Practices (GCP) as described in the Inter- of tick infestations is essential to reduce the national Cooperation on Harmonisation of risk of pathogen transmission and its geo- Technical Requirements for Registration of graphical expansion. Veterinary Medicinal Products (VICH).29 Frontline Tri-Act, a spot-on formula- The containment of the dogs complied with tion which combines 6.76 % w/v fipronil, the South African National Standard SANS and 50.48 % w/v permethrin, is effective 10386: latest edition “The care and use of for the treatment and prevention of flea animals for scientific purposes” and the infestations, including Ctenocephalides felis protocol was approved by the Institutional 16, 17 and Ctenocephalides canis, and ticks, Animal Care and Use Committee (IACUC) including Dermacentor reticulatus, Rhipi- prior to conduct the study. cephalus sanguineus (sensu lato) and Ixodes Animals ricinus.18, 19, 20, 21, 22 The product has shown repellent and insecticidal activities against Eighteen purpose-bred Mongrel and Beagle dogs were initially selected and acclimatized 94 Vol. 17, No.2, 2019 • Intern J Appl Res Vet Med. to the study conditions for 7 days. Dur- gicornis ticks on Day -7 for randomization ing the acclimation period, each dog was purposes, on Day -2 to assess immediate cu- examined by a veterinarian. Weighing of rative efficacy and on Days 7, 14, 21, and 28 all dogs and measuring of hair length were to assess sustained (i.e. persistent) efficacy. also performed. All dogs were confirmed The tick strain originated from a partheno- to be healthy. In addition, all dogs were genetic laboratory colony of H. longicornis dewormed prior to the initiation of the study (Okayama strain, Japanese origin). Tick and did not harbor resident ticks or fleas. infestations were performed by sedating the None of the dogs used in this study had been dogs with medetomidine, placing the se- treated with any long acting topical or sys- dated animals in an infestation chambers for temic acaricide/insecticide in the 12 weeks approximately 4 hours following tick infes- prior to the acclimation period. An initial H. tation. Ticks were released on multiple sites longicornis tick infestation was conducted of the dogs’ flank, being careful to avoid to determine individual tolerance of tick application sites. In addition, the light in the infestations and for random allocation to the room was turned off during the tick infesta- study groups. tion procedure to encourage attachment. After allocation, 14 healthy dogs, 8 On Day 0, dogs from Group 1 remained males and 6 females, aged of 13 months to untreated while dogs from Group 2 were 5.4 years and weighing 11.4 to 18.6 kg were treated once topically with Frontline Tri- included in the study. The animals were Act at the minimum effective dose of 0.1 kept individually in cages with visual and mL/kg (equivalent to 6.76 mg/kg fipronil auditory contact with conspecifics. At least bodyweight and 50.48 mg/kg permethrin) one toy was made available to each dog (re- according to label instructions. Dogs were plenished weekly). The dog cages were part observed hourly for four hours post treat- of an indoor animal unit, environmentally ment for any adverse reaction to treatment. controlled for temperature (20 °C ± 4 °C). On Day 2, all ticks were removed and Standard commercially available diet was counted to evaluate the curative efficacy of provided once a day at the recommended Frontline Tri-Act. manufacturer’s rates. Potable water was sup- In-situ tick thumb counts by palpation plied by the local municipality and provided and visual observation of the ticks onto the in stainless steel bowls and replenished at dogs were performed at 24 hours after each least twice daily. All the animals were ob- tick exposure on Days 8, 15, 22, and 29. served daily from Day -7 to Day 30 for their At 48 hours post-exposure, all ticks were general health. removed and counted using forceps. The Allocation to Treatment Groups numbers of live attached, live free, dead at- The study followed a randomized block tached, and dead free ticks were recorded. design based on pre-treatment tick count. Data Analysis Eighteen dogs were infested and the 14 dogs To evaluate the acaricidal efficacy, arith- with the highest live attached tick counts metic means of the live tick counts (free were ranked within sex in descending order and attached) were calculated by group at of individual pre-administration live, counts, each time-point. Percent effectiveness for and subsequently blocked into seven blocks the treated group was calculated using the of two dogs each. Within blocks, dogs were Abott formula [(C - T) / C] x 100, where C = randomly allocated to the two study groups: arithmetic mean for the control group and T Group 1, untreated control; Group 2 treated = arithmetic mean for the treated group. The with Frontline Tri-Act (fipronil, permethrin). groups were compared using an ANOVA Infestations, Treatment, and Tick Counts (Proc GLM procedure in SAS) with a treat- Each dog was experimentally infested with ment effect.
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