Efficacy of a Combination Product containing Sarolaner, Moxidectin and Pyrantel (Simparica Trio ®) against induced infestations of Haemaphysalis longicornis on dogs

Efficacy of a Combination Product containing Sarolaner, Moxidectin and Pyrantel (Simparica Trio ®) against induced infestations of Haemaphysalis longicornis on dogs | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Efficacy of a Combination Product containing Sarolaner, Moxidectin and Pyrantel (Simparica Trio ®) against induced infestations of Haemaphysalis longicornis on dogs Kristina Kryda, Masaya Naito, Takeshi Fuji, Andrew Hodge, Steven Maeder This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5455294/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 26 Mar, 2025 Read the published version in Parasites & Vectors → Version 1 posted 9 You are reading this latest preprint version Abstract Background The efficacy of Simparica Trio ® (Zoetis), an oral combination product for dogs containing sarolaner, moxidectin, and pyrantel pamoate, was evaluated against Haemaphysalis longicornis , the most common tick species affecting dogs in the East Asia / Pacific region and an exotic species now reported from multiple states within the USA. Methods Two groups of 8 dogs each were ranked based on pretreatment tick counts and randomly allocated to treatment on Day 0 with either placebo or Simparica Trio at the minimum label dose of 1.2 mg/kg sarolaner, 24 µg/kg moxidectin, and 5 mg/kg pyrantel (as pamoate salt). Dogs were infested with 50 viable adult H. longicornis on Days − 2, 5, 12, 19, 26, and 33. Tick counts were conducted for all dogs 48 hours after treatment and subsequent re-infestations. Results A single oral administration of Simparica Trio was 98.9% effective in treating existing H. longicornis infestation when considering live, attached (feeding) ticks. Efficacy remained > 98% in preventing re-infestation for at least 35 days, and the geometric mean live, attached tick counts for Simparica Trio-treated dogs were significantly lower than for placebo-treated dogs ( P < 0.0001) at all time points assessed in the study. Geometric mean total live (attached and free) tick counts for dogs treated with Simparica Trio were also significantly lower ( P < 0.0001) than those for placebo-treated dogs at each time point. No adverse events were reported for any dogs throughout the duration of the study. Conclusions One dose of Simparica Trio at minimum label dose was highly efficacious in treating and controlling adult H. longicornis in dogs for more than one month. Asian longhorned tick Dog Efficacy Haemaphysalis longicornis Moxidectin Oral Combination Pyrantel pamoate Sarolaner Simparica Trio® Background The three-host tick Haemaphysalis longicornis (Acari: Ixodidae; Asian longhorned tick; cattle tick; bush tick) is prevalent throughout the Asia Pacific region, being native to Japan, Korea, eastern China, and southeast Russia and an established introduced species in Australia, New Zealand, and several islands in the Pacific [ 1 , 2 , 3 , 4 , 5 ]. In 2017, H. longicornis was also identified outside of quarantine in the USA, with large numbers of ticks recovered from a sheep in New Jersey [ 6 ]. Within 12 months, new populations were identified within the state of New Jersey as well as in New York, North Carolina, West Virginia, Virginia, and Arkansas [ 7 ], and retrospective analyses of collected samples revealed H. longicornis in West Virginia as early as 2010 [ 7 ]. The highly invasive tick species continues to spread across the USA and has now been confirmed in 19 states [ 8 ]. The proficiency with which H. longicornis is able to rapidly infiltrate and establish in new areas is strongly tied to its ability to feed on a wide range of mammalian and avian hosts, its parthenogenetic reproduction, and its toleration of a wide range of environmental temperatures [ 6 , 9 , 10 ]. Modeling suggests large areas of habitat around the world are suitable for the geographic expansion of H. longicornis , including countries in which the tick has never before been recorded [ 11 ]. Suitable habitats in North America extend along the East Coast from Arkansas-South Carolina to Southern Quebec-Nova Scotia and along the West Coast from California to British Columbia [ 12 , 13 ], and additional potential exists for establishment in Mexico and Central America [ 14 ], Africa, and most European countries [ 11 ]. The geographic expansion of tick species over the years has paralleled a global increase in the incidence of tick-borne diseases [ 15 ]. Pathogens known to be transmitted by H. longicornis include species of Anaplasma , Borrelia , Babesia , and Rickettsia , as well as the causative agents of theileriosis ( Theileria buffeli , T. orientalis, T. mutans ), and Q fever ( Coxiella burnetii ) [ 2 , 5 , 11 , 16 , 17 , 18 , 19 ]. In some areas of Asia, H. longicornis is thought to transmit the virus that causes severe fever with thrombocytopenia syndrome in humans [ 20 , 21 , 22 ], an emerging disease of considerable concern in China due to its increasing incidence and high mortality rate [ 21 ]. The invasive expansion of H. longicornis into new geographic locales brings with it the potential introduction of novel pathogens to local hosts. One apparent example of this is the situation facing some cattle producers in the USA, who previously dealt only with nonpathogenic native T. orientalis genotypes but now must manage disease and resulting economic loss caused by the T. orientalis Ikeda genotype transmitted by recently invasive H. longicornis populations [ 2 , 10 , 23 , 24 ]. While cattle are generally recognized as the dominant primary host for H. longicornis , additional hosts include other livestock (sheep, goats), companion animals (dogs, cats), wild mammals (deer, foxes, rabbits), and birds [ 1 , 3 , 6 , 10 , 25 , 26 , 27 , 28 ]. One review of recorded H. longicornis infestations reports 77 different host species across eight countries, including many animals native to each country affected [ 29 ]. Dogs positive for H. longicornis have been identified in China, Japan, South Korea, Australia, and the USA [ 11 ], with H. longicornis being the most frequently found tick species on dogs in Japan [ 28 ] and increasingly found on dogs in Australia [ 30 ]. In the USA, where H. longicornis is a relatively new invasive species, dogs comprise the majority (85%) of reported companion animal infestations [ 7 ]. That H. longicornis can transmit B. gibsoni , a protozoan parasite that is able to induce a spectrum of clinical symptoms in dogs [ 31 , 32 ], is well documented in parts of Asia [ 1 , 2 , 3 , 28 , 30 , 31 ]. In Australia, invasive H. longicornis has also been implicated in the spread of B. gibsoni , which was not identified in Australian dogs until 2002 [ 2 , 30 , 33 ]. Similarly, as the incidence of canine B. gibsoni infections in the USA continues to rise [ 34 , 35 ] it is not unreasonable to conjecture that invasive H. longicornis populations may become integrated in the life cycle of this pathogen. Protection of dogs against H. longicornis infestation and the subsequent transmission of disease agents requires a parasiticide that will act rapidly to interrupt infestation and subsequent feeding. Additionally, to promote the greatest possible owner compliance, a product should be easy to administer and possess sustained efficacy, preferably against a broad spectrum of target parasites [ 36 ]. The past ten years has seen the emergence of a series of novel parasiticides from the isoxazoline drug class highly effective in protecting dogs against multiple tick and flea species [ 37 , 38 , 39 , 40 ]. Sarolaner (Simparica®, Zoetis, NJ, USA), an isoxazoline approved for oral administration to dogs, has demonstrated efficacy against many tick species around the world [ 41 , 42 , 43 , 44 , 45 ], including H. longicornis [ 46 ]. Simparica Trio® is a monthly chewable preventative that combines sarolaner with moxidectin and pyrantel pamoate, allowing the convenient oral delivery of these three molecules from distinct drug classes as a single-dose means of treating and controlling important internal and external parasites in dogs [ 47 ]. The study reported here evaluated the efficacy of a single oral administration of Simparica Trio administered to dogs at the minimum label dose [1.2 mg/kg sarolaner, 24 µg/kg moxidectin, and 5 mg/kg pyrantel (as pamoate salt)] in treating an existing adult H. longicornis infestation and preventing weekly re-infestations for a period of 5 weeks following a single dose. Methods The placebo-controlled, randomized, laboratory comparative efficacy study was conducted by Shokukanken Inc. (Gunma, Japan). All study procedures were in accordance with the World Association for the Advancement of Veterinary Parasitology guidelines for evaluating the efficacy of parasiticides for the treatment, prevention, and control of flea and tick infestation on dogs and cats [ 48 ]. Animals Uniquely identified purpose-bred Beagle dogs (8 males, 8 females) aged 21 to 25 weeks and weighing 5.6 to 8.0 kg were included in the study. At enrollment, a veterinarian examined all dogs to ensure they were in good health and suitable for inclusion in the study. Dogs underwent an adequate wash-out period to ensure that no residual tick efficacy remained from any previously administered treatments. Animals were housed in individual indoor enclosures (18 to 29 ºC) that conformed to accepted animal welfare guidelines while ensuring no direct contact. All dogs were acclimatized to these conditions for at least 10 days prior to the study starting on Day-7. Throughout the study, dogs were fed an appropriate ration of a commercial dry laboratory canine feed and were given water ad libitum . General health observations were performed daily for each dog starting on Day − 17 through the duration of the study. Design A randomized complete block design was employed. Host suitability was determined prior to inclusion in the study by infesting dogs on Day − 7 with 50 viable, unfed adult H. longicornis ticks. After 48 ± 2 hours, live attached ticks were counted and removed, and 16 eligible dogs were ranked into blocks of two based on pre-treatment tick counts. Two dogs within each block were then randomly allocated to treatment with either placebo or Simparica Trio with each treatment group containing 8 dogs. Dogs were weighed and infested with 50 viable adult ticks on Day − 2. On Day 0, all dogs were assessed for overall health and then orally administered either the placebo or Simparica Trio. Tick infestations of 50 viable H. longicornis adults were repeated on Days 5, 12, 19, 26, and 33, and live tick counts were performed 48 ± 2 hours after treatment on Day 0 and again after each infestation (on Days 7, 14, 21, 28, and 35). Treatment All dogs were treated on Day 0 with either placebo tablets (inert formulation ingredients) or with Simparica Trio chewable tablets, with doses calculated using body weights recorded on Day − 2. Each dog in the Simparica Trio group received either one or two tablets of the combination product to provide as close as possible to the minimum label dosage of 1.2 mg/kg sarolaner, 24 µg/kg moxidectin and 5 mg/kg pyrantel (as pamoate salt). Dogs in the placebo group received the equivalent number of placebo tablets. Placebo and Simparica Trio tablet presentations were similar in appearance, and doses were administered by hand to ensure accurate pilling. All dogs were observed for several minutes after dosing to confirm that the dose was swallowed. Dogs were not fed within 12 hours prior to dosing and for at least 4 hours after treatment. Tick infestation and assessment The study used viable, unfed adult H. longicornis ticks sourced from a laboratory colony first established at the laboratories of Shokukanken within one year of the study being conducted using wild, engorged ticks collected in the Gunma Prefecture, Japan. At each infestation, H. longicornis adult ticks were placed on the auricle to mimic the natural preferred attachment sites. To facilitate infestation, dogs were sedated with xylazine hydrochloride (2% injectable formulation) and were also fitted with Elizabethan collars to reduce grooming for the duration of each infestation period. To ensure the dogs were acceptable hosts and for allocation purposes, dogs were first infested on Day − 7 and live, attached ticks were counted on Day − 5. To determine efficacy against an existing infestation, dogs were then infested on Day − 2 (2 days before treatment on Day 0) and tick counts were conducted on Day 2. Subsequent weekly infestations were conducted on Days 5, 12, 19, 26 and 33, and counts were performed 48 ± 2 hours after each infestation (on Days 7, 14, 21, 28 and 35) to confirm the duration of preventative efficacy. Tick counts Dogs were sedated with xylazine hydrochloride (2% injectable formulation) at each tick count. Counts were conducted by trained personnel by systematically inspecting from head to tail while parting the hair by hand, with observed ticks being counted and removed. The dogs were then combed thoroughly with a fine-toothed comb. Each dog was examined for a minimum of 10 minutes, and if ticks were found in the final minute of counting, the examination was continued in 1-minute increments until no further ticks were found. All live and dead ticks were classified as either (1) unattached or free; (2) attached but un-engorged; or (3) attached and engorged. Personnel conducting counts were masked to treatment assignments and changed gloves between dogs. Statistical analysis The experimental unit was the individual dog. Tick counts were transformed using the log e (count + 1) transformation prior to analysis to decrease variance and normalize the data. Transformed counts were analyzed according to a mixed linear model for repeated measures using the PROC MIXED procedure (SAS 9.4, Cary NC). The fixed effects of the model were treatment, time point and the interaction between treatment and time point. The random effects included block, animal, and error. Two-sided testing was performed at the significance level of α = 0.05. Efficacy was evaluated separately for (i) total live tick counts (free and attached ticks) and (ii) live, attached tick counts. Efficacy assessment was based on the percent reduction in the arithmetic and geometric mean live tick counts for Simparica Trio-treated dogs relative to those for placebo-treated dogs using Abbott’s formula: % reduction = 100 × [mean count (placebo) – mean count (treated)]/mean count (placebo). Results and Discussion The sixteen dogs enrolled in the study proved to be acceptable hosts for H. longicornis , demonstrating a geometric mean (arithmetic mean; range) adult live tick count of 17.1 (17.4; range 13 to 23) on Day − 5. All eight dogs treated with the placebo maintained robust adult tick infestations for the duration of the study. On Days 2, 7, 14, 21, 28, and 35 geometric mean live (free and attached) adult tick counts were 29.6, 26.8, 31.3, 27.9, 29.0, and 27.5, respectively, and arithmetic mean live (free and attached) tick counts were 29.8, 27.0, 31.5, 28.1, 29.0, and 27.5, respectively. More than 50% (range 51.1 to 63.3%) of the live adult ticks observed on placebo-treated dogs at each time point were attached. On Days 2, 7, 14, 21, 28, and 35 geometric mean live, attached adult tick counts were 16.6, 16.8, 15.7, 15.8, 17.4, and 16.6, respectively, and arithmetic mean live, attached tick counts were 16.9, 17.1, 16.1, 16.0, 17.6, and 16.8, respectively. A few dead ticks (0 to 2) were found on placebo-treated dogs at each time point. Actual administered doses of Simparica Trio ranged from 1.23 to 1.58 mg/kg sarolaner, 25 to 32 µg/kg moxidectin, and 5.14 to 6.58 mg/kg pyrantel (as pamoate salt). The single oral dose of Simparica Trio was well tolerated by all eight treated dogs, with no adverse reactions to treatment recorded during daily general health observations. On Days 2, 7, 14, 21, 28, and 35 post-treatment, arithmetic mean live (free and attached) tick counts in Simparica Trio-treated dogs were 8.8, 11.9, 9.1, 7.9, 10.9, and 7.1, respectively, and geometric mean live (free and attached) tick counts were 8.0, 11.6, 8.8, 7.3, 10.5, and 6.3, respectively. The eight dogs treated with Simparica Trio showed significantly reduced ( P < 0.0001) geometric mean live (free and attached) adult tick counts compared to placebo-treated dogs at each time point examined (5.64 ≤ t(50.8) ≤ 9.62, P < 0.0001). In these same Simparica Trio-treated dogs, the number of killed adult ticks collected from each dog at each timepoint ranged from 14 to 27 (28 to 54% of the infestation). Almost all (98%) live adult ticks recovered from Simparica Trio-treated dogs were unattached, with geometric (arithmetic) mean live, attached tick counts for Simparica Trio-treated dogs being ≤ 0.3 (≤ 0.4) at each time point (Table 1 ). A total of only nine live, attached adult ticks were recovered from the Simparica Trio-treated dogs over the 35 days: two dogs never had live, attached ticks; three dogs were observed to have a single live, attached tick at one time point; and three dogs were observed to have a single live, attached tick at two different time points. Table 1 Geometric (arithmetic) mean live, attached Haemaphysalis longicornis counts and ranges for dogs treated orally (Day 0) with placebo or Simparica Trio chewable tablets and percent efficacy relative to placebo at 48 hours after treatment and subsequent weekly re-infestations Placebo Simparica Trio 1 Percent Efficacy 2 Day Geometric Mean (Arithmetic mean) Range No. of Dogs with ticks Geometric Mean (Arithmetic mean) Range No of Dogs with ticks 2 16.6 (16.9) 12–22 8/8 0.2 (0.3) 0–1 2/8 98.9 (98.5) 7 16.8 (17.1) 12–22 8/8 0.1 (0.1) 0–1 1/8 99.5 (99.3) 14 15.7 (16.1) 11–21 8/8 0.2 (0.3) 0–1 2/8 98.8 (98.5) 21 15.8 (16.0) 13–19 8/8 0.0 (0.0) 0–0 0/8 100 (100) 28 17.4 (17.6) 13–21 8/8 0.3 (0.4) 0–1 3/8 98.3 (97.9) 35 16.6 (16.8) 13–20 8/8 0.1 (0.1) 0–1 1/8 99.5 (99.3) 1 Treatment with placebo or Simparica Trio (minimum dose 1.2 mg/kg sarolaner, 24 µg/kg moxidectin, 5 mg/kg pyrantel (as pamoate salt)) occurred on Day 0. 2 Geometric (arithmetic) efficacy of Simparica Trio versus Placebo. Geometric mean live, attached tick counts significantly lower compared to placebo at each timepoint (22.66 ≤ t(71.6) ≤ 24.18, P < 0.0001) It is logical to assume that live, unattached adult ticks recovered from dogs during this study were not feeding and thus had not been exposed to Simparica Trio. Therefore, efficacy of Simparica Trio against H. longicornis was calculated using the live, attached adult tick counts recorded for dogs in each treatment group (Table 1 ). In this study, geometric mean counts of live, attached adult ticks for Simparica Trio-treated dogs were significantly lower at each time point compared with placebo-treated dogs (22.66 ≤ t(71.6) ≤ 24.18, P < 0.0001, Table 1 ). One treatment with Simparica Trio provided dogs with 98.9% efficacy against existing adult H. longicornis infestations and was ≥ 98.3% effective in protecting dogs against re-infestation with adult H. longicornis for at least 35 days after treatment. The impacts of tick infestations on dogs are well known [ 49 , 50 ], and the negative consequences of H. longicornis infestation have been documented across multiple host species, ranging from irritation and anemia to diarrhea, production losses, and even death [ 2 , 10 ]. Laboratory studies on H. longicornis have demonstrated that all life stages are able to feed on dogs, with mean larval, nymphal, and adult feeding periods similar to those recorded when feeding on cattle [ 2 , 51 , 52 ]. Additionally, environmental sampling has recovered host-seeking, engorged H. longicornis , providing evidence for repeated successful questing events and highlighting the role this tick species may play in the spread of vector-borne agents [ 53 ]. The need for an effective means of treating and controlling H. longicornis is apparent considering its importance as a vector of human and animal disease agents, the negative impacts of infestation, the species’ wide host range, and its invasive ability. Previous studies show that a single oral dose of sarolaner can rapidly and effectively control multiple canine tick species [ 41 , 42 , 44 , 45 , 54 , 55 , 56 ], including H. longicornis [ 46 ]. Here, we show sarolaner administered in a combination product, as Simparica Trio, remained highly effective in rapidly treating existing adult attached, live H. longicornis infestations in dogs and in preventing re-infestation for at least 35 days. This level of protection provides interruption of the H. longicornis life cycle and a reduction in the likelihood of pathogen transmission between hosts. When administering traditional pill or tablet medications, many dog owners worry about inaccurate dosing and damaging the animal-human bond during unsuccessful or stressful pilling events [ 57 ]. In contrast, dog owners generally view chewable medications positively due to their increased ease of administration and formulation palatability. The Simparica Trio chewable tablet has been shown to be well-accepted by dogs, with one study reporting the majority (91.9%) of 517 doses offered to dogs by owners in their own homes taken by free choice without food or in food (and only 8.1% of tablets requiring pilling) [ 58 ]. Moreover, as a combination product containing both moxidectin and pyrantel in addition to sarolaner, Simparica Trio provides dog owners seeking month-long efficacy against ticks and fleas the additional benefits of effective treatment for roundworm and hookworm infections and protection against lungworm and heartworm disease [ 56 , 58 , 59 , 60 , 61 ]. When one considers the important roles of palatability, ease of administration, and broad-spectrum protection in owner compliance [ 36 , 57 , 62 ], it is clear that Simparica Trio makes an important contribution in the field of canine health by providing comprehensive protection in a single convenient product. Conclusions A single oral dose of the combination antiparasiticide Simparica Trio administered at the minimum label dose of 1.2 mg/kg sarolaner, 24 µg/kg moxidectin, and 5 mg/kg pyrantel (as pamoate salt) effectively treated an existing infestation of H. longicornis adults and prevented re-infestation for up to 35 days. Simparica Trio is an easy and convenient means of providing dogs with rapid protection against many important tick species. Abbreviations US: United States of America Declarations Ethics Approval and Consent to Participate The protocol was reviewed and approved by the Institutional Animal Care and Use Committee at Shokukanken, Inc. prior to implementation. Consent for Publication Not applicable Availability of Data and Materials The dataset supporting the conclusions of this article are included within the article. Competing Interests This study was funded by Zoetis Japan. KK, TF, AH, and SM were employees of Zoetis at the time of the study. 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McTier TL, Six RH, Fourie JJ, Pullins A, Hedges L, Mahabir SP, et al. Determination of the effective dose of a novel oral formulation of sarolaner (Simparica™) for the treatment and month-long control of fleas and ticks on dogs. Vet Parasitol. 2016;222:12-7; doi: 10.1016/j.vetpar.2016.02.016. Six RH, Everett WR, Young DR, Carter L, Mahabir SP, Honsberger NA, et al. Efficacy of a novel oral formulation of sarolaner (Simparica TM ) against five common tick species infesting dogs in the United States. Vet Parasitol. 2016;222:28-32; doi: 10.1016/j.vetpar.2015.12.023. Geurden T, Becskei C, Grace S, Strube C, Doherty P, Liebenberg J, et al. Efficacy of a novel oral formulation of sarolaner (Simparica TM ) against four common tick species infesting dogs in Europe. Vet Parasitol. 2016;222:33-6; doi: 10.1016/j.vetpar.2016.03.024. Becskei C, De Bock F, Illambas J, Mahabir SP, Farkas R, Six RH. Efficacy and safety of a novel oral isoxazoline, sarolaner (Simparica™) in the treatment of naturally occurring flea and tick infestations in dogs presented as veterinary patients in Europe. Vet Parasitol. 2016;222:49-55; doi: 10.1016/j.vetpar.2016.02.007. Scott F, Franz L, Campos DR, Azevedo TRC, Cunha D, Six RH, et al. Efficacy of sarolaner (Simparic) against induced infestations of Amblyomma cajennense on dogs. Parasit Vectors. 2017;10 1:390; doi: 10.1186/s13071-017-2324-0. Oda K, Yonetake W, Fujii T, Hodge A, Six RH, Maeder S, et al. Efficacy of sarolaner (Simparica®) against induced infestations of Haemaphysalis longicornis on dogs. Parasit Vectors. 2019;12:509; doi: 10.1186/s13071-019-3765-4. https://doi.org/10.1186/s13071-019-3765-4. Zoetis: Simparica Trio® (sarolaner, moxidectin and pyrantel pamoate chewable tablets) package insert. https://www.zoetisus.com/content/_assets/docs/Petcare/simparica-trio-pi.pdf: Zoetis Inc; 2022. Marchiondo AA, Holdsworth PA, Fourie LJ, Rugg D, Hellmann K, Snyder DE, et al. World Association for the Advancement of Veterinary Parasitology (W.A.A.V.P.) second edition: guidelines for evaluating the efficacy of parasiticides for the treatment, prevention and control of flea and tick infestations on dogs and cats. Vet Parasitol. 2013;194 1:84-97. Blagburn BL, Dryden MW. Biology, treatment, and control of flea and tick infestations. Vet Clin North Am Small Anim Pract. 2009;39:1173-200; doi: 10.1016/j.cvsm.2009.07.001. Berrada ZL, Telford SR, 3rd. Burden of tick-borne infections on American companion animals. Top Companion Anim Med. 2009;24:175-81; doi: 10.1053/j.tcam.2009.06.005. Marendy D, Baker K, Emery D, Rolls P, Stutchbury R. Haemaphysalis longicornis : the life-cycle on dogs and cattle, with confirmation of its vector status for Theileria orientalis in Australia. Vet Parasitol. 2020;277s:100022; doi: 10.1016/j.vpoa.2019.100022. Liu JungZe LJ, Jiang ZaiJie JZ. Studies on the bionomics of Haemaphysalis longicornis Neumann (Acari: Ixodidae) under laboratory conditions. Acta Entomol Sinica. 1998;41 3:280-3. https://search.ebscohost.com/login.aspx?direct=true&db=lah&AN=19980505603&site=eds-live. Price KJ, Witmier BJ, Eckert RA, Boyer CN. Recovery of Partially Engorged Haemaphysalis longicornis (Acari: Ixodidae) Ticks from Active Surveillance. J Med Entomol. 2022;59 5:1842-6; doi: 10.1093/jme/tjac099. Packianathan R, Hodge A, Bruellke N, Jackson C, Maeder S. Efficacy of combination products containing sarolaner, moxidectin and pyrantel (Simparica Trio™) or afoxolaner and milbemycin (NexGard Spectra®) against induced infestations of Ixodes holocyclus in dogs. Parasit Vectors. 2020;13 1:448; doi: 10.1186/s13071-020-04323-8. https://doi.org/10.1186/s13071-020-04323-8. Becskei C, Liebenberg J, Thys M, Mahabir SP. Efficacy of a novel chewable tablet containing sarolaner, moxidectin and pyrantel (Simparica Trio™) against four common tick species infesting dogs in Europe. Parasit Vectors. 2020;13 1:100; doi: 10.1186/s13071-020-3949-y. https://doi.org/10.1186/s13071-020-3949-y. Kryda K, Mahabir SP, Chapin S, Holzmer SJ, Bowersock L, Everett WR, et al. Efficacy of a novel orally administered combination product containing sarolaner, moxidectin and pyrantel (Simparica Trio™) against induced infestations of five common tick species infesting dogs in the USA. Parasit Vectors. 2020;13:77; doi: 10.1186/s13071-020-3945-2. Wright A, Hillier A, Lambert J, Mwacalimba K, Lloyd N, Kagiwada T, et al. Dog Owners' Perceptions of the Convenience and Value of Chewable Oclacitinib: Quantitative Survey Data from an International Survey. Animals (Basel). 2024;14 6; doi: 10.3390/ani14060952. Kryda K, Mahabir SP, Inskeep T, Rugg J. Safety and efficacy of a novel oral chewable combination tablet containing sarolaner, moxidectin and pyrantel (Simparica Trio™) against natural flea infestations in client-owned dogs in the USA. Parasit Vectors. 2020;13 1:98; doi: 10.1186/s13071-020-3952-3. https://doi.org/10.1186/s13071-020-3952-3. Kryda K, Six RH, Walsh KF, Holzmer SJ, Chapin S, Mahabir SP, et al. Laboratory and field studies to investigate the efficacy of a novel, orally administered combination product containing moxidectin, sarolaner and pyrantel for the prevention of heartworm disease ( Dirofilaria immitis ) in dogs. Parasit Vectors. 2019;12 1:445; doi: 10.1186/s13071-019-3702-6. https://doi.org/10.1186/s13071-019-3702-6. Becskei C, Kryda K, Thys M, Holzmer S, Bowersock L, Fernandes T, et al. Efficacy of a new oral chewable tablet containing sarolaner, moxidectin and pyrantel (Simparica Trio™) against induced ascarid infections in dogs. Parasit Vectors. 2020;13 1:71; doi: 10.1186/s13071-020-3950-5. Becskei C, Thys M, Kryda K, Meyer L, Martorell S, Geurden T, et al. Efficacy of Simparica Trio™, a novel chewable tablet containing sarolaner, moxidectin and pyrantel, against induced hookworm infections in dogs. Parasit Vectors. 2020;13 1:99; doi: 10.1186/s13071-020-3951-4. Thombre AG. Oral delivery of medications to companion animals: palatability considerations. Adv Drug Deliv Rev. 2004;56 10:1399-413; doi: https://doi.org/10.1016/j.addr.2004.02.012. https://www.sciencedirect.com/science/article/pii/S0169409X04000729. Additional Declarations Competing interest reported. This study was funded by Zoetis Japan. KK, TF, AH, and SM were employees of Zoetis at the time of the study. MN was the contracted study investigator. Cite Share Download PDF Status: Published Journal Publication published 26 Mar, 2025 Read the published version in Parasites & Vectors → Version 1 posted Editorial decision: Revision requested 13 Dec, 2024 Reviews received at journal 11 Dec, 2024 Reviews received at journal 09 Dec, 2024 Reviewers agreed at journal 19 Nov, 2024 Reviewers agreed at journal 18 Nov, 2024 Reviewers invited by journal 18 Nov, 2024 Editor assigned by journal 16 Nov, 2024 Submission checks completed at journal 16 Nov, 2024 First submitted to journal 14 Nov, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5455294","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":390106744,"identity":"a6a73864-4be4-4c22-9e1b-f3db77688d9b","order_by":0,"name":"Kristina Kryda","email":"data:image/png;base64,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","orcid":"","institution":"Zoetis, Veterinary Medicine Research and Development","correspondingAuthor":true,"prefix":"","firstName":"Kristina","middleName":"","lastName":"Kryda","suffix":""},{"id":390106745,"identity":"ebe8d589-43ad-4ec3-ae56-bf08488363bf","order_by":1,"name":"Masaya Naito","email":"","orcid":"","institution":"Shokukanken Inc","correspondingAuthor":false,"prefix":"","firstName":"Masaya","middleName":"","lastName":"Naito","suffix":""},{"id":390106746,"identity":"00f906b6-4077-4ed3-b813-60de4efac56b","order_by":2,"name":"Takeshi Fuji","email":"","orcid":"","institution":"Zoetis Japan KK","correspondingAuthor":false,"prefix":"","firstName":"Takeshi","middleName":"","lastName":"Fuji","suffix":""},{"id":390106747,"identity":"94fa2153-a81b-42d8-b930-4f514b1f4aef","order_by":3,"name":"Andrew Hodge","email":"","orcid":"","institution":"Zoetis Australia Research and Manufacturing","correspondingAuthor":false,"prefix":"","firstName":"Andrew","middleName":"","lastName":"Hodge","suffix":""},{"id":390106748,"identity":"c97150d3-6e35-4387-9980-8c4d1570ced0","order_by":4,"name":"Steven Maeder","email":"","orcid":"","institution":"Zoetis, Veterinary Medicine Research and Development","correspondingAuthor":false,"prefix":"","firstName":"Steven","middleName":"","lastName":"Maeder","suffix":""}],"badges":[],"createdAt":"2024-11-14 16:08:18","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5455294/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5455294/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s13071-025-06747-6","type":"published","date":"2025-03-26T15:57:07+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":79605634,"identity":"026956e1-745d-4bb2-90c3-5621259f7601","added_by":"auto","created_at":"2025-03-31 16:11:16","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":716950,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5455294/v1/e473e63e-d0c6-4660-9212-6897ff716c52.pdf"}],"financialInterests":"Competing interest reported. This study was funded by Zoetis Japan. KK, TF, AH, and SM were employees of Zoetis at the time of the study. MN was the contracted study investigator.","formattedTitle":"Efficacy of a Combination Product containing Sarolaner, Moxidectin and Pyrantel (Simparica Trio ®) against induced infestations of Haemaphysalis longicornis on dogs","fulltext":[{"header":"Background","content":"\u003cp\u003eThe three-host tick \u003cem\u003eHaemaphysalis longicornis\u003c/em\u003e (Acari: Ixodidae; Asian longhorned tick; cattle tick; bush tick) is prevalent throughout the Asia Pacific region, being native to Japan, Korea, eastern China, and southeast Russia and an established introduced species in Australia, New Zealand, and several islands in the Pacific [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. In 2017, \u003cem\u003eH. longicornis\u003c/em\u003e was also identified outside of quarantine in the USA, with large numbers of ticks recovered from a sheep in New Jersey [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Within 12 months, new populations were identified within the state of New Jersey as well as in New York, North Carolina, West Virginia, Virginia, and Arkansas [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e], and retrospective analyses of collected samples revealed \u003cem\u003eH. longicornis\u003c/em\u003e in West Virginia as early as 2010 [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. The highly invasive tick species continues to spread across the USA and has now been confirmed in 19 states [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The proficiency with which \u003cem\u003eH. longicornis\u003c/em\u003e is able to rapidly infiltrate and establish in new areas is strongly tied to its ability to feed on a wide range of mammalian and avian hosts, its parthenogenetic reproduction, and its toleration of a wide range of environmental temperatures [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Modeling suggests large areas of habitat around the world are suitable for the geographic expansion of \u003cem\u003eH. longicornis\u003c/em\u003e, including countries in which the tick has never before been recorded [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. Suitable habitats in North America extend along the East Coast from Arkansas-South Carolina to Southern Quebec-Nova Scotia and along the West Coast from California to British Columbia [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e], and additional potential exists for establishment in Mexico and Central America [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e], Africa, and most European countries [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe geographic expansion of tick species over the years has paralleled a global increase in the incidence of tick-borne diseases [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. Pathogens known to be transmitted by \u003cem\u003eH. longicornis\u003c/em\u003e include species of \u003cem\u003eAnaplasma\u003c/em\u003e, \u003cem\u003eBorrelia\u003c/em\u003e, \u003cem\u003eBabesia\u003c/em\u003e, and \u003cem\u003eRickettsia\u003c/em\u003e, as well as the causative agents of theileriosis (\u003cem\u003eTheileria buffeli\u003c/em\u003e, \u003cem\u003eT. orientalis, T. mutans\u003c/em\u003e), and Q fever (\u003cem\u003eCoxiella burnetii\u003c/em\u003e) [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. In some areas of Asia, \u003cem\u003eH. longicornis\u003c/em\u003e is thought to transmit\u003c/p\u003e \u003cp\u003ethe virus that causes severe fever with thrombocytopenia syndrome in humans [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e], an emerging disease of considerable concern in China due to its increasing incidence and high mortality rate [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. The invasive expansion of \u003cem\u003eH. longicornis\u003c/em\u003e into new geographic locales brings with it the potential introduction of novel pathogens to local hosts. One apparent example of this is the situation facing some cattle producers in the USA, who previously dealt only with nonpathogenic native \u003cem\u003eT. orientalis\u003c/em\u003e genotypes but now must manage disease and resulting economic loss caused by the \u003cem\u003eT. orientalis\u003c/em\u003e Ikeda genotype transmitted by recently invasive \u003cem\u003eH. longicornis\u003c/em\u003e populations [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eWhile cattle are generally recognized as the dominant primary host for \u003cem\u003eH. longicornis\u003c/em\u003e, additional hosts include other livestock (sheep, goats), companion animals (dogs, cats), wild mammals (deer, foxes, rabbits), and birds [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e, \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. One review of recorded \u003cem\u003eH. longicornis\u003c/em\u003e infestations reports 77 different host species across eight countries, including many animals native to each country affected [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]. Dogs positive for \u003cem\u003eH. longicornis\u003c/em\u003e have been identified in China, Japan, South Korea, Australia, and the USA [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e], with \u003cem\u003eH. longicornis\u003c/em\u003e being the most frequently found tick species on dogs in Japan [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e] and increasingly found on dogs in Australia [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. In the USA, where \u003cem\u003eH. longicornis\u003c/em\u003e is a relatively new invasive species, dogs comprise the majority (85%) of reported companion animal infestations [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. That \u003cem\u003eH. longicornis\u003c/em\u003e can transmit \u003cem\u003eB. gibsoni\u003c/em\u003e, a protozoan parasite that is able to induce a spectrum of clinical symptoms in dogs [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e, \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e], is well documented in parts of Asia [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e, \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. In Australia, invasive \u003cem\u003eH. longicornis\u003c/em\u003e has also been implicated in the spread of \u003cem\u003eB. gibsoni\u003c/em\u003e, which was not identified in Australian dogs until 2002 [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e]. Similarly, as the incidence of canine \u003cem\u003eB. gibsoni\u003c/em\u003e infections in the USA continues to rise [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e] it is not unreasonable to conjecture that invasive \u003cem\u003eH. longicornis\u003c/em\u003e populations may become integrated in the life cycle of this pathogen.\u003c/p\u003e \u003cp\u003eProtection of dogs against \u003cem\u003eH. longicornis\u003c/em\u003e infestation and the subsequent transmission of disease agents requires a parasiticide that will act rapidly to interrupt infestation and subsequent feeding. Additionally, to promote the greatest possible owner compliance, a product should be easy to administer and possess sustained efficacy, preferably against a broad spectrum of target parasites [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. The past ten years has seen the emergence of a series of novel parasiticides from the isoxazoline drug class highly effective in protecting dogs against multiple tick and flea species [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e, \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e, \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e, \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. Sarolaner (Simparica\u0026reg;, Zoetis, NJ, USA), an isoxazoline approved for oral administration to dogs, has demonstrated efficacy against many tick species around the world [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e, \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e, \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e, \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e, \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e], including \u003cem\u003eH. longicornis\u003c/em\u003e [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e]. Simparica Trio\u0026reg; is a monthly chewable preventative that combines sarolaner with moxidectin and pyrantel pamoate, allowing the convenient oral delivery of these three molecules from distinct drug classes as a single-dose means of treating and controlling important internal and external parasites in dogs [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e]. The study reported here evaluated the efficacy of a single oral administration of Simparica Trio administered to dogs at the minimum label dose [1.2 mg/kg sarolaner, 24 \u0026micro;g/kg moxidectin, and 5 mg/kg pyrantel (as pamoate salt)] in treating an existing adult \u003cem\u003eH. longicornis\u003c/em\u003e infestation and preventing weekly re-infestations for a period of 5 weeks following a single dose.\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003eThe placebo-controlled, randomized, laboratory comparative efficacy study was conducted by Shokukanken Inc. (Gunma, Japan). All study procedures were in accordance with the World Association for the Advancement of Veterinary Parasitology guidelines for evaluating the efficacy of parasiticides for the treatment, prevention, and control of flea and tick infestation on dogs and cats [\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e].\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eAnimals\u003c/h2\u003e \u003cp\u003eUniquely identified purpose-bred Beagle dogs (8 males, 8 females) aged 21 to 25 weeks and weighing 5.6 to 8.0 kg were included in the study. At enrollment, a veterinarian examined all dogs to ensure they were in good health and suitable for inclusion in the study. Dogs underwent an adequate wash-out period to ensure that no residual tick efficacy remained from any previously administered treatments. Animals were housed in individual indoor enclosures (18 to 29 \u0026ordm;C) that conformed to accepted animal welfare guidelines while ensuring no direct contact. All dogs were acclimatized to these conditions for at least 10 days prior to the study starting on Day-7. Throughout the study, dogs were fed an appropriate ration of a commercial dry laboratory canine feed and were given water \u003cem\u003ead libitum\u003c/em\u003e. General health observations were performed daily for each dog starting on Day \u0026minus;\u0026thinsp;17 through the duration of the study.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eDesign\u003c/h3\u003e\n\u003cp\u003eA randomized complete block design was employed. Host suitability was determined prior to inclusion in the study by infesting dogs on Day \u0026minus;\u0026thinsp;7 with 50 viable, unfed adult \u003cem\u003eH. longicornis\u003c/em\u003e ticks. After 48\u0026thinsp;\u0026plusmn;\u0026thinsp;2 hours, live attached ticks were counted and removed, and 16 eligible dogs were ranked into blocks of two based on pre-treatment tick counts. Two dogs within each block were then randomly allocated to treatment with either placebo or Simparica Trio with each treatment group containing 8 dogs. Dogs were weighed and infested with 50 viable adult ticks on Day \u0026minus;\u0026thinsp;2. On Day 0, all dogs were assessed for overall health and then orally administered either the placebo or Simparica Trio. Tick infestations of 50 viable \u003cem\u003eH. longicornis\u003c/em\u003e adults were repeated on Days 5, 12, 19, 26, and 33, and live tick counts were performed 48\u0026thinsp;\u0026plusmn;\u0026thinsp;2 hours after treatment on Day 0 and again after each infestation (on Days 7, 14, 21, 28, and 35).\u003c/p\u003e\n\u003ch3\u003eTreatment\u003c/h3\u003e\n\u003cp\u003eAll dogs were treated on Day 0 with either placebo tablets (inert formulation ingredients) or with Simparica Trio chewable tablets, with doses calculated using body weights recorded on Day \u0026minus;\u0026thinsp;2. Each dog in the Simparica Trio group received either one or two tablets of the combination product to provide as close as possible to the minimum label dosage of 1.2 mg/kg sarolaner, 24 \u0026micro;g/kg moxidectin and 5 mg/kg pyrantel (as pamoate salt). Dogs in the placebo group received the equivalent number of placebo tablets. Placebo and Simparica Trio tablet presentations were similar in appearance, and doses were administered by hand to ensure accurate pilling. All dogs were observed for several minutes after dosing to confirm that the dose was swallowed. Dogs were not fed within 12 hours prior to dosing and for at least 4 hours after treatment.\u003c/p\u003e\n\u003ch3\u003eTick infestation and assessment\u003c/h3\u003e\n\u003cp\u003eThe study used viable, unfed adult \u003cem\u003eH. longicornis\u003c/em\u003e ticks sourced from a laboratory colony first established at the laboratories of Shokukanken within one year of the study being conducted using wild, engorged ticks collected in the Gunma Prefecture, Japan. At each infestation, \u003cem\u003eH. longicornis\u003c/em\u003e adult ticks were placed on the auricle to mimic the natural preferred attachment sites. To facilitate infestation, dogs were sedated with xylazine hydrochloride (2% injectable formulation) and were also fitted with Elizabethan collars to reduce grooming for the duration of each infestation period. To ensure the dogs were acceptable hosts and for allocation purposes, dogs were first infested on Day \u0026minus;\u0026thinsp;7 and live, attached ticks were counted on Day \u0026minus;\u0026thinsp;5. To determine efficacy against an existing infestation, dogs were then infested on Day \u0026minus;\u0026thinsp;2 (2 days before treatment on Day 0) and tick counts were conducted on Day 2. Subsequent weekly infestations were conducted on Days 5, 12, 19, 26 and 33, and counts were performed 48\u0026thinsp;\u0026plusmn;\u0026thinsp;2 hours after each infestation (on Days 7, 14, 21, 28 and 35) to confirm the duration of preventative efficacy.\u003c/p\u003e\n\u003ch3\u003eTick counts\u003c/h3\u003e\n\u003cp\u003eDogs were sedated with xylazine hydrochloride (2% injectable formulation) at each tick count. Counts were conducted by trained personnel by systematically inspecting from head to tail while parting the hair by hand, with observed ticks being counted and removed. The dogs were then combed thoroughly with a fine-toothed comb. Each dog was examined for a minimum of 10 minutes, and if ticks were found in the final minute of counting, the examination was continued in 1-minute increments until no further ticks were found. All live and dead ticks were classified as either (1) unattached or free; (2) attached but un-engorged; or (3) attached and engorged. Personnel conducting counts were masked to treatment assignments and changed gloves between dogs.\u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eThe experimental unit was the individual dog. Tick counts were transformed using the log\u003csub\u003ee\u003c/sub\u003e(count\u0026thinsp;+\u0026thinsp;1) transformation prior to analysis to decrease variance and normalize the data. Transformed counts were analyzed according to a mixed linear model for repeated measures using the PROC MIXED procedure (SAS 9.4, Cary NC). The fixed effects of the model were treatment, time point and the interaction between treatment and time point. The random effects included block, animal, and error. Two-sided testing was performed at the significance level of α\u0026thinsp;=\u0026thinsp;0.05.\u003c/p\u003e \u003cp\u003eEfficacy was evaluated separately for (i) total live tick counts (free and attached ticks) and (ii) live, attached tick counts. Efficacy assessment was based on the percent reduction in the arithmetic and geometric mean live tick counts for Simparica Trio-treated dogs relative to those for placebo-treated dogs using Abbott\u0026rsquo;s formula:\u003c/p\u003e \u003cp\u003e% reduction\u0026thinsp;=\u0026thinsp;100 \u0026times; [mean count (placebo) \u0026ndash; mean count (treated)]/mean count (placebo).\u003c/p\u003e \u003c/div\u003e"},{"header":"Results and Discussion","content":"\u003cp\u003eThe sixteen dogs enrolled in the study proved to be acceptable hosts for \u003cem\u003eH. longicornis\u003c/em\u003e, demonstrating a geometric mean (arithmetic mean; range) adult live tick count of 17.1 (17.4; range 13 to 23) on Day \u0026minus;\u0026thinsp;5. All eight dogs treated with the placebo maintained robust adult tick infestations for the duration of the study. On Days 2, 7, 14, 21, 28, and 35 geometric mean live (free and attached) adult tick counts were 29.6, 26.8, 31.3, 27.9, 29.0, and 27.5, respectively, and arithmetic mean live (free and attached) tick counts were 29.8, 27.0, 31.5, 28.1, 29.0, and 27.5, respectively. More than 50% (range 51.1 to 63.3%) of the live adult ticks observed on placebo-treated dogs at each time point were attached. On Days 2, 7, 14, 21, 28, and 35 geometric mean live, attached adult tick counts were 16.6, 16.8, 15.7, 15.8, 17.4, and 16.6, respectively, and arithmetic mean live, attached tick counts were 16.9, 17.1, 16.1, 16.0, 17.6, and 16.8, respectively. A few dead ticks (0 to 2) were found on placebo-treated dogs at each time point.\u003c/p\u003e \u003cp\u003eActual administered doses of Simparica Trio ranged from 1.23 to 1.58 mg/kg sarolaner, 25 to 32 \u0026micro;g/kg moxidectin, and 5.14 to 6.58 mg/kg pyrantel (as pamoate salt). The single oral dose of Simparica Trio was well tolerated by all eight treated dogs, with no adverse reactions to treatment recorded during daily general health observations. On Days 2, 7, 14, 21, 28, and 35 post-treatment, arithmetic mean live (free and attached) tick counts in Simparica Trio-treated dogs were 8.8, 11.9, 9.1, 7.9, 10.9, and 7.1, respectively, and geometric mean live (free and attached) tick counts were 8.0, 11.6, 8.8, 7.3, 10.5, and 6.3, respectively. The eight dogs treated with Simparica Trio showed significantly reduced (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) geometric mean live (free and attached) adult tick counts compared to placebo-treated dogs at each time point examined (5.64\u0026thinsp;\u0026le;\u0026thinsp;t(50.8)\u0026thinsp;\u0026le;\u0026thinsp;9.62, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.0001). In these same Simparica Trio-treated dogs, the number of killed adult ticks collected from each dog at each timepoint ranged from 14 to 27 (28 to 54% of the infestation). Almost all (98%) live adult ticks recovered from Simparica Trio-treated dogs were unattached, with geometric (arithmetic) mean live, attached tick counts for Simparica Trio-treated dogs being \u0026le;\u0026thinsp;0.3 (\u0026le;\u0026thinsp;0.4) at each time point (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). A total of only nine live, attached adult ticks were recovered from the Simparica Trio-treated dogs over the 35 days: two dogs never had live, attached ticks; three dogs were observed to have a single live, attached tick at one time point; and three dogs were observed to have a single live, attached tick at two different time points.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eGeometric (arithmetic) mean live, attached \u003cem\u003eHaemaphysalis longicornis\u003c/em\u003e counts and ranges for dogs treated orally (Day 0) with placebo or Simparica Trio chewable tablets and percent efficacy relative to placebo at 48 hours after treatment and subsequent weekly re-infestations\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"8\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c4\" namest=\"c1\"\u003e \u003cp\u003ePlacebo\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"3\" nameend=\"c7\" namest=\"c5\"\u003e \u003cp\u003eSimparica Trio\u003csup\u003e1\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003ePercent Efficacy\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eDay\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGeometric Mean\u003c/p\u003e \u003cp\u003e(Arithmetic mean)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eRange\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNo. of Dogs with ticks\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eGeometric Mean\u003c/p\u003e \u003cp\u003e(Arithmetic mean)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eRange\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eNo of Dogs with ticks\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e16.6\u003c/p\u003e \u003cp\u003e(16.9)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e12\u0026ndash;22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.2\u003c/p\u003e \u003cp\u003e(0.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u0026ndash;1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e2/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e98.9\u003c/p\u003e \u003cp\u003e(98.5)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e16.8\u003c/p\u003e \u003cp\u003e(17.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e12\u0026ndash;22\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.1\u003c/p\u003e \u003cp\u003e(0.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u0026ndash;1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e1/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e99.5\u003c/p\u003e \u003cp\u003e(99.3)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e15.7\u003c/p\u003e \u003cp\u003e(16.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11\u0026ndash;21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.2\u003c/p\u003e \u003cp\u003e(0.3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u0026ndash;1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e2/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e98.8\u003c/p\u003e \u003cp\u003e(98.5)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e15.8\u003c/p\u003e \u003cp\u003e(16.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e13\u0026ndash;19\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.0\u003c/p\u003e \u003cp\u003e(0.0)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u0026ndash;0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e100\u003c/p\u003e \u003cp\u003e(100)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e17.4\u003c/p\u003e \u003cp\u003e(17.6)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e13\u0026ndash;21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.3\u003c/p\u003e \u003cp\u003e(0.4)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u0026ndash;1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e3/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e98.3\u003c/p\u003e \u003cp\u003e(97.9)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e16.6\u003c/p\u003e \u003cp\u003e(16.8)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e13\u0026ndash;20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e8/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.1\u003c/p\u003e \u003cp\u003e(0.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0\u0026ndash;1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e1/8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e99.5\u003c/p\u003e \u003cp\u003e(99.3)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"8\"\u003e\u003csup\u003e1\u003c/sup\u003eTreatment with placebo or Simparica Trio (minimum dose 1.2 mg/kg sarolaner, 24 \u0026micro;g/kg moxidectin, 5 mg/kg pyrantel (as pamoate salt)) occurred on Day 0. \u003csup\u003e2\u003c/sup\u003eGeometric (arithmetic) efficacy of Simparica Trio versus Placebo. Geometric mean live, attached tick counts significantly lower compared to placebo at each timepoint (22.66\u0026thinsp;\u0026le;\u0026thinsp;t(71.6)\u0026thinsp;\u0026le;\u0026thinsp;24.18, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.0001)\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eIt is logical to assume that live, unattached adult ticks recovered from dogs during this study were not feeding and thus had not been exposed to Simparica Trio. Therefore, efficacy of Simparica Trio against \u003cem\u003eH. longicornis\u003c/em\u003e was calculated using the live, attached adult tick counts recorded for dogs in each treatment group (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). In this study, geometric mean counts of live, attached adult ticks for Simparica Trio-treated dogs were significantly lower at each time point compared with placebo-treated dogs (22.66\u0026thinsp;\u0026le;\u0026thinsp;t(71.6)\u0026thinsp;\u0026le;\u0026thinsp;24.18, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.0001, Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). One treatment with Simparica Trio provided dogs with 98.9% efficacy against existing adult \u003cem\u003eH. longicornis\u003c/em\u003e infestations and was \u0026ge;\u0026thinsp;98.3% effective in protecting dogs against re-infestation with adult \u003cem\u003eH. longicornis\u003c/em\u003e for at least 35 days after treatment.\u003c/p\u003e \u003cp\u003eThe impacts of tick infestations on dogs are well known [\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e, \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e], and the negative consequences of \u003cem\u003eH. longicornis\u003c/em\u003e infestation have been documented across multiple host species, ranging from irritation and anemia to diarrhea, production losses, and even death [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Laboratory studies on \u003cem\u003eH. longicornis\u003c/em\u003e have demonstrated that all life stages are able to feed on dogs, with mean larval, nymphal, and adult feeding periods similar to those recorded when feeding on cattle [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e, \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e]. Additionally, environmental sampling has recovered host-seeking, engorged \u003cem\u003eH. longicornis\u003c/em\u003e, providing evidence for repeated successful questing events and highlighting the role this tick species may play in the spread of vector-borne agents [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e]. The need for an effective means of treating and controlling \u003cem\u003eH. longicornis\u003c/em\u003e is apparent considering its importance as a vector of human and animal disease agents, the negative impacts of infestation, the species\u0026rsquo; wide host range, and its invasive ability. Previous studies show that a single oral dose of sarolaner can rapidly and effectively control multiple canine tick species [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e, \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e, \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e, \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e, \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e, \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e, \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e], including \u003cem\u003eH. longicornis\u003c/em\u003e [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e]. Here, we show sarolaner administered in a combination product, as Simparica Trio, remained highly effective in rapidly treating existing adult attached, live \u003cem\u003eH. longicornis\u003c/em\u003e infestations in dogs and in preventing re-infestation for at least 35 days. This level of protection provides interruption of the \u003cem\u003eH. longicornis\u003c/em\u003e life cycle and a reduction in the likelihood of pathogen transmission between hosts.\u003c/p\u003e \u003cp\u003eWhen administering traditional pill or tablet medications, many dog owners worry about inaccurate dosing and damaging the animal-human bond during unsuccessful or stressful pilling events [\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e]. In contrast, dog owners generally view chewable medications positively due to their increased ease of administration and formulation palatability. The Simparica Trio chewable tablet has been shown to be well-accepted by dogs, with one study reporting the majority (91.9%) of 517 doses offered to dogs by owners in their own homes taken by free choice without food or in food (and only 8.1% of tablets requiring pilling) [\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e]. Moreover, as a combination product containing both moxidectin and pyrantel in addition to sarolaner, Simparica Trio provides dog owners seeking month-long efficacy against ticks and fleas the additional benefits of effective treatment for roundworm and hookworm infections and protection against lungworm and heartworm disease [\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e, \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e, \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e, \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e, \u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e]. When one considers the important roles of palatability, ease of administration, and broad-spectrum protection in owner compliance [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e, \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e, \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e], it is clear that Simparica Trio makes an important contribution in the field of canine health by providing comprehensive protection in a single convenient product.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eA single oral dose of the combination antiparasiticide Simparica Trio administered at the minimum label dose of 1.2 mg/kg sarolaner, 24 \u0026micro;g/kg moxidectin, and 5 mg/kg pyrantel (as pamoate salt) effectively treated an existing infestation of \u003cem\u003eH. longicornis\u003c/em\u003e adults and prevented re-infestation for up to 35 days. Simparica Trio is an easy and convenient means of providing dogs with rapid protection against many important tick species.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003e\u003cstrong\u003eUS:\u003c/strong\u003e United States of America\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics Approval and Consent to Participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe protocol was reviewed and approved by the Institutional Animal Care and Use Committee at Shokukanken, Inc. prior to implementation.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for Publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of Data and Materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe dataset supporting the conclusions of this article are included within the article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting Interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was funded by Zoetis Japan. KK, TF, AH, and SM were employees of Zoetis at the time of the study. MN was the contracted study investigator.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was funded by Zoetis Japan.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors’ Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors were involved in protocol development, data interpretation and preparing the manuscript. AH was the biometrician responsible for the study design and statistical analysis. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors would like to thank the support staff at Shokukanken Inc. for their assistance with the conduct of the study and Sharon Chirgwin DeRosa for assistance with manuscript preparation.\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eChomel B. Tick-borne infections in dogs-an emerging infectious threat. Vet Parasitol.\u003cem\u003e \u003c/em\u003e2011;179:294-301; doi: 10.1016/j.vetpar.2011.03.040.\u003c/li\u003e\n\u003cli\u003eHoogstraal H, Roberts FH, Kohls GM, Tipton VJ. Review of \u003cem\u003eHaemaphysalis\u003c/em\u003e (kaiseriana)\u003cem\u003e longicornis\u003c/em\u003e Neumann (resurrected) of Australia, New Zealand, New Caledonia, Fiji, Japan, Korea, and Northeastern China and USSR, and its parthenogenetic and bisexual populations (Ixodoidea, Ixodidae). 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Recovery of Partially Engorged \u003cem\u003eHaemaphysalis longicornis\u003c/em\u003e (Acari: Ixodidae) Ticks from Active Surveillance. J Med Entomol.\u003cem\u003e \u003c/em\u003e2022;59 5:1842-6; doi: 10.1093/jme/tjac099.\u003c/li\u003e\n\u003cli\u003ePackianathan R, Hodge A, Bruellke N, Jackson C, Maeder S. Efficacy of combination products containing sarolaner, moxidectin and pyrantel (Simparica Trio\u0026trade;) or afoxolaner and milbemycin (NexGard Spectra\u0026reg;) against induced infestations of \u003cem\u003eIxodes holocyclus\u003c/em\u003e in dogs. Parasit Vectors.\u003cem\u003e \u003c/em\u003e2020;13 1:448; doi: 10.1186/s13071-020-04323-8. https://doi.org/10.1186/s13071-020-04323-8.\u003c/li\u003e\n\u003cli\u003eBecskei C, Liebenberg J, Thys M, Mahabir SP. Efficacy of a novel chewable tablet containing sarolaner, moxidectin and pyrantel (Simparica Trio\u0026trade;) against four common tick species infesting dogs in Europe. 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Adv Drug Deliv Rev.\u003cem\u003e \u003c/em\u003e2004;56 10:1399-413; doi: https://doi.org/10.1016/j.addr.2004.02.012. https://www.sciencedirect.com/science/article/pii/S0169409X04000729.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"parasites-and-vectors","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"parv","sideBox":"Learn more about [Parasites \u0026 Vectors](http://parasitesandvectors.biomedcentral.com/)","snPcode":"13071","submissionUrl":"https://submission.nature.com/new-submission/13071/3","title":"Parasites \u0026 Vectors","twitterHandle":"@bugbittentweets","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Asian longhorned tick, Dog, Efficacy, Haemaphysalis longicornis, Moxidectin, Oral Combination, Pyrantel pamoate, Sarolaner, Simparica Trio®","lastPublishedDoi":"10.21203/rs.3.rs-5455294/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5455294/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eThe efficacy of Simparica Trio\u003cb\u003e\u0026reg;\u003c/b\u003e (Zoetis), an oral combination product for dogs containing sarolaner, moxidectin, and pyrantel pamoate, was evaluated against \u003cem\u003eHaemaphysalis longicornis\u003c/em\u003e, the most common tick species affecting dogs in the East Asia / Pacific region and an exotic species now reported from multiple states within the USA.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eTwo groups of 8 dogs each were ranked based on pretreatment tick counts and randomly allocated to treatment on Day 0 with either placebo or Simparica Trio at the minimum label dose of 1.2 mg/kg sarolaner, 24 \u0026micro;g/kg moxidectin, and 5 mg/kg pyrantel (as pamoate salt). Dogs were infested with 50 viable adult \u003cem\u003eH. longicornis\u003c/em\u003e on Days \u0026minus;\u0026thinsp;2, 5, 12, 19, 26, and 33. Tick counts were conducted for all dogs 48 hours after treatment and subsequent re-infestations.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eA single oral administration of Simparica Trio was 98.9% effective in treating existing \u003cem\u003eH. longicornis\u003c/em\u003e infestation when considering live, attached (feeding) ticks. Efficacy remained\u0026thinsp;\u0026gt;\u0026thinsp;98% in preventing re-infestation for at least 35 days, and the geometric mean live, attached tick counts for Simparica Trio-treated dogs were significantly lower than for placebo-treated dogs (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) at all time points assessed in the study. Geometric mean total live (attached and free) tick counts for dogs treated with Simparica Trio were also significantly lower (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) than those for placebo-treated dogs at each time point. No adverse events were reported for any dogs throughout the duration of the study.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e \u003cp\u003eOne dose of Simparica Trio at minimum label dose was highly efficacious in treating and controlling adult \u003cem\u003eH. longicornis\u003c/em\u003e in dogs for more than one month.\u003c/p\u003e","manuscriptTitle":"Efficacy of a Combination Product containing Sarolaner, Moxidectin and Pyrantel (Simparica Trio ®) against induced infestations of Haemaphysalis longicornis on dogs","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-12-17 14:22:25","doi":"10.21203/rs.3.rs-5455294/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-12-13T19:08:25+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-12-11T18:48:47+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-12-09T18:36:04+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"48325178896144192767560056510749941639","date":"2024-11-19T13:48:12+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"20148705881390636280031152303380424576","date":"2024-11-18T19:30:36+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-11-18T19:07:17+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-11-16T11:33:50+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-11-16T11:11:00+00:00","index":"","fulltext":""},{"type":"submitted","content":"Parasites \u0026 Vectors","date":"2024-11-14T15:59:39+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"parasites-and-vectors","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"parv","sideBox":"Learn more about [Parasites \u0026 Vectors](http://parasitesandvectors.biomedcentral.com/)","snPcode":"13071","submissionUrl":"https://submission.nature.com/new-submission/13071/3","title":"Parasites \u0026 Vectors","twitterHandle":"@bugbittentweets","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"664bf5cb-c446-4210-9c40-bd4ad61e78ca","owner":[],"postedDate":"December 17th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-03-31T16:09:00+00:00","versionOfRecord":{"articleIdentity":"rs-5455294","link":"https://doi.org/10.1186/s13071-025-06747-6","journal":{"identity":"parasites-and-vectors","isVorOnly":false,"title":"Parasites \u0026 Vectors"},"publishedOn":"2025-03-26 15:57:07","publishedOnDateReadable":"March 26th, 2025"},"versionCreatedAt":"2024-12-17 14:22:25","video":"","vorDoi":"10.1186/s13071-025-06747-6","vorDoiUrl":"https://doi.org/10.1186/s13071-025-06747-6","workflowStages":[]},"version":"v1","identity":"rs-5455294","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5455294","identity":"rs-5455294","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}