Safety evaluation of a novel topical combination of esafoxolaner, eprinomectin and praziquantel, in reproducing female cats.

NexGard® Combo, a novel topical endectoparasiticide product for cats, is a combination of esafoxolaner, eprinomectin and praziquantel. The safety of this novel combination administered to females during reproduction and lactation was evaluated per analysis of breeding parameters and adverse reactions observed on females and offspring. Females with successful breeding history were randomized to three groups, a placebo group and groups treated with the novel formulation at 1× or 3× multiples of the maximum exposure dose. Females were dosed at 28-day intervals, at least twice before mating, then during a period including mating, pregnancy, whelping and 56 days of lactation. In the placebo, 1× and 3× groups, 10, 9 and 10 females, respectively completed the study (nine, seven and nine females achieved pregnancy), and were dosed 7.1 times on average. Breeding parameters included success of mating, success of gestation, length of gestation, abortion rate, number of live, dead and stillborn kittens at birth, number of kittens with abnormalities, weight of kittens after birth and at weaning, growth of kittens, proportion of male and female kittens, and proportion of kittens born alive and weaned. No significant adverse reactions related to the novel combination were observed on females and on kittens; no significant and adverse effects on breeding parameters were observed.

TITLE: Évaluation de l'innocuité d'une nouvelle combinaison topique d'esafoxolaner, d'éprinomectine et de praziquantel chez les chattes reproductrices. ABSTRACT: NexGard® Combo, un nouvel endectoparasiticide topique pour chats, est une combinaison d'esafoxolaner, d'éprinomectine et de praziquantel. La sécurité de cette nouvelle association administrée aux chattes pendant la reproduction et la lactation a été évaluée par analyse des paramètres d'élevage et des effets indésirables observés sur les femelles et les descendants. Les chattes ayant des antécédents de reproduction réussie ont été randomisées en trois groupes, un groupe placebo et des groupes traités avec la nouvelle formulation à des multiples de 1× ou 3× la dose d'exposition maximale. Les femelles ont reçu des doses à 28 jours d'intervalle, au moins deux fois avant l'accouplement, puis pendant une période comprenant l'accouplement, la gestation, la mise bas et 56 jours de lactation. Dans les groupes placebo, 1× et 3×, repectivement dix, neuf et dix chattes ont terminé l'étude (neuf, sept et neuf chattes ont été gestantes) et ont été traitées 7,1 fois en moyenne. Les paramètres d'élevage comprenaient le succès de l'accouplement, le succès de la gestation, la durée de la gestation, le taux d'avortement, le nombre de chatons vivants, morts et mort-nés à la naissance, le nombre de chatons présentant des anomalies, le poids des chatons après la naissance et au sevrage, la croissance des chatons, la proportion de chatons mâles et femelles et la proportion de chatons nés vivants et sevrés. Aucun effet indésirable significatif lié à la nouvelle association n'a été observé chez les femelles et les chatons et aucun effet indésirable significatif sur les paramètres d'élevage n'a été observé.

Parasite specific 7SL-derived small RNA is an effective target for diagnosis of active trypanosomiasis infection.

Human and animal African trypanosomiasis (HAT & AAT, respectively) remain a significant health and economic issue across much of sub-Saharan Africa. Effective control of AAT and potential eradication of HAT requires affordable, sensitive and specific diagnostic tests that can be used in the field. Small RNAs in the blood or serum are attractive disease biomarkers due to their stability, accessibility and available technologies for detection. Using RNAseq, we have identified a trypanosome specific small RNA to be present at high levels in the serum of infected cattle. The small RNA is derived from the non-coding 7SL RNA of the peptide signal recognition particle and is detected in the serum of infected cattle at significantly higher levels than in the parasite, suggesting active processing and secretion. We show effective detection of the small RNA in the serum of infected cattle using a custom RT-qPCR assay. Strikingly, the RNA can be detected before microscopy detection of parasitaemia in the blood, and it can also be detected during remission periods of infection when no parasitaemia is detectable by microscopy. However, RNA levels drop following treatment with trypanocides, demonstrating accurate prediction of active infection. While the small RNA sequence is conserved between different species of trypanosome, nucleotide differences within the sequence allow generation of highly specific assays that can distinguish between infections with Trypanosoma brucei, Trypanosoma congolense and Trypanosoma vivax. Finally, we demonstrate effective detection of the small RNA directly from serum, without the need for pre-processing, with a single step RT-qPCR assay. Our findings identify a species-specific trypanosome small RNA that can be detected at high levels in the serum of cattle with active parasite infections. This provides the basis for the development of a cheap, non-invasive and highly effective diagnostic test for trypanosomiasis.

Comparative efficacy of oral administrated afoxolaner (NexGard™) and fluralaner (Bravecto™) with topically applied permethrin/imidacloprid (Advantix(®)) against transmission of Ehrlichia canis by infected Rhipicephalus sanguineus ticks to dogs.

BACKGROUND: The ability of the topical spot-on Advantix(®) (50 % permethrin/10 % imidacloprid) to prevent transmission of Ehrlichia canis by infected Rhipicephalus sanguineus ticks to dogs has previously been reported. The recent market introduction of chewable tablets containing the novel compounds, afoxolaner (NexGard™) and fluralaner (Bravecto™) enabled us to conduct a comparative efficacy study with respect to the ability of these three products to block transmission of E. canis by ticks to dogs. The speed of kill, immediate drop-off rate and anti-attachment efficacy of the respective products were also studied. METHODS: The study was a blinded parallel group design, wherein 32 dogs were randomised into four different groups of eight dogs. Group 1 served as negative placebo control, group 2 and 3 were treated on Days 0, 28 and 56 with NexGard™ and Advantix(®), respectively. Group 4 was dosed once on Day 0 with Bravecto™. For tick efficacy assessments 50 non-infected ticks were placed onto the dogs on Days 30, 35, 42, 49, 56, 63, 70, 77 and 84 and on animal tick counts were performed at 3 h, 6 h and 12 h after infestation. To evaluate the ability to block transmission of E. canis, each dog was challenged by releasing 80 adult E. canis-infected R. sanguineus ticks into their sleeping kennels on Days 31, 38, 45 and 52. The animals were monitored for clinical signs of monocytic ehrlichiosis (pyrexia and thrombocytopenia) and were tested for E. canis DNA by PCR and for specific antibodies using IFA. A dog was considered infected with E. canis if both PCR and IFA yielded positive test results up to Day 84. RESULTS: Mean arithmetic tick counts on dogs treated with the Advantix(®) spot-on were significantly (P < 0.0005) lower throughout the study as compared with the negative controls and was, with respect to the speed of kill and resulting onset of acaricidal efficacy, superior over NexGard™ and Bravecto™ at all time points in the 12 h period observed (3 h, 6 h and 12 h). None of the dogs treated with the Advantix(®) spot-on became infected with E. canis, whereas six out of eight untreated control dogs acquired the infection. Furthermore, E. canis infection was diagnosed in four out of eight dogs treated with NexGard™ and in two out of eight dogs treated with Bravecto™. CONCLUSIONS: The speed of kill of the two recently registered systemic compounds against R. sanguineus was not sufficiently fast to prevent transmission of E. canis and resulted in only low partial blocking and protection capacity while Advantix(®) effectively blocked transmission of E. canis to dogs in the challenge period and thus provided adequate protection for dogs against monocytic ehrlichiosis.

Comparative speed of kill after treatment with Simparica™ (sarolaner) and Advantix®(imidacloprid + permethrin) against induced infestations of Dermacentor reticulatus on dogs.

BACKGROUND: Ticks are common ectoparasites that infest dogs globally. Acaricides with rapid and sustained speed of kill are critical to control infestations and to reduce the risk of disease transmission. This study evaluated the speed of kill for 5 weeks after a single dose of orally administered Simparica™ (sarolaner) against induced infestations with Dermacentor reticulatus on dogs, compared to Advantix® Spot-on solution for dogs (imidacloprid + permethrin). METHODS: Twenty four dogs were randomly allocated to treatment with either a placebo tablet, a sarolaner tablet (at 2 to 4 mg/kg) or with Advantix® as per label instructions. Dogs were treated on Day 0 and tick counts were performed in situ at 8 and 12 hours and with removal of the ticks at 24 hours after treatment and subsequent re-infestations on Days 7, 14, 21, 28 and 35. Acaricidal efficacy was determined at each time point relative to live tick counts from the placebo-treated dogs. RESULTS: Based on arithmetic (geometric) mean tick counts, the efficacy of sarolaner was ≥75.6 % (89.6 %) within 8 hours of treatment and tick counts were significantly lower than placebo and imidacloprid + permethrin-treated dogs (P < 0.0001), while imidacloprid + permethrin had no significant reduction (P ≥ 0.3990) at 8 or 12 hours after treatment. Sarolaner killed all ticks on the dogs within 24 hours after treatment, while imidacloprid + permethrin efficacy was only 48.1 %. After weekly re-infestations sarolaner significantly reduced the tick counts versus placebo within 8 hours on Days 7, 14 and 35 (P ≤ 0.0239), and at 12 hours and 24 hours (P ≤ 0.0079) until Day 35.Sarolaner efficacy was ≥95.8 % within 24 hours for 35 days. Significantly more live ticks (P ≤ 0.0451) were recovered from imidacloprid + permethrin-treated dogs than from sarolaner-treated dogs at 24 hours after infestation on all days. There were no sarolaner-related adverse reactions during the study. CONCLUSIONS: This study demonstrated that Simparica™ had a faster and more consistent speed of kill against D. reticulatus compared to Advantix®. The rapid and consistent efficacy within 24 hours for 5 weeks after a single oral dose of Simparica™ provides effective and reliable control of D. reticulatus and reduces the risk of transmission of tick-borne diseases.

The efficacy of a generic doxycycline tablet in the treatment of canine monocytic ehrlichiosis.

The objective of the present study was to evaluate the therapeutic efficacy of a generic doxycycline tablet (DoxyVet) against Ehrlichia canis infection in dogs. Canine monocytic ehrlichiosis is caused by the bacterium E. canis and transmitted by the brown kennel tick (Rhipicephalus sanguineus). Six disease-free and tick-free dogs were infested with E. canis infected ticks. Once diagnosed (with polymerase chain reaction [PCR] analysis and platelet counts) as positive for infection, doxycycline tablets were administered orally once a day for 20 consecutive days, at a target dose level of 10 mg/kg. The actual dose administered was calculated as ranging between 10 mg/kg and 11.7 mg/kg. The PCR analysis, 28 days after the first administration of the tablets, failed to detect E. canis in any of the dogs. On Day 56 of the study, four of the dogs were diagnosed with E. canis for the second time and a fifth dog was diagnosed on Day 70. The platelet counts of the sixth dog remained within normal levels and it was discharged from the study on Day 84. Doxycycline tablets were then administered to the remaining five infected dogs for 28 consecutive days. Four of these dogs had no positive PCR results during the following 3 months. The fifth dog was diagnosed with E. canis for the third time 58 days after the last tablets of the second treatment had been administered, after which it was rescue treated (doxycycline for a further 28 days). The results indicate that doxycycline administered in tablet form (DoxyVet) at 10 mg/kg - 11.7 mg/kg body mass once daily for 28 consecutive days clears most dogs of infection. The importance of a concomitant tick-control programme is therefore stressed.

Efficacy, acceptability and tolerability of the new oral phosphate binder Lenziaren® in healthy cats fed a standard diet.

BACKGROUND: The efficacy, acceptability and tolerability of the new oral phosphate binder Lenziaren® (SBR759) were evaluated in a randomized parallel-group design study in 36 healthy cats (n = 6 per group). Five groups were fed once daily with a commercial diet containing 0.2% phosphorus ("standard diet") into which was mixed Lenziaren® at 0.25, 0.5, 1.0 or 2.0 g/day or no treatment (control group) daily for 30 days. A sixth group was fed a commercial diet containing lower amounts (0.12%) of phosphorus ("renal diet") and no treatment. RESULTS: When compared to the control group, Lenziaren® produced significant dose-related reductions in urine phosphate concentrations, urine phosphate excretion and fractional urinary phosphate excretion. Significant effects versus the control group were observed at the 0.5, 1.0 and 2.0 g/day dosages. Lenziaren® was well tolerated and was associated with higher food consumption and serum iron concentrations versus the control. When compared to the control group, the renal diet was associated with significantly lower urine phosphate concentrations and loss of body weight. Lenziaren® had similar effects on urine phosphate concentrations compared to the renal diet, but was not associated with loss of body weight. CONCLUSIONS: Lenziaren® was effective as an oral phosphate binder in cats fed with a standard diet containing 0.2% phosphorus. The acceptability and tolerability were good. Dosages of 0.5-1.0 g/cat per day are recommended for clinical testing in cats fed with a standard diet.

The ability of an oral formulation of afoxolaner to block the transmission of Babesia canis by Dermacentor reticulatus ticks to dogs.

BACKGROUND: Canine babesiosis due to Babesia canis is an endemic disease in many European countries. A vaccine is available in some countries, but it does not prevent the infection and just helps in reducing the gravity of clinical signs. Therefore, the major way to help preventing the disease is by controlling tick infestations on dogs.To assess the preventive efficacy of afoxolaner (NexGard®), a new oral anti- flea and tick product, against Babesia canis infected adult Dermacentor reticulatus in an experimentally controlled study. METHODS: Sixteen healthy mixed breed adult dogs, negative for Babesia canis antibodies were included in a single centre, randomized, blinded and controlled study to evaluate the impact of treatment with afoxolaner on the transmission of Babesia canis to dogs exposed to Dermacentor reticulatus. The dogs were randomly allocated into two groups of 8 dogs each. One group remained untreated. In the other group, dogs were treated orally with a novel formulation of afoxolaner (NexGard®) on day 0. All dogs were infested each by 50 adult Dermacentor reticulatus ticks (equal sex ratio) at days 7, 14, 21 and 28. The Dermacentor reticulatus ticks were confirmed to harbour Babesia canis by Polymerase Chain Reaction (PCR). RESULTS: The treatment was well tolerated by all dogs without any adverse effects. Babesia canis was transmitted by D. reticulatus to all untreated control dogs, confirmed following demonstration of hyperthermia, detection of B. canis parasites in blood smears and PCR assay from blood and serology. These confirmed infected dogs were subsequently treated with imidocarb and diminazene. The treated dogs remained negative based on all criteria until the last study, Day 56, confirming that the oral treatment of dogs with NexGard® prevented transmission of Babesia canis and development of clinical babesiosis for up to 28 days. CONCLUSION: This is the first demonstration that an oral acaricidal treatment may prevent the transmission of a pathogen despite the need for the tick to attach and start feeding before being killed by the acaricide.

Development and evaluation of an ITS1 "Touchdown" PCR for assessment of drug efficacy against animal African trypanosomosis.

Animal African trypanosomoses (AAT) are caused by flagellated protozoa of the Trypanosoma genus and contribute to considerable losses in animal production in Africa, Latin America and South East Asia. Trypanosoma congolense is considered the economically most important species. Drug resistant T. congolense strains present a threat to the control of AAT and have triggered research into discovery of novel trypanocides. In vivo assessment of trypanocidal efficacy relies on monitoring of treated animals with microscopic parasite detection methods. Since these methods have poor sensitivity, follow-up for up to 100 days after treatment is recommended to increase the chance of detecting recurrent parasitaemia waves. Molecular techniques are more amendable to high throughput processing and are generally more sensitive than microscopic detection, thus bearing the potential of shortening the 100-day follow up period. The study presents a "Touchdown" PCR targeting the internal transcribed spacer 1 of the ribosomal DNA (ITS1 TD PCR) that enables detection and discrimination of different Trypanosoma taxa in a single run due to variations in PCR product sizes. The assay achieves analytical sensitivity of 10 parasites per ml of blood for detection of T. congolense savannah type and T. brucei, and 100 parasites per ml of blood for detection of T. vivax in infected mouse blood. The ITS1 TD PCR was evaluated on cattle experimentally infected with T. congolense during an investigational new veterinary trypanocide drug efficacy study. ITS1 TD PCR demonstrated comparable performance to microscopy in verifying trypanocide treatment success, in which parasite DNA became undetectable in cured animals within two days post-treatment. ITS1 TD PCR detected parasite recrudescence three days earlier than microscopy and had a higher positivity rate than microscopy (84.85% versus 57.58%) in 66 specimens of relapsing animals collected after treatments. Therefore, ITS1 TD PCR provides a useful tool in assessment of drug efficacy against T. congolense infection in cattle. As the assay bears the potential for detection of mixed infections, it may be applicable for drug efficacy studies and diagnostic discrimination of T. vivax and T. congolense against other pathogenic trypanosomes, including T. brucei, T. evansi and T. equiperdum.