Asymptomatic Anticoagulant Rodenticide Exposure in Dogs and Cats-A French and Belgian Rural and Urban Areas Study.

Anticoagulant rodenticides (ARs) are important tools for controlling rodent pests, but they also pose a health threat to non-target species. ARs are one of the most common causes of pet poisoning. However, exposure of domestic animals to subclinical doses of ARs is poorly documented. To study the random exposure of dogs and cats to ARs, feces from animals showing no clinical signs of rodenticide poisoning were collected from a network of French and Belgian veterinarians. We analyzed fresh feces from 304 dogs and 289 cats by liquid chromatography-tandem mass spectrometry. This study showed a limited prevalence of AR exposure in dogs and cats of 2.6 and 4.5% respectively. In both species, access to the outdoors is a risk factor for ARs exposure. In contrast, the sex of the animals did not affect the ARs exposure status. The observation of the ratio of cis and trans isomers suggested primary exposure in dogs, but also in some cats. While primary exposure in dogs appears to be related to the use of ARs as plant protection products, primary exposure in cats may be malicious, as warfarin, an anticoagulant formerly used as a rodenticide and now used only in humans, was found in 4 of 13 exposed cats. Secondary exposure may also occur in cats.Our study showed reduced exposure in dogs and cats, compared to wildlife, which often has high exposure, especially in areas where rodent control is important.

A large-scale field randomized trial demonstrates safety and efficacy of the vaccine LetiFend® against canine leishmaniosis.

Canine leishmaniosis is a zoonotic disease caused by Leishmania infantum. Extensive research is currently ongoing to develop safe and effective vaccines to protect from disease development. The European Commission has granted a marketing authorization for LetiFend®, a new vaccine containing recombinant Protein Q. The efficacy of LetiFend® vaccination in a large-scale dog population of both sexes, different breeds and ages in endemic areas is reported in this multicenter, randomized, double-blind, placebo-controlled field trial. Dogs (n = 549) living in France and Spain were randomly selected to receive a single subcutaneous dose of LetiFend® or placebo per year, and were naturally exposed to two L. infantum transmission seasons. Clinical examinations, blood and lymphoid organ sampling to evaluate serological, parasitological and disease status of the dogs were performed at different time points during the study. LetiFend® was very well tolerated and clearly reduced the incidence of clinical signs related to leishmaniosis. The number of confirmed cases of leishmaniosis was statistically significantly lower in the vaccine group. The number of dogs with parasites was close to be significantly reduced in the vaccine group (p = 0.0564). Re-vaccination of seropositive dogs demonstrated to be safe and not to worsen the course of the disease. The likelihood that a dog vaccinated with LetiFend® develops a confirmed case or clinical signs of leishmaniosis in areas with high pressure is, respectively, 5 and 9.8 time less than that for an unvaccinated dog. Thus, the overall efficacy of the LetiFend® vaccine in the prevention of confirmed cases of leishmaniosis in endemic areas with high disease pressure was shown to be 72%. In conclusion, this field trial demonstrates that LetiFend® is a novel, safe and effective vaccine for the active immunization of non-infected dogs from 6 months of age in reducing the risk of developing clinical leishmaniosis after natural infection with Leishmania infantum.

Validation of a new liquid chromatography- tandem mass spectrometry ion-trap technique for the simultaneous determination of thirteen anticoagulant rodenticides, drugs, or natural products.

The purpose of this study was to develop and validate a liquid chromatography-tandem mass spectrometry method for the identification and quantification of anticoagulant (anti-vitamin K or AVK) compounds, including rodenticides, drugs, and natural products because no published method could be found. The proposed method is based on ion-trap technology with electrospray ionization (ESI) and multiple reaction monitoring (MRM) technique. Each AVK is identified by means of its retention time, precursor ion, and two product ions. Plasma samples are extracted by liquid-liquid partition on Toxi-tube B((R)). The method was validated on dog plasma and gave good results in terms of specificity, linearity, and percent recovery for the 14 AVK tested (warfarin, acenocoumarol, bromadiolone, brodifacoum, chlorophacinone, coumatetralyl, dicoumarol, difenacoum, difethialone, flocoumafen, fluindione, phenindione, and tioclomarol). The limits of detection ranged from 5 to 25 ng/mL. Intraday repeatability was good, but interday repeatability was more variable though still sufficient for our diagnostic purposes. The technique was successfully applied in a series of clinical investigations to demonstrate its applicability in various animal species and gave very high sensitivity and specificity results.

Osteomyelitis and discospondylitis due to Scedosporium apiospermum in a dog.

A 6-year-old, 30-kg, female German Shepherd Dog, living in a leishmaniasis enzootic area, was presented with a severe rear limb motor disorder and a medical history of acute onset of fever. Routine hematology indicated neutrophilia. Spinal survey radiographs were consistent with osteomyelitis and discospondylitis. Because of the poor clinical prognosis and the painful nature of the lesions, the dog was euthanized at the owners' request. At necropsy, T13-L1 vertebrae had large areas of necrosis within the vertebral bodies. Histopathological findings were consistent with chronic, severe, fungal osteomyelitis and discospondylitis. Polymerase chain reaction identified Scedosporium apiospermum, a eutrophic filamentous fungus now recognized as an emerging agent of severe infections in immunosuppressed human patients.

Multiple oral dosing of ketoconazole increases dog exposure to ivermectin.

PURPOSE: The parasiticide ivermectin and the antimicrobial drug ketoconazole are macrolides that interact with P-glycoprotein. We investigated the effects of ketoconazole at a clinical dose on the pharmacokinetics of ivermectin, a CYP3A substrate with low hepatic clearance. METHODS: Beagle dogs received a single subcutaneous injection of ivermectin at 0.05 mg/kg alone (n=6) or in combination with a daily oral dose of ketoconazole 10 mg/kg over 5 days before and after ivermectin administration (n=6). The plasma kinetics of ivermectin and its metabolite were followed over 15 days by HPLC analysis. RESULTS: Co-administered ketoconazole induced a higher plasma concentration and longer residence time of ivermectin in dogs, leading to a substantial increase in the overall exposure of the animal to the drug. Ketoconazole does not interfere with the production of the ivermectin metabolite but it may rather inhibit the elimination of the parental drug by interfering with P-gp transport. CONCLUSIONS: Multiple oral dosing of ketoconazole dramatically altered the pharmacokinetics of ivermectin in dogs leading to an increase in systemic exposure to the drug. Neurotoxicity of ivermectin means that inhibition of the P-gp function at the blood-brain barrier during polytherapy using P-gp inhibitors must be taken into consideration.

Long-lasting protection against canine visceral leishmaniasis using the LiESAp-MDP vaccine in endemic areas of France: double-blind randomised efficacy field trial.

Vaccination against visceral leishmaniasis has received limited attention compared with cutaneous leishmaniasis, although the need for an effective vaccine against visceral leishmaniasis is pressing. Dogs constitute the major reservoir of Leishmania infantum/chagasi responsible for human visceral leishmaniasis. We have recently demonstrated that the combination of naturally excreted/secreted antigens, easily purified from culture supernatant of Leishmania infantum promastigotes (LiESAp) as vaccine antigen in formulation with muramyl dipeptide (MDP) as adjuvant, conferred 100% protection to dogs experimentally infected with L. infantum by inducing in vaccinees a significant, stable and long-lasting Th1-type cell response [Lemesre JL, Holzmuller P, Cavaleyra M, Bras Gonçalves R, Hottin G, Papierok G. Protection against experimental visceral leishmaniasis infection in dogs immunised with purified excreted secreted antigens of L. infantum promastigotes. Vaccine 2005; 23:2825-2840; Holzmuller P, Cavaleyra M, Moreaux J, Kovacic R, Vincendeau P, Papierok G, Lemesre JL. Lymphocytes of dogs immunised with purified excreted secreted antigens of L. infantum co-incubated with Leishmania-infected macrophages produce IFN-gamma resulting in nitric oxide-mediated amastigote apoptosis. Vet. Immunol. Immunopathol. 2005, 106:247-257]. In this report, protection against visceral leishmaniasis is investigated in naturally exposed dogs of endemic areas of the South of France vaccinated with LiESAp/MDP vaccine. A double-blind randomised efficacy field trial was developed on a large-scale dog population composed of vaccinees (n=205) and placebo-treated animals (n=209), which were prospectively studied for a 2-year period. 0f the initial 414 enrolled dogs, 340 (175 controls and 165 vaccinees) were analysed for clinical, serological and parasitological studies at 24 months post-vaccination, after two sand fly seasons. Strong seroconversion disclosed by an L. infantum indirect immunofluorescence antibody test (IFAT) associated with suspicious clinical symptoms, considered an indication that the animals had an established progressive infection, was only observed in the placebo group. The seropositive and/or symptomatic dogs were selected for further examination for possible Leishmania infection by culturing parasites from bone-marrow aspirate. The presence of leishmanial infection was also evaluated by means of the PCR analysis of bone marrow samples in all enrolled dogs prior to vaccination and in all evaluated animals (175 controls and 165 vaccinees) at 24 months post-vaccination. After two transmission cycles completed, the Leishmania infection rate was 0.61% (1/165) in vaccinated dogs and 6.86% (12/175) in the placebo group. The efficacy of the vaccine was calculated to be 92% (P=0.002). A clear difference between the dogs that received vaccine and those that received placebo was also established by the results of their immune status. Increased anti-LiESAp IgG2 reactivity and significant enhanced NO-mediated anti-leishmanial activity of canine macrophages in response to higher IFN-gamma production by T cells were almost exclusively revealed in vaccinees. The LiESAp-MDP vaccine induced a significant, long-lasting and strong protective effect against canine visceral leishmaniasis in the field.