Prevention of transmission of Borrelia burgdorferi sensu lato and Anaplasma phagocytophilum by Ixodes spp. ticks to dogs treated with the Seresto® collar (imidacloprid 10% + flumethrin 4.5%).

The capability of imidacloprid 10% + flumethrin 4.5% (Seresto®) collars to prevent transmission of Borrelia burgdorferi sensu lato (Bbsl) and Anaplasma phagocytophilum (Ap) by naturally infected ticks was evaluated in two studies with 44 dogs. In each study, one group served as non-treated control, whereas the other groups were treated with the Seresto® collar. All dogs were exposed to naturally Bbsl- and Ap-infected hard ticks (Ixodes ricinus, Ixodes scapularis). In study 1, tick infestation was performed on study day (SD) 63 (2 months post-treatment [p.t.]); in study 2, it was performed on SD 32 (one month p.t.) respectively SD 219 (seven months p.t.). In situ tick counts were performed 2 days after infestation. Tick counts and removals followed 6 (study 1) or 5 days (study 2) later. Blood sampling was performed for the detection of specific Bbsl and Ap antibodies and, in study 1, for the documentation of Ap DNA by PCR. Skin biopsies were examined for Bbsl by PCR and culture (only study 1). The efficacy against Ixodes spp. was 100% at all time points. In study 1, two of six non-treated dogs became infected with Bbsl, and four of six tested positive for Ap; none of the treated dogs tested positive for Bbsl or Ap. In study 2, ten of ten non-treated dogs became infected with Bbsl and Ap; none of the treated dogs tested positive for Bbsl or Ap; 100% acaricidal efficacy was shown in both studies. Transmission of Bbsl and Ap was successfully blocked for up to 7 months.

Efficacy of four commercially available heartworm preventive products against the JYD-34 laboratory strain of Dirofilaria immitis.

BACKGROUND: Heartworm disease in dogs can be severe and life threatening. Resistance to available heartworm preventives was considered among potential causes of increased reports of failed heartworm prevention in dogs. The objective of the present study was to compare the efficacy of four commercially available heartworm disease preventives against the JYD-34 strain of D. immitis. METHODS: Forty laboratory-reared dogs approximately 6 months old were used. Each dog was infected with fifty, third-stage heartworm larvae on study day (SD) -30. On SD-1, the dogs were randomized to five groups of eight dogs each. On SD-0, dogs in groups 1-4 were treated as follows: Group 1: ivermectin/pyrantel pamoate chewable tablets; Group 2: milbemycin oxime/spinosad tablets; Group 3: selamectin topical solution; and Group 4: imidacloprid/moxidectin topical solution. Dogs in Group 5 were not treated and served as controls. The dogs were treated according to their current body weights and labelled dose banding for each product. Groups 1, 2, and 3 were retreated with their respective products and current body weights on SD 31 and 60. On SDs 124-126 the dogs were euthanized and necropsied for recovery of adult heartworms. RESULTS: Adult heartworms were recovered at necropsy from each of the dogs in the control group (13-32 worms/dog, geometric mean (GM) = 18.4 worms/dog). Adult heartworms and/or worm fragments were also recovered from each of the dogs treated with ivermectin/pyrantel pamoate, milbemycin oxime/spinosad or selamectin. Geometric means of worms recovered from dogs in each of these groups were 13.1, 8.8, and 13.1, resulting in efficacies compared to controls of 29.0, 52.2, and 28.8 %, respectively. All dogs in Group 4 (imidacloprid/moxidectin) were free of adult heartworms (100 % efficacy). CONCLUSIONS: The combination of imidacloprid/moxidectin was 100 % effective in this study in preventing development of JYD-34 laboratory strain of D. immitis in dogs following a single treatment, while three monthlytreatments of the three other commercial products provided less than 100 % efficacy. The high efficacy achieved with imidacloprid/moxidectin was likely due to the unique pharmacokinetic properties of the topical formulation delivering greater and sustained drug concentrations necessary to prevent development of D. immitis larvae.

Real-time PCR of the mammalian hydroxymethylbilane synthase (HMBS) gene for analysis of flea (Ctenocephalides felis) feeding patterns on dogs.

BACKGROUND: Precise data on quantitative kinetics of blood feeding of fleas, particularly immediately after contact with the host, are essential for understanding dynamics of flea-borne disease transmission and for evaluating flea control strategies. Standard methods used are inadequate for studies that simulate early events after real-life flea access to the host. METHODS: Here, we developed a novel quantitative polymerase chain reaction targeting mammalian DNA within fleas to quantify blood consumption with high sensitivity and specificity. We used primers and fluorescent probes that amplify the hydroxymethylbilane synthase (HMBS) gene, an evolutionary divergent gene that is unlikely to be detected in insects by mammalian-specific primers and probes. To validate this assay, fleas were placed on dogs, allowed to distribute in the hair, and removed at specific time points with single-use combs. Fleas were then immediately homogenized by vigorous shaking with ceramic beads in guanidinium-based DNA preservation buffer for DNA extraction. RESULTS: The specificity of this assay was ascertained by amplification of canine, feline and equine blood with differential product melting temperatures (Tm), and lack of amplification of bovine and porcine blood and of adult fleas reared from larvae fed with bovine blood. Sensitivity of the assay was established by limiting dilution and detection of single copies of HMBS DNA equivalent to 0.043 nL blood. Application of the assay indicated that after 15 minutes on a dog, male and female fleas had ingested low, but similar amounts of approximately 1.1. nL blood. Saturation uptake of 118 and 100 nL blood per flea was found at 30 and 60 min on the dog, respectively. CONCLUSIONS: The HMBS PCR method developed here offers the advantages of both exquisite sensitivity and specificity that make it superior to other approaches for quantification of blood ingested by fleas. The capability to detect minute quantities of blood in single fleas, particularly immediately after colonization of the host, will provide a superior tool for studying flea-host interactions, flea-borne disease transmission, and flea control strategies.

Age-related changes in cytokine expression by neutrophils of foals stimulated with virulent Rhodococcus equi in vitro.

Although evidence exists that neutrophils play a vital role in resistance to infection with Rhodococcus equi, the means by which neutrophils exert their effects have not been clearly defined. In the present study we evaluated differences in cytokine expression by unstimulated and R. equi-stimulated neutrophils obtained from newborn foals and subsequently at 2-, 4-, and 8-weeks of age. Stimulation with virulent R. equi induced significantly (P<0.05) greater expression of IFNgamma, TNFalpha, IL-6, IL-8, IL-12p40, IL-12p35, and IL-23p19 mRNA relative to expression by unstimulated neutrophils, and there were significant effects of age on expression of IL-6, IL-8, IL-12p40 and IL-23p19. Neutrophil expression of IL-6 and IL-8 in newborn foals was significantly greater than expression at 2-, 4-, and 8-weeks of age. Expression of IL-12p40 by R. equi-stimulated neutrophils from newborn and 2-week-old foals did not differ from that of unstimulated neutrophils; however, expression of IL-12p40 by neutrophils from 4- and 8-week-old foals was significantly greater when stimulated by R. equi than without stimulation. These results demonstrate that foal neutrophils increase mRNA expression of many pro-inflammatory cytokines, including IFNgamma, in response to in vitro stimulation with R. equi, and that the magnitude of this expression with respect to IL-6, IL-8, IL-12p40 and IL-23p19 is influenced by age. The clinical importance of the age-related difference in R. equi-induced expression of IL-12p40 to susceptibility to R. equi pneumonia remains to be determined.

Use of imidacloprid-permethrin to prevent transmission of Anaplasma phagocytophilum from naturally infected Ixodes scapularis ticks to dogs.

One group of eight beagles was treated with a combination of imidacloprid and permethrin 7 days before exposure to Ixodes scapularis ticks that were naturally infected with Anaplasma phagocytophilum. A second group of eight beagles was not treated and was also exposed to infected ticks. Seven of eight non-treated dogs--but none of the treated dogs--developed specific antibodies to A. phagocytophilum. Results of this study indicate that a combination of imidacloprid and permethrin can prevent transmission of A. phagocytophilum to dogs if administered before exposure to infected ticks.