ABSTRACT: Canine vector-borne disease transmission can be reduced with regular use of repellent insecticides. The objective of this year-long experimental study was to assess the efficacy of a topical formulation of fipronil/permethrin (Frontline Tri-Act®) in preventing transmission of Leishmania infantum by sandflies. This clinical field trial was conducted in Xanthi (Northern Greece), an area highly endemic for canine leishmaniosis, from April 2018 to March 2019. Forty purpose-bred Beagle dogs, testing negative for L. infantum prior to study initiation, were enrolled in the study, which consisted of three phases: Phase 1 (field exposure phase) took place from Day 0-196. The dogs were randomly allocated to two groups, group 1 (sham-treated topically with sterile water) and group 2 (treated topically with Frontline Tri-Act®). Dogs in both groups were housed in two subunits of an open-air kennel for a period of 7 months, spanning the Leishmania transmission season. All dogs were treated or sham-treated on Days 0, 28, 56, 84, 112, 140 and 168. Clinical examinations, PCR analysis of conjunctival swabs, and serological tests were performed on a monthly basis. Sandflies were collected every 2 weeks, during a 12 -h period overnight using light traps. Each collection was placed in a container and kept refrigerated until speciation and PCR analysis could be performed. In the second phase of the study, from Day 197 to 252, the dogs were moved into a protected environment (insect-proof protected environment phase). CDC light traps were activated every 2 weeks inside and outside the kennels to ensure the vector-free status of the facility. Monthly clinical examinations, including PCR analysis of conjunctival swabs, and serological tests continued. At the end of the phase 2, bone marrow samples were collected on all dogs. Phase 3 (the final post-winter check) took place from Day 253 to 350. Dogs were released and adopted by individual private owners on Day 253. Follow up analyses included blood collection for SNAP tests and conjunctival swaps for PCR analysis on Days 304 and 350. Additionally, bone marrow collections were also performed on Day 350. Presence of sandflies was observed only in the phase 1 exposure period, and 1714 sandflies were collected (1427 females and 287 males). Two species were identified, Phlebotomus perniciosus var. tobbi and Phlebotomus neglectus. Out of the 62 pooled samples of sandflies assessed by PCR, three were considered positive (4.8 %). By the end of the study, 35 % of the Group 1 dogs (7/20) became positive based on PCR (conjunctival swab and bone marrow) and 30 % (6/20) based on SNAP/IFAT and ELISA tests, while all the dogs in the Frontline Tri-Act® treated group 2 remained negative for all tests (G1 vs G2, p = 0.008). All tests identified the same positive animals, and PCR allowed the detection of one additional infected dog. This clinical field trial demonstrated that monthly administration of Frontline Tri-Act® to dogs exposed to Leishsmania infection in a high endemic area provided 100 % preventive efficacy against transmission of L. infantum.
