Control Of Canine Leishmaniasis Research Articles

Impact of Vaccination, Insecticide-Impregnated Collar, and Treatment on the Canine Leishmaniasis

Leishmaniasis is a parasite disease transmitted by the bites of sandflies. There have been found more than 70 animal species that are natural reservoir hosts of Leishmania. Among the reservoirs, dogs are the most important ones due to their proximity to the human habitat. In this work, we formulate a model to assess the impact of vaccination, insecticide-impregnated collar use, and treatment on the control of canine leishmaniasis. To this end, we calculated the Basic Reproduction Number of the disease and carried out a sensitivity analysis of this parameter concerning the epidemiological and demographic parameters. The numerical simulations show the correlation between the disease prevalence and the strategies' effectiveness. Control of infection on dogs can be obtained by protecting around 35 percent of dogs with vaccination and insecticide-impregnated collar.

Serodiagnosis of leishmaniasis in asymptomatic and symptomatic dogs by use of the recombinant dynamin-1-like protein from Leishmania infantum: A preliminary study

Visceral leishmaniasis (VL) is a fatal manifestation of an infection caused by intracellular protozoa of the Leishmania genus. In New World countries, VL is classified as a zoonotic disease with domestic dogs acting as its main reservoir. Asymptomatic dogs are as competent to transmit Leishmania to the vectors as symptomatic dogs, however current diagnostic tests are limited and present low sensitivity for this important group. The development of accurate tests is fundamental to the early diagnosis, treatment, and control of canine leishmaniasis. In this study, we investigated the use of a recombinant protein (dynamin-1-like protein, Dyn-1) from L. infantum, as a potential target antigen for leishmaniasis serodiagnosis in both symptomatic and asymptomatic dogs. The antigenic performance of the protein was evaluated by means of ELISA assays using sera from symptomatic (n = 25), asymptomatic (n = 34) and non-infected dogs (n = 36) using ELISA. In addition, sera from dogs experimentally infected with Trypanosoma cruzi (n = 49) and naturally infected with Babesia sp. (n = 8) were tested to evaluate possible cross-reactivity. A crude soluble antigen (CSA) of Leishmania was used as an antigen control and K39 and K26 were used as reference antigens because they are already widely used in commercial tests. rDyn-1-based assay showed the highest sensitivity (97%) compared to the antigens K39 (88%), K26 (86%) and crude extract (95%). The highest specificity among the tests was also obtained with the protein rDyn-1 (94%), compared with the other antigens K39 (81%), K26 (87%), and crude extract (77%). This study showed that the rDyn-1 ELISA assay was able to identify 100% of asymptomatic dogs, establishing its potential as a target for the diagnosis of canine leishmaniasis.

Detection of Leishmania tarentolae in lizards, sand flies and dogs in southern Italy, where Leishmania infantum is endemic: hindrances and opportunities

BackgroundLeishmania tarentolae is a protozoan isolated from geckoes (Tarentola annularis, Tarentola mauritanica), which is considered non-pathogenic and is transmitted by herpetophilic Sergentomyia spp. sand flies. This species occurs in sympatry with Leishmania infantum in areas where canine leishmaniasis is endemic. In the present study, we investigated the circulation of L. tarentolae and L. infantum in sand flies, dogs and lizards in a dog shelter in southern Italy, where canine leishmaniasis by L. infantum is endemic.MethodsSheltered dogs (n = 100) negative for Leishmania spp. (March 2020) were screened by immunofluorescence antibody test (IFAT) using promastigotes of both species at two time points (June 2020 and March 2021). Whole blood from dogs, tissues of Podarcis siculus lizards (n = 28) and sand flies (n = 2306) were also sampled and tested by a duplex real-time PCR (dqPCR). Host blood meal was assessed in sand flies by PCR.ResultsOverall, 16 dogs became positive for L. infantum and/or L. tarentolae by IFAT at one or both sampling periods. One canine blood sample was positive for L. infantum, whilst two for L. tarentolae by dqPCR. At the cytology of lizard blood, Leishmania spp. amastigote-like forms were detected in erythrocytes. Twenty-two tissue samples, mostly lung (21.4%), scored molecularly positive for L. tarentolae, corresponding to 10 lizards (i.e., 35.7%). Of the female Sergentomyia minuta sampled (n = 1252), 158 scored positive for L. tarentolae, four for L. infantum, and one co-infected. Two Phlebotomus perniciosus (out of 29 females) were positive for L. tarentolae. Engorged S. minuta (n = 10) fed on humans, and one P. perniciosus, positive for L. tarentolae, on lagomorphs.ConclusionsDogs and lacertid lizards (Podarcis siculus) were herein found for the first time infected by L. tarentolae. The detection of both L. tarentolae and L. infantum in S. minuta and P. perniciosus suggests their sympatric circulation, with a potential overlap in vertebrate hosts. The interactions between L. tarentolae and L. infantum should be further investigated in both vectors and vertebrate hosts to understand the potential implications for the diagnosis and control of canine leishmaniasis in endemic areas.Graphical abstract

Longitudinal monitoring of anti-saliva antibodies as markers of repellent efficacy against Phlebotomus perniciosus and Phlebotomus papatasi in dogs.

A 2-year longitudinal study of enzyme-linked immunosorbent assay (ELISA) antibodies against Phlebotomus perniciosus and Phlebotomus papatasi (Diptera: Psychodidae) sandfly saliva was performed in 32 Beagle dogs treated preventively with an imidacloprid-permethrin topical insecticide in an endemic area in Spain. Dogs were grouped into three sandfly exposure groups according to the time of inclusion in the study. Assays analysed immunoglobulin G (IgG) against salivary gland homogenates (SGH) of both species and recombinant P. papatasi rSP32 and P. perniciosus rSP03B proteins in serum. The dogs were participating in a Leishmania infantum (Kinetoplastida: Trypanosomatidae) vaccine trial and were experimentally infected with the parasite in the second year. No dog acquired natural L. infantum infections during the first year, but most developed anti-saliva antibodies, and median log-transformed optical densities (LODs) were seasonal, mimicking those of local sandflies. This indicates that the repellent efficacy of the insecticide used is below 100%. Multi-level modelling of LODs revealed variability among dogs, autocorrelation and differences according to the salivary antigen and the dog's age. However, dog seroprevalence, estimated using pre-exposure LODs as cut-offs, was relatively low. This, and the fact that dogs did not become naturally infected with L. infantum, would support the efficacy and usefulness of this imidacloprid-permethrin topical insecticide in canine leishmaniasis control.

Canine Leishmaniasis Progression is Associated with Vitamin D Deficiency

The relationship between vitamin D deficiency and the risk of suffering from a plethora of health disorders, ranging from autoimmune processes to infectious diseases has been widely described. Nonetheless, the potential role of vitamin D in visceral leishmaniasis remains uncharacterized. In the Mediterranean basin, where the dog is leishmania’s main peri-domestic reservoir, control measures against the canine disease have shown beneficial effects on the incidence of human leishmaniasis. In this study, we measured the vitamin D levels in serum samples from a cohort of 68 healthy and disease dogs from a highly endemic area and we have also studied the relationship of these levels with parasitological and immunological parameters. The sick dogs presented significantly lower (P < 0.001) vitamin D levels (19.6 ng/mL) than their non-infected (31.8 ng/mL) and the asymptomatic counterparts (29.6 ng/mL). In addition, vitamin D deficiency correlated with several parameters linked to leishmaniasis progression. However, there was no correlation between vitamin D levels and the Leishmania-specific cellular immune response. Moreover, both the leishmanin skin test and the IFN-γ levels displayed negative correlations with serological, parasitological and clinical signs. Further studies to determine the functional role of vitamin D on the progression and control of canine leishmaniasis are needed.

Xenodiagnosis on dogs with visceral leishmaniasis: Canine and sand fly aspects related to the parasite transmission

One of the main limitations for the effective control of canine leishmaniasis in endemic areas is the difficulty in identifying infectious dogs. The objective of this study was to determine factors, related to dogs and to parasite detection in sand flies, which are associated with the positive xenodiagnosis of Leishmania infantum using the vector Lutzomyia longipalpis. The xenodiagnosis was performed in 50 owned dogs residing in endemic areas, which were divided into three different groups: G1—26 dogs proved to be infected and classified by severity of VL clinical signs on physical examination; G2—15 dogs proved to be infected and classified by severity of clinical signs and intensity of laboratory abnormalities; G3—nine dogs that were seropositive for anti-Leishmania IgG in ELISA tests. Parasite search in the sand flies after having fed on dogs in the xenodiagnosis was performed by both methodologies, PCR and dissection followed by microscopy. In G1, 58% (15/26) of dogs were able to transmit Leishmania to the vector, when parasite detection in sand flies were performed by PCR technique, 5 days after blood meal, whereas in G2, 53% (8/15) transmitted the parasite to the vector, however, confirmation was performed by direct observation of parasite through optical miscroscopy held 10 days after blood meal. Rate of infectiousness of dogs to sand flies was positively associated to severity of disease (p=0.042 and p=0.040), regardless the method used for clinical classification or for parasite detection in sand flies after xenodiagnosis. In G1 30% (3/10) of dogs with subclinical infection were infectious to the vector, while 80% (12/16) of dogs with clinical disease were also infectious. Even more, 17% (1/6) of dogs that had moderate disease were infectious to the sand flies, while 78% (7/9) of dogs with severe disease were infectious in G2. Still in G2, the proportion of sand flies infected (grade of infectiousness) was significantly lower (p=0.0098) when they fed on dogs with moderate disease (1%) in comparison with dogs with severe disease (38%). The dogs from G3 presented a rate of infectiousness of 11% (1/9), demonstrating that the indirect ELISA is not a good indicator of infectiousness and, therefore, should not be used as a confirmatory test for the euthanasia of dogs, as it is currently done in Brazil.

The protective immune response produced in dogs after primary vaccination with the LiESP/QA-21 vaccine (CaniLeish®) remains effective against an experimental challenge one year later.

Control of canine leishmaniasis is an important objective for the benefit of dogs living in or visiting endemic areas and for public health because of the zoonotic nature of this disease. Resistance or susceptibility to developing canine leishmaniasis after exposure to Leishmania infantum is primarily determined by the ability of the immune system to develop an appropriate Th1-dominated specific response to the parasite. For this reason there is a need for effective canine vaccines that can decrease the number of dogs developing progressive infections. In this study, we followed the impact of the LiESP/QA-21 canine vaccine (composed of excreted-secreted proteins of L. infantum and the QA-21 saponin adjuvant), recently launched commercially in Europe, on selected humoral and cellular immune parameters following an infectious intravenous challenge with L. infantum promastigotes administered one year after the primary vaccine course. We also followed parasitological parameters to determine the parasitological status of the challenged dogs. In contrast to controls, vaccinated dogs retained significantly stronger cell-mediated immune responses against the parasite despite a virulent challenge and had significantly lower mean parasite burdens at the end of the study, associated with a lower probability of developing active infections. These results confirm that the immune responses generated by vaccination with LiESP/QA-21 are still effective against an intravenous challenge one year after the primary vaccine course.

Occurrence of Trypanosoma caninum in areas overlapping with leishmaniasis in Brazil: what is the real impact of canine leishmaniasis control?

Trypanosoma caninum is a parasite of the Trypanosoma genus recently described in the natural infection of dogs in the municipality of Rio de Janeiro, Brazil. Suspecting the existence of a natural cycle and the circulation of this new species, the objective of this study was the taxonomic identification of samples of Trypanosoma spp. isolated from dogs in different Brazilian regions. Parasites were solely obtained from skin fragments culture and characterized by nested-PCR targeting the partial sequence of 18S rRNA gene and PCR products were sequenced. Thirty-three samples, obtained in São Paulo, Minas Gerais, Goiás, Mato Grosso and Rio de Janeiro states were analyzed. PCR and sequencing showed that the isolates were genetically identical or closely similar and confirmed T. caninum identity. This report broadens the geographical distribution of T. caninum in Brazil and discusses the impact of the presence of this parasite in areas of canine leishmaniasis occurrence.

Genetic control of canine leishmaniasis: genome-wide association study and genomic selection analysis.

BackgroundThe current disease model for leishmaniasis suggests that only a proportion of infected individuals develop clinical disease, while others are asymptomatically infected due to immune control of infection. The factors that determine whether individuals progress to clinical disease following Leishmania infection are unclear, although previous studies suggest a role for host genetics. Our hypothesis was that canine leishmaniasis is a complex disease with multiple loci responsible for the progression of the disease from Leishmania infection.Methodology/Principal FindingsGenome-wide association and genomic selection approaches were applied to a population-based case-control dataset of 219 dogs from a single breed (Boxer) genotyped for ∼170,000 SNPs. Firstly, we aimed to identify individual disease loci; secondly, we quantified the genetic component of the observed phenotypic variance; and thirdly, we tested whether genome-wide SNP data could accurately predict the disease.Conclusions/SignificanceWe estimated that a substantial proportion of the genome is affecting the trait and that its heritability could be as high as 60%. Using the genome-wide association approach, the strongest associations were on chromosomes 1, 4 and 20, although none of these were statistically significant at a genome-wide level and after correcting for genetic stratification and lifestyle. Amongst these associations, chromosome 4: 61.2–76.9 Mb maps to a locus that has previously been associated with host susceptibility to human and murine leishmaniasis, and genomic selection estimated markers in this region to have the greatest effect on the phenotype. We therefore propose these regions as candidates for replication studies. An important finding of this study was the significant predictive value from using the genomic information. We found that the phenotype could be predicted with an accuracy of ∼0.29 in new samples and that the affection status was correctly predicted in 60% of dogs, significantly higher than expected by chance, and with satisfactory sensitivity-specificity values (AUC = 0.63).

Diagnosis of canine leishmaniasis: Conventional and molecular techniques using different tissues

Serology, bone marrow (BM)-, lymph node (LN)- and whole blood-polymerase chain reaction (PCR) were evaluated as potential reference tests for the diagnosis of canine leishmaniasis. A high degree of agreement (⩾91.0%) was observed between Leishmania cultures and serology or BM/LN-PCR. In the light of these results as well as the access to biological test material and the cost of each method, LN-PCR is recommended for the diagnosis or therapeutic control of canine leishmaniasis, but BM-PCR is a suitable alternative in dogs without detectable adenomegaly. For large-scale epidemiological field studies, antibody detection is appropriate and whole blood-PCR can be used to complement the serological results.

PCR identification of Leishmania in diagnosis and control of canine leishmaniasis

Leishmaniases are endemic in many countries, mainly in rural areas. In Brazil, Leishmania infection is responsible for many cases of Leishmaniases, including recent reports in urban regions. Despite their sensitivity, traditional serological and parasitological methods for detecting Leishmaniases have proven inadequate for species discrimination. This study aimed to identify Leishmania species in biological samples by a fast methodology, avoiding “in vitro” cultivation. Knowledge of the Leishmania species is an important tool in regions where both New World visceral leishmaniasis (VL) and cutaneous leishmaniasis (CL) are prevalent. As these new foci appear in areas not traditionally endemic for VL, the main problem is to distinguish between true autochthonous infections and infections acquired in other well-known endemic areas. Since, domestic dogs are known to be the main VL and CL reservoir, they are regularly investigated in endemic areas to prevent, principally, severe and often fatal VL in humans. However, several infected dogs present no clinical signs or clinical signs similar to other canine diseases. Here, we evaluated the ability of PCR to diagnose VL and distinguish L. ( L.) chagasi from other Leishmania species in domestic dogs. Samples from 114 dogs from 30 cities (Sao Paulo, Brazil) were divided into two groups: 44 symptomatic and 70 asymptomatic. They were assayed by parasitological methods (culture and microscopic examination) and PCR to determine L. ( L.) chagasi, L. ( V.) braziliensis; and in some cases, Leishmania spp. Parasitological tests and PCR- L. chagasi were concordant in 105 samples (92%). VL was confirmed in 49 dogs, while 56 had negative results. Of the 114 samples, 9 had discordant results, but were further tested by PCR- Leishmania spp. with positive results. VL was also confirmed in 4 dogs having negative parasitological tests and positive PCR- L. chagasi. Consequently, this PCR was positive for 100% (53/49) of dogs with parasites detected in parasitological tests. Also, PCR demonstrated high specificity detecting 61 dogs negative for VL. Leishmania infection was negative in 56 dogs, and 5 with positive culture and PCR- Leishmania spp. had CL since they were positive in PCR- L. braziliensis. This study shows the importance of including PCR in diagnosis of Leishmaniases by differential diagnosis contributing to the surveillance and control of VL programs.

Deltamethrin-impregnated collars for the control of canine leishmaniasis: Evaluation of the protective effect and influence on the clinical outcome of Leishmania infection in kennelled stray dogs

A 2-year field study on kennelled stray dogs living in a highly endemic area of leishmaniasis was designed to evaluate whether deltamethrin-impregnated collars (Scalibor ® Protector Band) could confer protection against leishmaniasis in this peculiar setting, and to assess differences in clinical outcomes between collared and uncollared dogs. A cohort of 120 clinically healthy and Leishmania-seronegative dogs was enrolled, 50% of which were collared before the 2003 transmission season, and then re-collared before the subsequent season. Collared and uncollared animals were allowed to live with infected dogs in same groups within the kennel. Follow-up included serological (IFAT) assessment twice a year with parasitological Leishmania confirmation, and clinical evaluation performed every 3 months on seroconverted dogs from both groups. Collar losses during the two seasons were high (35%). About 50% of enrolled dogs were lost at follow-up because of death or they were moved to other locations. After the 2003 season, cross-sectional serological examinations tested positive in 5 out of 44 collared animals (11.4%) and in 14 out of 34 controls (41.2%), with 72.3% estimated protection ( P < 0.005). After the 2004 season, 7/31 seronegative collared dogs seroconverted (22.6%) compared with 7/17 seronegative controls (41.2%), with 45.1% protection ( P = 0.15). At the end of the study, the cumulative rate of protection was 50.8% ( P = 0.005). At the clinical evaluation of 21 seroconverted dogs from both groups, canine leishmaniasis signs were significantly more frequent (90% versus 36%, P = 0.017) and rapidly progressive in uncollared than in collared dogs. Reasons for such partial clinical protection in collared dogs may be found in the vector anti-feeding effect of protector bands, resulting in a lower number of infectious bites and, probably, in the reduction of antigenic stimuli necessary to shift toward a non-protective immune response.

Canine visceral leishmaniasis: Comparison of in vitro leishmanicidal activity of marbofloxacin, meglumine antimoniate and sodium stibogluconate

The control of canine leishmaniasis largely depends on the success of treatment. Drugs currently available to treat this disease are toxic and partially effective. The curative effect of marbofloxacin, a third-generation fluoroquinolone developed for veterinarian individual treatment, was evaluated in vitro in the presence of Leishmania infantum promastigotes and dog-monocyte-derived macrophages; meglumine antimoniate and sodium stibogluconate were used as comparative treatments. We observed that the killing of Leishmania promastigotes and intracellular amastigotes by marbofloxacin was dose-dependent. We demonstrated that successful treatment of canine infected macrophages for 48 h was possible with 500 μg/ml of marbofloxacin. Leishmanicidal activity acted through a TNF-α and nitric oxide pathway and correlated with the generation of nitric oxide (NO 2) production by monocytes derived macrophages from infected (23 ± 5 μM) or healthy (21 ± 6 μM) dogs, in comparison with NO 2 concentration in infected/non-treated macrophages (<3 μM, P < 0.01). This significant induced parasiticidal effect correlated with extensive elimination of amastigotes by macrophages derived from infected (11 ± 5) and healthy dogs (6 ± 2), when compared to infected/non-treated macrophages (530 ± 105 and 472 ± 86 amastigotes, respectively, P < 0.01). Marbofloxacin was shown to be non-toxic at 500 μg/ml in vitro and no cell apoptosis was observed. The molecule was able to induce a parasitic process after significant elimination of amastigotes in leishmania-infected dog macrophages. We propose that marbofloxacin, compared to standard chemotherapeutic agents (meglumine antimoniate and sodium stibogluconate), could be an effective and pragmatic oral route alternative to treat canine leishmaniasis.

Protection of dogs from bites of phlebotomine sandflies by deltamethrin collars for control of canine leishmaniasis.

Dog collars made of PVC plastic impregnated with the pyrethroid insecticide deltamethrin at 40 mg/g were investigated for their protective efficacy against phlebotomine sandflies. Collared dogs were kept separately (two untreated control dogs lived together) in outdoor enclosures, each with a kennel, in the Cévennes, southern France. To measure sandfly mortality and anti-feeding effects due to the deltamethrin-impregnated collars worn continuously by the dogs for up to 8 months, each dog was periodically sedated and exposed for 2h to 150-200 laboratory-reared Phlebotomus perniciosus females (plus c. 25 males) inside a net (1.2 m square, 1.8 m high) indoors. After dogs were removed from the nets, allowed to recover and returned to their kennels, any dead sandflies were collected from inside the net and counted. Surviving flies were kept overnight, then scored according to whether they were still alive or dead, unfed or blood-fed. From tests 2, 3, 4, 13, 20, 26 and 34 weeks after the dogs began wearing collars, the overall numbers of blood-fed female sandflies recaptured were 75 from two dogs with collars, compared with 1911 from two collarless dogs. Thus, for every 100 flies which fed on collarless dogs, only 4 fed on collared dogs, i.e. the collars protected dogs from 96% of the bites and this activity was maintained for up to 34 weeks. During the same period, the percentage of recaptured female sandflies that had fed on collared dogs was 0-12% compared to 55-95% on collarless dogs. Immediately after dogs were taken out of the nets, 21-60% of flies confined with the collared dogs were found dead, compared to 0-12% with the controls. It is concluded that, at least in the Mediterranean subregion, this insecticidal collar would protect a dog from the majority of sandfly bites and retain a killing effect for a complete sandfly season. Moreover, it seems likely that the use of collars on all dogs in a focus of Leishmania infantum would reduce contact between sandfly vectors and canine reservoir hosts sufficiently to diminish the risk of infection for humans as well as dogs.

Studies on canine leishmaniasis control. 1. Evolution of infection of different clinical forms of canine leishmaniasis following antimonial treatment

81 dogs naturally infected with Leishmania infantum in the Isle of Elba, Italy, were treated with meglumine antimoniate (Glucantime ®). 36 of them (45 · 5%) were asymptomatic cases. 4–24 months after treatment the dogs were clinically and serologically examined; the recovery rates were 47 · 2% for asymptomatic cases, 33 · 3% for oligosymptomatic cases, and 11 · 1% for symptomatic cases. Furthermore, treatment had prevented the development of patent disease in 90% of non-recovered asymptomatic cases, whereas it had produced only slight improvement of clinical condition in patent dogs which were still infected after drug administration. Treatment with antimonial drugs is therefore recommended in canine leishmaniasis control if non-patent or sub-clinical forms of the disease are detected by seroepidemiological surveys.

Studies on canine leishmaniasis control. 2. Effectiveness of control measures against canine leishmaniasis in the Isle of Elba, Italy

A serological and parasitological survey carried out in 1985 in the Isle of Elba, Italy, revealed a high prevalence of canine leishmaniasis. This focus was considered ideal for the evaluation of effectiveness of treating infected dogs with meglumine antimoniate, applied as a control measure during 1985 and 1986. New data on prevalence were obtained for the years 1986 and 1987. Incidence of new canine leishmaniasis cases after the transmission seasons 1984, 1985 and 1986 were determined by examining 2 cohorts: cohort I, dogs born within 2 transmission seasons, and cohort II, adult dogs examined and found to be negative before each transmission season. Over 2000 tests were carried out on 1500 dogs. Prevalence analysis showed a reduction of infective dogs (symptomatic and oligosymptomatic) from 14 · 4% in 1985 to 5 · 2% in 1987. Incidence analysis showed a decrease of new cases from 12 · 4% after transmission season 1984 to 4 · 6% after transmission season 1987. The results indicate a two-thirds reduction of the disease frequency in dogs of the Isle of Elba after a 2-year period of control measures.

Leishmania infantum infection rates in Phlebotomus perniciosus fed on naturally infected dogs under antimonial treatment.

Dogs naturally infected with Leishmania infantum Nicolle were treated with three courses of meglumine antimoniate. Changes were observed in the dogs' clinical signs, antibody titres and in infection rates of Phlebotomus perniciosus Newstead fed on the dogs. A large reduction in the sandfly infection rate was observed for 4-5 months after the first treatment. The use of antimonial drugs is advocated for the control of canine leishmaniasis and to reduce risks of L.infantum transmission.

Canine leishmaniasis in the Mediterranean area and its implications for human leishmaniasis

The distribution of canine leishmaniasis and figures on infection frequency in countries bordering the Mediterranean Sea are given. The clinical picture and most common laboratory findings are reviewed. Leishmania infantum is the aetiological agent of both human and canine forms of visceral leishmaniasis. The proved or suspected vectors of L. infantum are listed and mention is made of wild animals (fox, rat) as possible primary reservoirs of the parasite. Finally, the methods of investigation and control of canine leishmaniasis are discussed.