Study of ocular manifestations and humoral immune response in eyes of dogs with leishmaniasis.

BACKGROUND: Ocular manifestations in dogs with leishmaniasis are frequent and complications in affected tissues can lead to blindness. Immune processes play a very important role in the pathogenesis of ocular inflammation. Therefore, the immunology of ocular manifestations in dogs with leishmaniasis remains complex and poorly understood. OBJECTIVES: Estimation and characterisation of ocular and periocular manifestations in dogs naturally infected with Leishmania infantum and investigation of the production site of specific anti-Leishmania infantum IgG. METHODS: The present investigation used 53 confirmed dogs infected with Leishmania infantum, presenting ocular and periocular lesions, and 10 control non-infected dogs. Complete macroscopic ophthalmic examination of eyelids and globes was performed. Both total and anti-Leishmania infantum IgG antibodies were studied in sera and aqueous humour (AH) of all dogs by ELISA technique. A Goldmann-Witmer coefficient (C value) was calculated. RESULTS: The main ophthalmological findings were keratoconjunctivitis (71.7%; 38/53), hyperplasia of conjunctival lymphoid follicles (54.7%; 29/53), blepharitis (50.9%; 27/53) and uveitis (20.7%; 11/53). Ocular production of anti-Leishmania infantum IgG was detected in 73.6% (39/53) of infected dogs. There was no correlation between the antibody levels in AH and sera of the same dog. The mean anti-Leishmania infantum IgG in AH was higher in uveitis, followed by lesions affecting only the adnexa (p < 0.0001). The highest mean C values were observed for uveitis, conjunctivitis and keratitis. CONCLUSIONS: Our findings suggest that production of anti-Leishmania IgG in dogs infected with Leishmania infantum with ocular manifestations begin in situ and follows by a transfer of antibodies from the bloodstream to the AH.

Use of an Interferon Gamma Release Assay (IGRA) to test T-cell responsiveness to soluble Leishmania infantum antigen in whole blood of dogs from endemic areas.

Interferon-Gamma (IFN-γ) Release Assays (IGRAs) are easy tests that allow rapid screening of primed memory T-cells immunity in response to antigen. The aim of this study was to use IGRA to assess IFN-γ release in response to Soluble Leishmania infantum antigen (SLA) in whole blood of dogs living in endemic area of visceral leishmaniasis and to interpret IGRA results according to clinical examination, specific anti-Leishmania humoral response and presence of L. infantum DNA in blood. The study was carried out on 56 dogs living in greater Tunis area. Physical examination, quantitative serology and PCR on blood were used to characterize dogs' status in relation to Leishmania infection and disease. IGRA consisted on testing by ELISA for IFN-γ-secretion in whole blood after a 20-h challenge with SLA. PBS and Phytohemagglutinin (PHA) stimulations were used as controls. Four groups of dogs were characterized: 31 were negative by both serology and PCR, two had doubtful serology, 10 presented no to mild clinical signs but low antibodies levels and 13 were affected by Canine Leishmaniasis (CanL). In seronegative dogs, IGRA was little contributory in 4 puppies (age <6months) and 5 old dogs (median age=72months, IQR: 45-84 months) that didn't respond to PHA stimulation, IGRA was negative in 19 and positive in three animals with lymph node enlargement. In dogs with doubtful serology, IGRA was positive in one dog and negative in the other. In infected dogs with no to mild clinical signs, one dog exhibited high level of IFN-γ in absence of antigenic stimulation and all the other were positive by IGRA. CanL dogs showed variable IGRA results. Negative IGRAs (n=4) were shown in animals with the highest parasitic burden whereas positive IGRAs (n=5) were shown in dogs with negative PCR or low parasitic load. The 4 remaining dogs either didn't respond to PHA (n=2) or showed non-specific secretion in PBS tube (n=2). The results of this study showed that IGRA is a useful new tool that can assess exposure to Leishmania in dogs with no to mild clinical signs in endemic area. Further comparative investigations using assays exploring cellular immunity are needed to determine its accuracy.