Interaction between Wolbachia pipientis and Leishmania infantum in heartworm infected dogs.

Background: Wolbachia is a Gram-negative endosymbiont associated with several species of arthropods and filarioid nematodes, including Dirofilaria immitis. This endosymbiont may elicit a Th1 response, which is a component of the immunity against Leishmania infantum.

Methods: To investigate the interactions between Wolbachia of D. immitis and L. infantum in naturally infected dogs and cytokine circulation, dogs without clinical signs (n = 187) were selected. Dogs were tested for microfilariae (mfs) by Knott, for female antigens of D. immitis by SNAP, and for anti-L. infantum antibodies by IFAT and assigned to four groups. Dogs of group 1 (G1) and 2 (G2) were positive for D. immitis and positive or negative to L. infantum, respectively. Dogs of group 3 (G3) and 4 (G4) were negative to D. immitis and positive or negative to L. infantum, respectively. Wolbachia and L. infantum DNA was quantified by real-time PCR (qPCR) in dog blood samples. A subset of dogs (n = 65) was examined to assess pro- and anti-inflammatory cytokine production using an ELISA test.

Results: Of 93 dogs positive to D. immitis with circulating mfs, 85% were positive to Wolbachia, with the highest amount of DNA detected in G1 and the lowest in dogs with low mfs load in G1 and G2. Among dogs positive to L. infantum, 66% from G1 showed low antibody titer, while 48.9% from G3 had the highest antibody titer. Of 37 dogs positive to Wolbachia from G1, 26 (70.3%) had low antibody titers to L. infantum (1:160). Among cytokines, TNFα showed the highest mean concentration in G1 (246.5 pg/ml), IFNγ being the one most represented (64.3%). IL-10 (1809.5 pg/ml) and IL-6 (123.5 pg/ml) showed the highest mean concentration in dogs from G1. A lower percentage of dogs producing IL-4 was observed in all groups examined, with the highest mean concentration (2794 pg/ml) recorded in G2.

Conclusion: Results show the association of D. immitis and Wolbachia with the lower antibody titers of L. infantum in co-infected dogs, suggesting the hypothesis that the endosymbiont may affect the development of the patent leishmaniosis. However, due to the limitations associated with the heterogeneity of naturally infected dogs in field conditions, results should be validated by investigation on experimental models.