Canine leishmaniosis caused by L. infantum is a severe zoonotic disease. Although macrophages are the definitive host cells, neutrophils are the first cells to encounter the parasite soon after its inoculation in the dermis by the phlebotomine vector. To study the interaction of dog neutrophils and L. infantum promastigotes, blood neutrophils were isolated from healthy donors and the infection was established in vitro. In the majority of the dogs, L. infantum was efficiently phagocytized by neutrophils, and oxidative (superoxide production) and non-oxidative (neutrophil elastase exocytosis) intracellular effector mechanisms were activated, but the release of neutrophil extracellular traps was minimized. Furthermore, promastigotes and culture supernatants induced neutrophil migration, but the prior contact with Leishmania inhibits chemotaxis, which might contribute to neutrophil retention at the inoculation site. Neutrophil-parasite interaction resulted in a decrease in parasite viability, although some intracellular promastigotes survive and maintain their proliferative capacity. These findings indicate that dog neutrophils are competent effector cells able to control the initial L. infantum infection. However, some parasites evade intracellular effector mechanisms and can be transferred to the definitive host cell, the macrophage, contributing to the development of canine leishmaniosis.

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Canine leishmaniosis (CanL) caused by Leishmania infantum is a zoonotic disease of global concern. Antileishmanial drug therapies commonly used to treat sick dogs improve their clinical condition, although when discontinued relapses can occur. Thus, the current study aims to evaluate the effect of CanL treatments in peripheral blood, lymph node, and bone marrow cytokine profile associated with clinical recovery. Two groups of six dogs diagnosed with CanL were treated with miltefosine combined with allopurinol and meglumine antimoniate combined with allopurinol (MT+A and MG+A), respectively. At diagnosis and after treatment, during a 3-month follow-up, clinical signs, hematological and biochemical parameters, urinalysis results and antileishmanial antibody titers were registered. Furthermore, peripheral blood, popliteal lymph node, and bone marrow samples were collected to assess the gene expression of IL-2, IL-4, IL-5, IL-10, IL-12, TNF-α, TGF-β, and IFN-γ by qPCR. In parallel, were also evaluated samples obtained from five healthy dogs. Both treatment protocols promoted the remission of clinical signs as well as normalization of hematological and biochemical parameters and urinalysis values. Antileishmanial antibodies returned to non-significant titers in all dogs. Sick dogs showed a generalized upregulation of IFN-γ and downregulation of IL-2, IL-4, and TGF-β, while gene expression of IL-12, TNF-α, IL-5, and IL-10 varied between groups and according to evaluated tissue. A trend to the normalization of cytokine gene expression was induced by both miltefosine and meglumine antimoniate combined therapies. However, IFN-γ gene expression was still up-regulated in the three evaluated tissues. Furthermore, the effect of treatment in the gene expression of cytokines that were not significantly changed by infection, indicates that miltefosine and meglumine antimoniate combined therapy directly affects cytokine generation. Both combined therapies are effective in CanL treatment, leading to sustained pro-inflammatory immune environments that can compromise parasite survival and favor dogs' clinical cure. In the current study, anti-inflammatory and regulatory cytokines do not seem to play a prominent role in CanL or during clinical recovery.

The clinical manifestation of leishmaniases depends on parasite species, host genetic background, and immune response. Manifestations of human leishmaniases are highly variable, ranging from self-healing skin lesions to fatal visceral disease. The scope of standard model hosts is insufficient to mimic well the wide disease spectrum, which compels the introduction of new model animals for leishmaniasis research. In this article, we study the susceptibility of three Asian rodent species (Cricetulus griseus, Lagurus lagurus, and Phodopus sungorus) to Leishmania major and L. donovani. The external manifestation of the disease, distribution, as well as load of parasites and infectiousness to natural sand fly vectors, were compared with standard models, BALB/c mice and Mesocricetus auratus. No significant differences were found in disease outcomes in animals inoculated with sand fly- or culture-derived parasites. All Asian rodent species were highly susceptible to L. major. Phodopus sungorus showed the non-healing phenotype with the progressive growth of ulcerative lesions and massive parasite loads. Lagurus lagurus and C. griseus represented the healing phenotype, the latter with high infectiousness to vectors, mimicking best the character of natural reservoir hosts. Both, L. lagurus and C. griseus were also highly susceptible to L. donovani, having wider parasite distribution and higher parasite loads and infectiousness than standard model animals.

Hepatocytes constitute the majority of hepatic cells, and play a key role in controlling systemic innate immunity, via pattern-recognition receptors (PRRs) and by synthesizing complement and acute phase proteins. Leishmania infantum, a protozoan parasite that causes human and canine leishmaniasis, infects liver by establishing inside the Kupffer cells. The current study proposes the elucidation of the immune response generated by dog hepatocytes when exposed to L. infantum. Additionally, the impact of adding leishmanicidal compound, meglumine antimoniate (MgA), to parasite-exposed hepatocytes was also addressed. L. infantum presents a high tropism to hepatocytes, establishing strong membrane interactions. The possibility of L. infantum internalization by hepatocytes was raised, but not confirmed. Hepatocytes were able to recognize parasite presence, inducing PRRs [nucleotide oligomerization domain (NOD)1, NOD2 and Toll-like receptor (TLR)2] gene expression and generating a mix pro- and anti-inflammatory cytokine response. Reduction of cytochrome P 450s enzyme activity was also observed concomitant with the inflammatory response. Addition of MgA increased NOD2, TLR4 and interleukin 10 gene expression, indicating an immunomodulatory role for MgA. Hepatocytes seem to have a major role in coordinating liver's innate immune response against L. infantum infection, activating inflammatory mechanisms, but always balancing the inflammatory response in order to avoid cell damage.

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