Chromosomal segments may explain the antibody response cooperation for canine leishmaniasis pathogenesis.

Visceral leishmaniasis (VL) is marked by hyperactivation of a humoral response secreting high quantity of immunoglobulins (Igs) that are inaccessible to intracellular parasites. Here we investigated the contributions of the antibody response to the canine leishmaniasis pathogenesis. Using correlation and genome-wide association analysis, we investigated the relationship of anti-Leishmania infantum immunoglobulin classes levels with parasite burden, clinical response, renal/hepatic biochemical, and oxidative stress markers in dogs from endemic areas of VL. Immunoglobulin G (IgG) and IgA were positively correlated with parasite burden on lymph node and blood. Increased IgG, IgA and IgE levels were associated with severe canine leishmaniasis (CanL) whereas IgM was elevated in uninfected exposed dogs. Correlations of IgM, IgG and IgA with creatinine, urea, AST and ALT levels in the serum were suggested an involvement of those Igs with renal and hepatic changes. The correlogram of oxidative radicals and antioxidants revealed a likely relationship of IgM, IgG and IgA with oxidative stress and lipid peroxidation in the blood, suggested as mechanisms mediating tissue damage and CanL worsening. The gene mapping on chromosomal segments associated with the quantitative variation of immunoglobulin classes identified genetic signatures involved with reactive oxygen species generation, phagolysosome maturation and rupture, free iron availability, Th1/Th2 differenciation and, immunoglobulin clearance. The findings demonstrated the roles of the antibody response as resistance or susceptibility markers and mediators of CanL pathogenesis. In addition we pinpointed candidate genes as potential targets for the therapy against the damage caused by exacerbated antibody response and parasitism in VL.

Canine leishmaniasis: Genome-wide analysis and antibody response to Lutzomyia longipalpis saliva.

The anti-inflammatory properties of sand fly saliva favor the establishment of the Leishmania infantum infection. In contrast, an antibody response against Lutzomyia longipalpis saliva is often associated with a protective cell-mediated response against canine visceral leishmaniasis. Genetic studies may demonstrate to what extent the ability to secrete anti-saliva antibodies depends on genetic or environmental factors. However, the genetic basis of canine antibody response against sand fly saliva has not been assessed. The aim of this study was to identify chromosomal regions associated with the anti-Lu. longipalpis salivary IgG response in 189 dogs resident in endemic areas in order to provide information for prophylactic strategies. Dogs were classified into five groups based on serological and parasitological diagnosis and clinical evaluation. Anti-salivary gland homogenate (SGH) IgG levels were assessed by Enzyme-Linked Immunosorbent Assay (ELISA). Genomic DNA was isolated from blood samples and genotyped using a SNP chip with 173,662 single nucleotide polymorphism (SNP) markers. The following linear regression model was fitted: IgG level = mean + origin + sex + age + use of a repellent collar, and the residuals were assumed as pseudo-phenotypes for the association test between phenotypes and genotypes (GWA). A component of variance model that takes into account polygenic and sample structure effects (EMMAX) was employed for GWA. Phenotypic findings indicated that anti-SGH IgG levels remained higher in exposed and subclinically infected dogs than in severely diseased dogs even in regression model residuals. Five associated markers were identified on chromosomes 2, 20 and 31. The mapped genes included CD180 (RP105) and MITF related to the rapid activation of B lymphocytes and differentiation into antibody-secreting plasma cells. The findings pointed to chromosomal segments useful for functional confirmation studies and a search for adjuvant molecules of the anti-saliva response.

Skin mycoflora of Cebus primates kept in captivity and semicaptivity.

BACKGROUND: This study was performed to determine the fungal mycoflora in healthy tufted capuchins primates (Cebus sp) kept in captivity and semicaptivity to allow a more realistic interpretation on the basis of fungi isolated from their skin. Furthermore, we aimed at evaluating the potential risk of infection to humans by fungi perpetuated in the tegument of monkeys of this genus. METHODS: For the collection of skin material, the carpet method was used, followed by seeding in Sabouraud dextrose agar culture media, Mycosel agar, Dermatophyte Test Medium and Sabouraud agar supplemented with olive oil. RESULTS: Seventeen genera of fungi were detected, being three of them isolated only in the captivity animals (Acremonium - Cephalosporium, Phoma and Trichosporon). The genera of fungi with the higher frequencies were identified in the semicaptivity capuchins (Fusarium, Aspergillus and Penicillium). CONCLUSIONS: Many of the genera of fungi identified are potential pathogens for immune-compromised monkeys and humans.

Temporal IgG subclasses response in dogs following vaccination against Leishmania with Leishmune®.

Canine Visceral Leishmaniasis (CVL) is a widespread zoonotic disease with mandatory euthanasia of infected dogs determined by the Brazilian Ministry of Health. Development of vaccines against CVL may provide a prophylactic barrier, but transitory peak of antibody response detected by standard diagnostic techniques in vaccinated dogs may be interpreted as natural infection. Accordingly, the aim of the present study was to sequentially evaluate total and IgG subclasses response between naturally Leishmania-infected and dogs vaccinated with Leishmune(®). A total of 172 mongrel dogs were divided in four groups: Group 1 (G1) with 45 clinically healthy dogs, Group 2 (G2) and Group 3 (G3) with 45 dogs naturally infected by Leishmania sp. each, symptomatic and asymptomatic respectively, and G4 (G4) with 37 healthy dogs submitted to a complete protocol of a commercially available vaccine against CVL, monitored and evaluated in 5 different chronological moments (M0-M4) up to 180 days after M0. Total IgG, IgG1 and IgG2 were unable to differentiate between infected (G2 and G3) and vaccinated (G4) dogs, demonstrating that polyclonal commercial antibodies do not distinguish these groups apart. Total and IgG subclasses antibodies were not detected until 21 days of the second vaccination dose; however, seroconversion was observed on 21 days and sustained positivity up to 6 months after the vaccination start. A peak of antibodies response was observed on 90 days (M3), when results for total IgG, IgG1, IgG2, IgG3 and IgG4 where highly significant when compared to M0 (P<0.0001). Neither total IgG nor IgG1 effectively differentiated between infected (G2 and G3) and vaccinated (G4) dogs. In conclusion, despite dogs may test serologically negative immediately after vaccination against CVL with Leishmune(®), subsequent seroconversion, antibody peak and positivity up to six months may lead vaccinated dogs to be mistakenly identified as naturally infected dogs during this period.

Diminished CD4+/CD25+ T cell and increased IFN-gamma levels occur in dogs vaccinated with Leishmune in an endemic area for visceral leishmaniasis.

The Leishmune vaccine has been used in endemic areas to prevent canine visceral leishmaniasis in Brazil, but cytokine production induced by vaccination has rarely been investigated in dogs. This study aimed to evaluate the immune response of dogs vaccinated with Leishmune FML vaccine (Fort Dodge) against total antigen of Leishmania (Leishmania) chagasi (TAg) and FML. Twenty healthy dogs from Araçatuba, São Paulo, Brazil, an endemic leishmaniasis area, received three consecutive subcutaneous injection of Leishmune vaccine at 21-day intervals. PBMC were isolated before and 10 days after completing vaccination and lymphoproliferative response and antibody production against FML or total promastigote antigen were tested. Cytokines IFN-gamma, IL-4 and TNF-alpha were measured in culture supernatant and CD4+/CD25+ and CD8+/CD25+ T cell presence was determined. Analysis of the data indicated that the vaccine conferred humoral responses (100%) against both antigens and cellular immunity to FML (85%) and total antigen (80%), the supernatant of cultured cells stimulated with TAg and FML showed an increase in IFN-gamma (P<0.05), and the vaccine reduced CD4+/CD25+ T cell presence compared to that observed before vaccination. These responses may constitute part of the immune mechanism induced by Leishmune.