Leishmania (L.) tropica is a causative agent of cutaneous leishmaniasis, and occasionally of visceral or viscerotropic leishmaniasis in humans. Murine models of Leishmania infection have been proven to be useful for elucidation of mechanisms for pathogenesis and immunity in leishmaniasis. The aim of this study was to establish a murine model for human viscerotropic leishmaniasis, and the growth pattern of L. tropica was studied in different tissues of BALB/c mice in order to find out whether the parasite visceralizes in this murine model. L. major was used as a control as this species is known to cause a progressive infection in BALB/c mice. L. tropica or L. major was injected into the footpad of mice, and thickness of footpad, parasite loads in different tissues, and the weight of the spleen and lymph node were determined at different intervals. Results showed that L. tropica visceralizes to the spleen and grows there while its growth is controlled in footpad tissues. Dissemination of L. tropica to visceral organs in BALB/c mice was similar to the growth patterns of this parasite in human viscerotropic leishmaniasis. The BALB/c model of L. tropica infection may be considered as a good experimental model for human diseases.
Animals ; *Disease Models, Animal ; Female ; Foot/parasitology ; Humans ; Leishmania major/growth & development ; Leishmania tropica/*growth & development ; Leishmaniasis/*parasitology ; Lymph Nodes/parasitology/pathology ; Mice ; Mice, Inbred BALB C ; Organ Size ; Spleen/parasitology/pathology

Objective: To study the role of antibodies in protection against Leishmania tropica (L. tropica) infection in the experimental model of BALB/c mice.Methods: BALB/c mice were vaccinated against L. tropica by soluble Leishmania antigen or recombinant L. tropica stress-inducible protein-1 (LtSTI1) of L. tropica, and against Leishmania major (L. major) by soluble Leishmania antigen. Monophosphoryl lipid A was used as an adjuvant. The L. tropica- or L. major-vaccinated mice were challenged by L. tropica or L. major, respectively. The levels of anti-Leishmania antibodies (IgG1 and IgG2a) were determined after vaccination and after challenge. Results: All vaccinated groups caused a higher antibody response in comparison with the control group. The L. major-vaccinated group showed lower IgG1 response than the control group after the challenge. Conversely, in L. tropica-vaccinated mice, the levels of antibodies were higher than the control group. Moreover, the group receiving rLtSTI1 and monophosphoryl lipid A showed higher levels of antibodies than those of the rLtSTI1 group. In vaccinated mice, antibody responses against L. tropica remained high until 16 weeks after the challenge. Conclusions: The higher levels of post-challenge antibodies are associated with protective vaccination against L. tropica infection of BALB/c mice. Our findings provide new insight into the association of antibody with vaccine-induced protective immunity against L. tropica infection. More studies are needed to clarify the role of antibody in protection against L. tropica.

Objective: To compare limiting dilution assay and real-time PCR methods in Leishmania tropica parasite load measurement in vaccinated mice.Methods: BALB/c mice were vaccinated by Leishmania tropica soluble Leishmania antigen or recombinant Leishmania tropica stress-inducible protein-1 with/without adjuvant. After three vaccinations, mice were challenged by Leishmania tropica promastigotes. Two months after challenge, the draining lymph nodes of mice footpad were removed and parasite load was assayed by limiting dilution assay and real-time PCR methods. Limiting dilution assay was done by diluting tissue samples to extinction in a biphasic medium. For real-time PCR, DNA of the lymph nodes was extracted, equal dilutions of each sample were prepared and hot-start real-time PCR was done using appropriate primers. The data of the two methods were compared by appropriate statistical methods. Results: Both methods were able to measure different levels of parasite load in vaccinated/unvaccinated mice. In addition, wherever parasite load of a group was estimated high (or low) by one method, the estimated parasite load by another method was the same, although statistically significant differences were found between some groups. Conclusions: Both methods lead to approximately similar results in terms of differentiating parasite load of the experimental groups. However, due to the lower errors and faster process, the real-time PCR method is preferred.

Leishmania tropica and L. major are etiologic agents of human cutaneous leishmaniasis. Delayed type hypersensitivity (DTH) is an immunologic response that has been frequently used as a correlate for protection against or sensitization to leishmania antigen. In BALB/c mice, L. tropica infection results in non-ulcerating disease, whereas L. major infection results in destructive lesions. In order to clarify the immunologic mechanisms of these 2 different outcomes, we compared the ability of these 2 leishmania species in induction of DTH response in this murine model. BALB/c mice were infected with L. major or L. tropica, and disease evolution and DTH responses were determined. The results show that the primary L. major infection can exacerbate the secondary L. major infection and is associated with DTH response. Higher doses of the primary L. major infection result in more disease exacerbation of the secondary L. major infection as well as higher DTH response. L. tropica infection induces lower DTH responses than L. major. We have previously reported that the primary L. tropica infection induces partial protection against the secondary L. major infection in BALB/c mice. Induction of lower DTH response by L. tropica suggests that the protection induced against L. major by prior L. tropica infection may be due to suppression of DTH response.
Animals ; Disease Models, Animal ; Ear/pathology ; Female ; Foot/pathology ; *Hypersensitivity, Delayed ; Leishmania major/*immunology ; Leishmania tropica/*immunology ; Leishmaniasis, Cutaneous/*immunology/*parasitology/pathology ; Mice ; Mice, Inbred BALB C

Objective: To study the role of antibodies in protection against Leishmania tropica (L. tropica) infection in the experimental model of BALB/c mice. Methods: BALB/c mice were vaccinated against L. tropica by soluble Leishmania antigen or recombinant L. tropica stress-inducible protein-1 (LtSTI1) of L. tropica, and against Leishmania major (L. major) by soluble Leishmania antigen. Monophosphoryl lipid A was used as an adjuvant. The L. tropica- or L. major-vaccinated mice were challenged by L. tropica or L. major, respectively. The levels of anti-Leishmania antibodies (IgG1 and IgG2a) were determined after vaccination and after challenge. Results: All vaccinated groups caused a higher antibody response in comparison with the control group. The L. major-vaccinated group showed lower IgG1 response than the control group after the challenge. Conversely, in L. tropica-vaccinated mice, the levels of antibodies were higher than the control group. Moreover, the group receiving rLtSTI1 and monophosphoryl lipid A showed higher levels of antibodies than those of the rLtSTI1 group. In vaccinated mice, antibody responses against L. tropica remained high until 16 weeks after the challenge. Conclusions: The higher levels of post-challenge antibodies are associated with protective vaccination against L. tropica infection of BALB/c mice. Our findings provide new insight into the association of antibody with vaccine-induced protective immunity against L. tropica infection. More studies are needed to clarify the role of antibody in protection against L. tropica.

Leishmania tropica is one of the causative agents of leishmaniasis in humans. Routes of infection have been reported to be an important variable for some species of Leishmania parasites. The role of this variable is not clear for L. tropica infection. The aim of this study was to explore the effects of route of L. tropica infection on the disease outcome and immunologic parameters in BALB/c mice. Two routes were used; subcutaneous in the footpad and intradermal in the ear. Mice were challenged by Leishmani major, after establishment of the L. tropica infection, to evaluate the level of protective immunity. Immune responses were assayed at week 1 and week 4 after challenge. The subcutaneous route in the footpad in comparison to the intradermal route in the ear induced significantly more protective immunity against L. major challenge, including higher delayed-type hypersensitivity responses, more rapid lesion resolution, lower parasite loads, and lower levels of IL-10. Our data showed that the route of infection in BALB/c model of L. tropica infection is an important variable and should be considered in developing an appropriate experimental model for L. tropica infections.
Animals ; Disease Models, Animal ; Female ; Leishmania major/*immunology ; Leishmania tropica/*immunology/*pathogenicity ; Leishmaniasis/*immunology/parasitology/*pathology ; Mice ; Mice, Inbred BALB C ; Treatment Outcome

OBJECTIVES: Despite all the efforts and increased knowledge of rabies, the exact mechanisms of infection and mortality from the rabies virus are not well understood. To understand the mechanisms underlying the pathogenicity of rabies virus infection, it is crucial to study the tissue that the rabies virus naturally infects in humans. METHODS: Cerebellum brain tissue from 9 human post mortem cases from Iran, who had been infected with rabies virus, were examined histopathologically and immunohistochemically to evaluate the innate immune responses against the rabies virus. RESULTS: Histopathological examination revealed inflammation of the infected cerebellum and immunohistochemical analyses showed an increased immunoreactivity of heat shock protein 70, interleukin-6, interleukin-1, tumor necrosis factor-alpha, caspase-3, caspase-9, toll-like receptor3 and toll-like receptor4 in the infected brain tissue. CONCLUSION: These results indicated the involvement of innate immunity in rabies infected human brain tissue, which may aggravate the progression of this deadly disease.
Brain ; Caspase 3 ; Caspase 9 ; Central Nervous System ; Cerebellum ; HSP70 Heat-Shock Proteins ; Humans ; Immunity, Innate ; Immunohistochemistry ; Inflammation ; Interleukin-1 ; Interleukin-6 ; Iran ; Mortality ; Pathology ; Rabies virus ; Rabies ; Tumor Necrosis Factor-alpha ; Virulence

Objective: To compare limiting dilution assay and real-time PCR methods in Leishmania tropica parasite load measurement in vaccinated mice. Methods: BALB/c mice were vaccinated by Leishmania tropica soluble Leishmania antigen or recombinant Leishmania tropica stress-inducibleprotein-1 with/without adjuvant. After three vaccinations, mice were challenged by Leishmania tropica promastigotes. Two months after challenge, the draining lymph nodes of mice footpad were removed and parasite load was assayed by limiting dilution assay and real-time PCR methods. Limiting dilution assay was done by diluting tissue samples to extinction in a biphasic medium. For real-time PCR, DNA of the lymph nodes was extracted, equal dilutions of each sample were prepared and hot-start real-time PCR was done using appropriate primers. The data of the two methods were compared by appropriate statistical methods. Results: Both methods were able to measure different levels of parasite load in vaccinated/unvaccinated mice. In addition, wherever parasite load of a group was estimated high (or low) by one method, the estimated parasite load by another method was the same, although statistically significant differences were found between some groups. Conclusions: Both methods lead to approximately similar results in terms of differentiating parasite load of the experimental groups. However, due to the lower errors and faster process, the real-time PCR method is preferred.