Leishsmania (Leishmania) amazonensis infection: Muscular involvement in BALB/c and C3H.HeN mice

Recent studies have provided some insights into Leishsmania (Leishmania) amazonensis muscular infection in dogs, although, muscular disease due to leishmaniasis has been poorly documented. The aim of our study was to evaluate involvement of Leishmania in muscular infection of two distinct mouse strains (BALB/c and C3H.He), with different genetic backgrounds. BALB/c mice, susceptible to Leishmania infection, showed, at the beginning of infection, a great number of infected macrophages among muscle fibers; however, in C3H.He resistant mice, muscle fibers were less damaged than in BALB/c mice, but some parasitized macrophages could be seen among them. A follow up of the infection showed an intense inflammatory infiltrate mainly composed of infected macrophages in BALB/c muscles and the presence of amastigotes within muscle fibers; while C3H.He mice exhibited a moderate inflammatory infiltrate among skeletal muscle fibers and an absence of amastigotes. Total destruction of muscles was observed in BALB/c mice in the late phase of infection (day 90) while C3H.He mice showed a process of muscle repair. We concluded that: (1) the muscles of BALB/c mice were more affected by leishmaniasis than those of C3/H.He mice; (2) Leishmania amastigotes are capable of infecting muscular fibers, as observed in BALB/c mice; (3) as inflammatory infiltrate is less intense in C3H.He mice these animals are capable of restoring muscular fibers.     

Infection-induced respiratory burst in BALB/c macrophages kills Leishmania guyanensis amastigotes through apoptosis: possible involvement in resistance to cutaneous leishmaniasis

The immune mechanisms that underlie resistance and susceptibility to leishmaniasis are not completely understood for all species of Leishmania. It is becoming clear that the immune response, the parasite elimination by the host and, as a result, the outcome of the disease depend both on the host and on the species of the infecting Leishmania. Here, we analyzed the outcome of the infection of BALB/c mice with L. guyanensis in vivo and in vitro. We showed that BALB/c mice, which are a prototype of susceptible host for most species of Leishmania, dying from these infections, develop insignificant or no cutaneous lesions and eliminate the parasite when infected with promastigotes of L. guyanensis. In vitro, we found that thioglycollate-elicited BALB/c peritoneal macrophages, which are unable to eliminate L. amazonensis without previous activation with cytokines or lipopolysaccharide, can kill L. guyanensis amastigotes. This is the first report showing that infection of peritoneal macrophages with stationary phase promastigotes efficiently triggers innate microbicidal mechanisms that are effective in eliminating the amastigotes, without exogenous activation. We demonstrated that L. guyanensis amastigotes die inside the macrophages through an apoptotic process that is independent of nitric oxide and is mediated by reactive oxygen intermediates generated in the host cell during infection. This innate killing mechanism of macrophages may account for the resistance of BALB/c mice to infection by L. guyanensis.     

Coadministration of the Three Antigenic Leishmania infantum Poly (A) Binding Proteins as a DNA Vaccine Induces Protection against Leishmania major Infection in BALB/c Mice

Background

Highly conserved intracellular proteins from Leishmania have been described as antigens in natural and experimental infected mammals. The present study aimed to evaluate the antigenicity and prophylactic properties of the Leishmania infantum Poly (A) binding proteins (LiPABPs).

Methodology/Principal Findings

Three different members of the LiPABP family have been described. Recombinant tools based on these proteins were constructed: recombinant proteins and DNA vaccines. The three recombinant proteins were employed for coating ELISA plates. Sera from human and canine patients of visceral leishmaniasis and human patients of mucosal leishmaniasis recognized the three LiPABPs. In addition, the protective efficacy of a DNA vaccine based on the combination of the three Leishmania PABPs has been tested in a model of progressive murine leishmaniasis: BALB/c mice infected with Leishmania major. The induction of a Th1-like response against the LiPABP family by genetic vaccination was able to down-regulate the IL-10 predominant responses elicited by parasite LiPABPs after infection in this murine model. This modulation resulted in a partial protection against L. major infection. LiPABP vaccinated mice showed a reduction on the pathology that was accompanied by a decrease in parasite burdens, in antibody titers against Leishmania antigens and in the IL-4 and IL-10 parasite-specific mediated responses in comparison to control mice groups immunized with saline or with the non-recombinant plasmid.

Conclusion/Significance

The results presented here demonstrate for the first time the prophylactic properties of a new family of Leishmania antigenic intracellular proteins, the LiPABPs. The redirection of the immune response elicited against the LiPABP family (from IL-10 towards IFN-γ mediated responses) by genetic vaccination was able to induce a partial protection against the development of the disease in a highly susceptible murine model of leishmaniasis.     

Evaluation of DNA/DNA and prime-boost vaccination using LPG3 against Leishmania major infection in susceptible BALB/c mice and its antigenic properties in human leishmaniasis

One of the main issues in vaccine development is implementation of new adjuvants to improve the antigen presentation and eliciting the protective immune response. Heat shock protein (HSP) molecules are known as natural adjuvants. They can stimulate the innate and adaptive immune response against infectious diseases and cancer. Lipophosphoglycan 3 (LPG3), the Leishmania homologous with GRP94 (glucose regulated protein 94), a member of HSP90 family, is involved in assembly of LPG as the most abundant macromolecule on the surface of Leishmania promastigotes. In the present study as a primary step, we tested LPG3 as a vaccine candidate in two regimens, DNA/DNA and prime-boost (DNA/Protein), against Leishmania major infection in BALB/c mice model. Our results showed that LPG3 and its fragment (rNT-LPG3) are highly immunogenic in BALB/c mice and can stimulate the production of both IgG1 and IgG2a. In prime-boost immunization strategy, the level of antibody response was higher compared with DNA/DNA immunization. The levels of IFN-γ in the supernatant of splenocytes from mice immunized with DNA/DNA and prime-boost regimens were significantly higher when compared to control groups. In fact, immunization with prime-boost vaccination has higher ratio of IFN-γ/IL-5, suggesting a shift towards a Th1 response.In addition, sera reactivity against LPG3 in visceral leishmaniasis (VL) patients was significantly higher in comparison with cutaneous leishmaniasis (CL) patients. Therefore, we recommend further investigations on the usage of LPG3 co-delivery with candidate antigens for vaccine development against leishmaniasis.     

Kinetics of antibody response in BALB/c and C57BL/6 mice bitten by Phlebotomus papatasi

Background

Phlebotomine sand flies are blood-sucking insects transmitting Leishmania parasites. In bitten hosts, sand fly saliva elicits specific immune response and the humoral immunity was shown to reflect the intensity of sand fly exposure. Thus, anti-saliva antibodies were suggested as the potential risk marker of Leishmania transmission. In this study, we examined the long-term kinetics and persistence of anti-Phlebotomus papatasi saliva antibody response in BALB/c and C57BL/6 mice. We also tested the reactivity of mice sera with P. papatasi salivary antigens and with the recombinant proteins.

Methodology/Principal Findings

Sera of BALB/c and C57BL/6 mice experimentally bitten by Phlebotomus papatasi were tested by ELISA for the presence of anti-saliva IgE, IgG and its subclasses. We detected a significant increase of specific IgG and IgG1 in both mice strains and IgG2b in BALB/c mice that positively correlated with the number of blood-fed P. papatasi females. Using western blot and mass spectrometry we identified the major P. papatasi antigens as Yellow-related proteins, D7-related proteins, antigen 5-related proteins and SP-15-like proteins. We therefore tested the reactivity of mice sera with four P. papatasi recombinant proteins coding for most of these potential antigens (PpSP44, PpSP42, PpSP30, and PpSP28). Each mouse serum reacted with at least one of the recombinant protein tested, although none of the recombinant proteins were recognized by all sera.

Conclusions

Our data confirmed the concept of using anti-sand fly saliva antibodies as a marker of sand fly exposure in Phlebotomus papatasi–mice model. As screening of specific antibodies is limited by the availability of salivary gland homogenate, utilization of recombinant proteins in such studies would be beneficial. Our present work demonstrates the feasibility of this implementation. A combination of recombinant salivary proteins is recommended for evaluation of intensity of sand fly exposure in endemic areas and for estimation of risk of Leishmania transmission.     

Vaccination with the Leishmania infantum ribosomal proteins induces protection in BALB/c mice against Leishmania chagasi and Leishmania amazonensis challenge

Leishmania chagasi and Leishmania amazonensis are the etiologic agents of different clinical forms of human leishmaniasis in South America. In an attempt to select candidate antigens for a vaccine protecting against different Leishmania species, the efficacy of vaccination using Leishmania ribosomal proteins and saponin as adjuvant was examined in BALB/c mice against challenge infection with both parasite species. Mice vaccinated with parasite ribosomal proteins purified from Leishmania infantum plus saponin showed a specific production of IFN-γ, IL-12 and GM-CSF after in vitro stimulation with L. infantum ribosomal proteins. Vaccinated mice showed a reduction in the liver and spleen parasite burdens after L. chagasi infection. After L. amazonensis challenge, vaccinated mice showed a decrease of the dermal pathology and a reduction in the parasite loads in the footpad and spleen. In both models, protection was correlated to an IL-12-dependent production of IFN-γ by CD4⁺ and CD8⁺ T cells that activate macrophages for the synthesis of NO. In the protected mice a decrease in the parasite-mediated IL-4 and IL-10 responses was also observed. In mice challenged with L. amazonensis, lower levels of anti-parasite-specific antibodies were detected. Thus, Leishmania ribosomal proteins plus saponin fits the requirements to compose a pan-Leishmania vaccine.     

Taenia crassiceps: Immunity to metacestodes in BALB/c and BDF1 mice

The kinetics of primary and secondary infections with Taenia crassiceps larvae and the effects of immune serum on T. crassiceps larvae were studied in BALB/c and BDF1 mice. In both strains of mice a substantial degree of resistance to reinfection comparable to that previously reported in C3H mice can be induced by subcutaneous injection of three larvae 3 weeks prior to intraperitoneal challenge infection. Both early immune damage in the absence of adherent host cells and encapsulation by host cells are involved in rejection of larvae by BALB/c and BDF1 mice, but in both of these strains early immune damage is less pronounced and the cellular encapsulation response considerably more prominent than in the C3H mice studied previously. This difference is also reflected in the effect of immune serum on T. crassiceps metacestodes in vitro: immune serum from BALB/c and BDF1 mice is less effective than immune serum taken from C3H mice at comparable times after challenge infection in mediating damage to T. crassiceps larvae in vitro in the absence of host cells. These results suggest that genetically determined differences in immune capability can alter the state of equilibrium existing among different immune effector mechanisms without producing measurable effects upon overall host resistance to reinfection.     

Influence of long-term treatment with pravastatin on the survival, evolution of cutaneous lesion and weight of animals infected by Leishmania amazonensis

The high toxicity of current drugs for treatment of leishmaniasis is a major hindrance for controlling the disease. Pravastatin is a well-known drug with anti-inflammatory and immunomodulatory properties that may modulate host defense mechanisms against Leishmania. We evaluated the influence of prolonged pravastatin treatment on the survival of Leishmania amazonensis-infected animals (BALB/c, C57BL6 mice and Syrian hamsters), including weekly measurement of cutaneous lesions (footpad thickness) and weight. Pravastatin improved survival of Leishmania-infected BALB/c mice but not of infected C57BL6 mice or hamsters. On the 50th week of follow-up, 71% of pravastatin-treated Leishmania-infected BALB/c mice were alive against 29% of control group (p < 0.01). Low footpad thickness was found on BALB/c pravastatin treated mice from the 14th week (p < 0.05), and 20th week onward for C57BL6 treated mice. Pravastatin treatment decreased weight loss in Leishmania-infected C57BL6 mice and Syrian hamsters, but not infected BALB/c mice. Our results points to beneficial effects of pravastatin on the evolution of the disease in the murine leishmaniasis model.     

C-Terminal Domain Deletion Enhances the Protective Activity of cpa/cpb Loaded Solid Lipid Nanoparticles against Leishmania major in BALB/c Mice

Background

We have demonstrated that vaccination with pDNA encoding cysteine proteinase Type II (CPA) and Type I (CPB) with its unusual C-terminal extension (CTE) can partially protect BALB/c mice against cutaneous leishmanial infection. Unfortunately, this protection is insufficient to completely control infection without booster injection. Furthermore, in developing vaccines for leishmaniasis, it is necessary to consider a proper adjuvant and/or delivery system to promote an antigen specific immune response. Solid lipid nanoparticles have found their way in drug delivery system development against intracellular infections and cancer, but not Leishmania DNA vaccination. Therefore, undefined effect of cationic solid lipid nanoparticles (cSLN) as an adjuvant in enhancing the immune response toward leishmanial antigens led us to refocus our vaccine development projects.

Methodology/Principal Findings

Three pDNAs encoding L. major cysteine proteinase type I and II (with or without CTE) were formulated by cSLN. BALB/c mice were immunized twice by 3-week interval, with cSLN-pcDNA-cpa/b, pcDNA-cpa/b, cSLN-pcDNA-cpa/b^-CTE, pcDNA-cpa/b^-CTE, cSLN, cSLN-pcDNA and PBS. Mice vaccinated with cSLN-pcDNA-cpa/b^-CTE showed significantly higher levels of parasite inhibition related to protection with specific Th1 immune response development, compared to other groups. Parasite inhibition was determined by different techniques currently available in exploration vacciation efficacy, i.e., flowcytometry on footpad and lymph node, footpad caliper based measurements and imaging as well as lymph node microtitration assay. Among these techniques, lymph node flowcytometry was found to be the most rapid, sensitive and easily reproducible method for discrimination between the efficacy of vaccination strategies.

Conclusions/Significance

This report demonstrates cSLN''s ability to boost immune response magnitude of cpa/cpb^-CTE cocktail vaccination against leishmaniasis so that the average parasite inhibition percent could be increased significantly. Hence, cSLNs can be considered as suitable adjuvant and/or delivery systems for designing third generation cocktail vaccines.     

Induction of autophagy correlates with increased parasite load of Leishmania amazonensis in BALB/c but not C57BL/6 macrophages

We investigated the role of autophagy in infection of macrophages by Leishmania amazonensis. Induction of autophagy by IFN-γ or starvation increased intracellular parasite load and the percentages of infected macrophages from BALB/c but not from C57BL/6 mice. In contrast, starvation did not affect the replication of either Leishmania major or Trypanosoma cruzi in BALB/c macrophages. In BALB/c macrophages, starvation resulted in increased monodansylcadaverine staining and in the appearance of double-membrane and myelin-like vesicles characteristic of autophagosomes. Increased parasite load was associated with a reduction in NO levels and was attenuated by wortmannin, an inhibitor of autophagy. In infected macrophages from BALB/c, but not from C57BL/6 mice, starvation increased the number of lipid bodies and the amounts of PGE₂ produced. Exogenous PGE₂ increased parasite load in macrophages from BALB/c, but not C57BL/6 mice. The cyclooxygenase inhibitor indomethacin prevented the increase of parasite load in starved BALB/c macrophages, and actually induced parasite killing. These results suggest that autophagy regulates the outcome of L. amazonensis infection in macrophages in a host strain specific manner.     

Down-Regulation of TLR and JAK/STAT Pathway Genes Is Associated with Diffuse Cutaneous Leishmaniasis: A Gene Expression Analysis in NK Cells from Patients Infected with Leishmania mexicana

An important NK-cell inhibition with reduced TNF-α, IFN-γ and TLR2 expression had previously been identified in patients with diffuse cutaneous leishmaniasis (DCL) infected with Leishmania mexicana. In an attempt to pinpoint alterations in the signaling pathways responsible for the NK-cell dysfunction in patients with DCL, this study aimed at identifying differences in the NK-cell response towards Leishmania mexicana lipophosphoglycan (LPG) between patients with localized and diffuse cutaneous leishmaniasis through gene expression profiling. Our results indicate that important genes involved in the innate immune response to Leishmania are down-regulated in NK cells from DCL patients, particularly TLR and JAK/STAT signaling pathways. This down-regulation showed to be independent of LPG stimulation. The study sheds new light for understanding the mechanisms that undermine the correct effector functions of NK cells in patients with diffuse cutaneous leishmaniasis contributing to a better understanding of the pathobiology of leishmaniasis.     

T-Cell Populations and Cytokine Expression Are Impaired in Thymus and Spleen of Protein Malnourished BALB/c Mice Infected with Leishmania infantum

Visceral leishmaniasis (VL) is a parasitic infectious disease that causes significant morbidity and mortality in the tropical and subtropical regions of the world. Although infections with visceralizing Leishmania may be asymptomatic, factors such as undernutrition increase the likelihood of progressing to clinical disease. Protein malnutrition, the most deleterious cause of malnutrition in developing countries, has been considered as a primary risk factor for the development of clinical VL. However, data regarding the immunological basis of this association are scarce. With the aim to analyze the effects of protein malnutrition on Leishmania infantum infection, we used BALB/c mice subjected to control or low protein isocaloric diets. Each animal group was divided into two subgroups and one was infected with L. infantum resulting in four study groups: animals fed 14% protein diet (CP), animals fed 4% protein diet (LP), animals fed 14% protein diet and infected (CPi), and animals fed 4% protein diet and infected (LPi).The susceptibility to L. infantum infection and immune responses were assessed in terms of body and lymphoid organ weight, parasite load, lymphocyte subpopulations, and cytokine expression. LPi mice had a significant reduction of body and lymphoid organ weight and exhibited a severe decrease of lymphoid follicles in the spleen. Moreover, LPi animals showed a significant decrease in CD4⁺CD8⁺ T cells in the thymus, whereas there was an increase of CD4⁺ and CD8⁺ T cells percentages in the spleen. Notably, the cytokine mRNA levels in the thymus and spleen of protein malnourished-infected animals were altered compared to the CP mice. Protein malnutrition results in a drastic dysregulation of T cells and cytokine expression in the thymus and spleen of L. infantum-infected BALB/c mice, which may lead to defective regulation of the thymocyte population and an impaired splenic immune response, accelerating the events of a normal course of infection.     

Anti-leishmanial Effects of Trinitroglycerin in BALB/C Mice Infected with Leishmania major via Nitric Oxide Pathway

This study investigated whether trinitroglycerine (TNG) as nitric oxide (NO) releasing agent had anti-leishmanial effects and mediated pathology in BALB/c mice infected with Leishmania major. Cutaneous leishmaniasis (CL), a zoonotic infection caused by leishmania protozoa is still one of the health problems in the world and in Iran. NO is involved in host immune responses against intracellular L. major, and leishmania killing by macrophages is mediated by this substance. Moreover, application of CL treatment with NO-donors has been recently indicated. In our study, TNG was used for its ability to increase NO and to modify CL infection in mice, in order to evaluate NO effects on lesion size and formation, parasite proliferation inside macrophages, amastigote visceralization in target organs, and NO induction in plasma and organ suspensions. Data obtained in this study indicated that TNG increased plasma and liver-NO, reduced lesion sizes, removed amastigotes from lesions, livers, spleens, and lymph nodes, declined proliferation of amastigotes, hepatomegaly, and increased survival rate. However, TNG reduced spleen-NO and had no significant effects on spelenomegaly. The results show that TNG therapy reduced leishmaniasis and pathology in association with raised NO levels. TNG had some antiparasitic activity by reduction of positive smears from lesions, livers, spleens, and lymph nodes, which could emphasize the role of TNG to inhibit visceralization of L. major in target organs.     

Sphingolipid Degradation in Leishmania (Leishmania) amazonensis

Background

Human leishmaniasis is caused by more than 20 Leishmania species and has a wide range of symptoms. Our recent studies have demonstrated the essential role of sphingolipid degradation in the virulence of Leishmania (Leishmania) major, a species responsible for localized cutaneous leishmaniasis in the Old World. In this study, we investigated the function of sphingolipid degradation in Leishmania (Leishmania) amazonensis, an etiological agent of localized and diffuse cutaneous leishmaniasis in South America.

Methodology/Principal Findings

First, we identified the enzyme LaISCL which is responsible for sphingolipid degradation in L. amazonensis. Primarily localized in the mitochondrion, LaISCL shows increased expression as promastigotes progress from replicative log phase to non-replicative stationary phase. To study its function, null mutants of LaISCL (Laiscl⁻) were generated by targeted gene deletion and complemented through episomal gene add-back. In culture, loss of LaISCL leads to hypersensitivity to acidic pH and poor survival in murine macrophages. In animals, Laiscl⁻ mutants exhibit severely attenuated virulence towards C57BL6 mice but are fully infective towards BALB/c mice. This is drastically different from wild type L. amazonensis which cause severe pathology in both BALB/c and C57BL 6 mice.

Conclusions/Significance

A single enzyme LaISCL is responsible for the turnover of sphingolipids in L. amazonensis. LaISCL exhibits similar expression profile and biochemical property as its ortholog in L. major. Deletion of LaISCL reduces the virulence of L. amazonensis and the outcome of Laiscl⁻-infection is highly dependent on the host''s genetic background. Therefore, compared to L. major, the role of sphingolipid degradation in virulence is substantially different in L. amazonensis. Future studies may reveal whether sphingolipid degradation is required for L. amazonensis to cause diffuse cutaneous infections in humans.     

Differential Gene Expression and Infection Profiles of Cutaneous and Mucosal Leishmania braziliensis Isolates from the Same Patient

BackgroundLeishmaniasis is a complex disease in which clinical outcome depends on factors such as parasite species, host genetics and immunity and vector species. In Brazil, Leishmania (Viannia) braziliensis is a major etiological agent of cutaneous (CL) and mucosal leishmaniasis (MCL), a disfiguring form of the disease, which occurs in ~10% of L. braziliensis-infected patients. Thus, clinical isolates from patients with CL and MCL may be a relevant source of information to uncover parasite factors contributing to pathogenesis. In this study, we investigated two pairs of L. (V.) braziliensis isolates from mucosal (LbrM) and cutaneous (LbrC) sites of the same patient to identify factors distinguishing parasites that migrate from those that remain at the primary site of infection.Conclusions/SignificanceDespite sharing high similarity at the genome structure and ploidy levels, the parasites exhibited divergent expressed genomes. The proteome and metabolome results indicated differential profiles between the cutaneous and mucosal isolates, primarily related to inflammation and chemotaxis. BALB/c infection revealed that the cutaneous isolates were more virulent than the mucosal parasites. Furthermore, our data suggest that the LbrPGF2S protein is a candidate to contribute to parasite virulence profiles in the mammalian host.     

Protective immunity against Leishmania major induced by Leishmania tropica infection of BALB/c mice

Leishmania (L.) tropica is a causative agent of human cutaneous and viscerotropic leishmaniasis. Immune response to L. tropica in humans and experimental animals are not well understood. We previously established that L. tropica infection induces partial protective immunity against subsequent challenge infection with Leishmania major in BALB/c mice. Aim of the present study was to study immunologic mechanisms of protective immunity induced by L. tropica infection, as a live parasite vaccine, in BALB/c mouse model. Mice were infected by L. tropica, and after establishment of the infection, they were challenged by L. major. Our findings shows that L. tropica infection resulted in protection against L. major challenge in BALB/c mice and this protective immunity is associated with: (1) a DTH response, (2) higher IFN-γ and lower IL-10 response at one week post-challenge, (3) lower percentage of CD4⁺ lymphocyte at one month post-challenge, and (4) the source of IFN-γ and IL-10 were mainly CD4⁻ lymphocyte up to one month post-challenge suggesting that CD4⁻ lymphocytes may be responsible for protection induced by L. tropica infection in the studied intervals.     

Histopathology,humoral and cellular immune response in the murine model of Leishmania (Viannia) shawi

Leishmania (Viannia) shawi was recently characterized and few studies concerning modifications in cellular and humoral immune responses in experimental leishmaniasis have been conducted. In this work, immunopathological changes induced by L. shawi in chronically infected BALB/c mice were investigated. Infected BALB/c mice developed increased lesion size associated with strong inflammatory infiltrate diffusely distributed in the dermis, with highly infected macrophages. The humoral immune response was predominantly directed toward the IgG1 isotype. The functional activity of CD4⁺ and CD8⁺ T cells showed significantly increased TNF-α mRNA levels associated with reduced IFN-γ expression by CD4⁺ T cells and the double negative (dn) CD4CD8 cell subset. High IL-4 levels expressed by CD8⁺ T cells and dnCD4CD8 and TGF-β by CD4⁺ and CD8⁺ T cells were detected, while IL-10 was highly expressed by all three cell subpopulations. Taken together, these results show an evident imbalance between TNF-α and IFN-γ that is unfavorable to amastigote replication control. Furthermore, L. shawi seems to regulate different cell populations to express deactivating cytokines to avoid its own destruction. This study indicates BALB/c mice as a potentially good experimental model for further studies on American cutaneous leishmaniosis caused by L. shawi.     

Characterization of a Leishmania stage‐specific mitochondrial membrane protein that enhances the activity of cytochrome c oxidase and its role in virulence

Leishmaniasis is caused by the dimorphic protozoan parasite Leishmania. Differentiation of the insect form, promastigotes, to the vertebrate form, amastigotes, and survival inside the vertebrate host accompanies a drastic metabolic shift. We describe a gene first identified in amastigotes that is essential for survival inside the host. Gene expression analysis identified a 27 kDa protein‐encoding gene (Ldp27) that was more abundantly expressed in amastigotes and metacyclic promastigotes than in procyclic promastigotes. Immunofluorescence and biochemical analysis revealed that Ldp27 is a mitochondrial membrane protein. Co‐immunoprecipitation using antibodies to the cytochrome c oxidase (COX) complex, present in the inner mitochondrial membrane, placed the p27 protein in the COX complex. Ldp27 gene‐deleted parasites (Ldp27^?/?) showed significantly less COX activity and ATP synthesis than wild type in intracellular amastigotes. Moreover, the Ldp27^?/? parasites were less virulent both in human macrophages and in BALB/c mice. These results demonstrate that Ldp27 is an important component of an active COX complex enhancing oxidative phosphorylation specifically in infectious metacyclics and amastigotes and promoting parasite survival in the host. Thus, Ldp27 can be explored as a potential drug target and parasites devoid of the p27 gene could be considered as a live attenuated vaccine candidate against visceral leishmaniasis.