Recombinant forms of Leishmania amazonensis excreted/secreted promastigote surface antigen (PSA) induce protective immune responses in dogs

International audiencePreventive vaccination is a highly promising strategy for interrupting leishmaniasis transmission that can, additionally, contribute to elimination. A vaccine formulation based on naturally excreted secreted (ES) antigens was prepared from L. infantum promastigote culture supernatant. This vaccine achieved successful results in Phase III trials and was licensed and marketed as CaniLeish. We recently showed that newly identified ES promastigote surface antigen (PSA), from both viable promastigotes and axenically-grown amastigotes, represented the major constituent and the highly immunogenic antigen of L. infantum and L. amazonensis ES products. We report here that three immunizations with either the recombi-nant ES LaPSA-38S (rPSA) or its carboxy terminal part LaPSA-12S (Cter-rPSA), combined with QA-21 as adjuvant, confer high levels of protection in naive L. infantum-infected Beagle dogs, as checked by bone marrow parasite absence in respectively 78.8% and 80% of vaccinated dogs at 6 months post-challenge. The parasite burden in infected vaccinated dogs was significantly reduced compared to placebo group, as measured by q-PCR. Moreover, our results reveal humoral and cellular immune response clear-cut differences between vaccinated and control dogs. An early increase in specific IgG2 antibodies was observed in rPSA/QA-21-and Cter-rPSA/QA-21-immunized dogs only. They were found functionally active in vitro and were highly correlated with vaccine protection. In vaccinated protected dogs, IFN-γ and NO productions, as well as anti-leishmanial macrophage activity, were increased. These data strongly suggest that ES PSA or its carboxy-terminal part, in recom-binant forms, induce protection in a canine model of zoonotic visceral leishmaniasis by inducing a Th1-dominant immune response and an appropriate specific antibody response. These data suggest that they could be considered as important active components in vaccine candidates

In Vitro Evaluation of a Soluble Leishmania Promastigote Surface Antigen as a Potential Vaccine Candidate against Human Leishmaniasis

International audiencePSA (Promastigote Surface Antigen) belongs to a family of membrane-bound and secreted proteins present in severalLeishmania (L.) species. PSA is recognized by human Th1 cells and provides a high degree of protection in vaccinated mice.We evaluated humoral and cellular immune responses induced by a L. amazonensis PSA protein (LaPSA-38S) produced in aL. tarentolae expression system. This was done in individuals cured of cutaneous leishmaniasis due to L. major (CCLm) or L.braziliensis (CCLb) or visceral leishmaniasis due to L. donovani (CVLd) and in healthy individuals. Healthy individuals weresubdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantuminfection) or non immune/naive individuals (HLR: Healthy Low Responders), depending on whether they produce high orlow levels of IFN-c in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detectedin sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-c, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-a in more. No significant cytokine response wasobserved in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increasein CD4+ T cells producing IFN-c after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between thepercentage of IFN-c-producing CD4+ T cells and the released IFN-c. We showed that the LaPSA-38S protein was able toinduce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infectionindicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacityof Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infectio

Serological detection of antibodies against Paracoccidioides brasiliensis in dogs with leishmaniasis

The aim of this study was to detect antibodies against Paracoccidioides brasiliensis in dogs seropositive and seronegative for leishmaniasis. Sera from 836 dogs (449 positive and 387 negative to leishmaniasis) were analysed by ELISA and the immunodiffusion test using gp43 and exoantigen, respectively. The analysis of the 836 serum samples by ELISA and the immunodiffusion test showed a positivity of 67.8 % and 7.3%, respectively, for P. brasiliensis infection. The dogs positive to leishmaniasis showed a higher reactivity to gp43 (79.9%) and exoantigen (12.7%) than the negative ones (54.0% and 1.0%, respectively). The higher reactivity to P. brasiliensis antigens may be due to cross-reactivity or a co-infection of dogs by Leishmania and P. brasiliensis. The lower correlation (0.187) observed between reactivity to gp43 and Leishmania antigen reinforces the latter hypothesis

Resistance of Leishmania (Viannia) braziliensis to nitric oxide: correlation with antimony therapy and TNF-α production

<p>Abstract</p> <p>Background</p> <p>Nitric oxide (NO) produced in macrophages plays a pivotal role as a leishmanicidal agent. A previous study has demonstrated that 20% of the <it>L. (V.) braziliensis </it>isolated from initial cutaneous lesions of patients from the endemic area of Corte de Pedra, Bahia, Brazil, were NO resistant. Additionally, 5 to 11% of the patients did not respond to three or more antimony treatments" (refractory patients). The aim of this study is to investigate if there is an association between the resistance of <it>L. (V.) braziliensis </it>to NO and nonresponsiveness to antimony therapy and cytokine production.</p> <p>Methods</p> <p>We evaluated the <it>in vitro </it>toxicity of NO against the promastigotes stages of <it>L. (V.) braziliensis </it>isolated from responsive and refractory patients, and the infectivity of the amastigote forms of these isolates against human macrophages. The supernatants from <it>Leishmania </it>infected macrophage were used to measure TNF-α and IL-10 levels.</p> <p>Results</p> <p>Using NaNO₂(pH 5.0) as the NO source, <it>L. (V.) braziliensis </it>isolated from refractory patients were more NO resistant (IC50 = 5.8 ± 4.8) than <it>L. (V.) braziliensis </it>isolated from responsive patients (IC50 = 2.0 ± 1.4). Four isolates were selected to infect human macrophages: NO-susceptible and NO-resistant <it>L. (V.) braziliensis </it>isolated from responsive and refractory patients. NO-resistant <it>L. (V.) braziliensis </it>isolated from refractory patients infected more macrophages stimulated with LPS and IFN-γ at 120 hours than NO-susceptible <it>L. (V.) braziliensis </it>isolated from refractory patients. Also, lower levels of TNF-α were detected in supernatants of macrophages infected with NO-resistant <it>L. (V.) braziliensis </it>as compared to macrophages infected with NO-susceptible <it>L. (V.) braziliensis </it>(p < 0.05 at 2, 24 and 120 hours), while no differences were detected in IL-10 levels.</p> <p>Conclusion</p> <p>These data suggest that NO resistance could be related to the nonresponsiveness to antimony therapy seen in American Tegumentary Leishmaniasis.</p

Proteins of Leishmania (Viannia) shawi confer protection associated with Th1 immune response and memory generation

<p>Abstract</p> <p>Background</p> <p><it>Leishmania (Viannia) shawi </it>parasite was first characterized in 1989. Recently the protective effects of soluble leishmanial antigen (SLA) from <it>L. (V.) shawi </it>promastigotes were demonstrated using BALB/c mice, the susceptibility model for this parasite. In order to identify protective fractions, SLA was fractionated by reverse phase HPLC and five antigenic fractions were obtained.</p> <p>Methods</p> <p>F1 fraction was purified from L. (V.) shawi parasite extract by reverse phase HPLC. BALB/c mice were immunized once a week for two consecutive weeks by subcutaneous routes in the rump, using 25 μg of F1. After 1 and 16 weeks of last immunization, groups were challenged in the footpad with L. (V.) shawi promastigotes. After 2 months, those same mice were sacrificed and parasite burden, cellular and humoral immune responses were evaluated.</p> <p>Results</p> <p>The F1 fraction induced a high degree of protection associated with an increase in IFN-γ, a decrease in IL-4, increased cell proliferation and activation of CD8⁺T lymphocytes. Long-term protection was acquired in F1-immunized mice, associated with increased CD4⁺central memory T lymphocytes and activation of both CD4⁺and CD8⁺T cells. In addition, F1-immunized groups showed an increase in IgG2a levels.</p> <p>Conclusions</p> <p>The inductor capability of antigens to generate memory lymphocytes that can proliferate and secrete beneficial cytokines upon infection could be an important factor in the development of vaccine candidates against American Tegumentary Leishmaniasis.</p

LeishVet guidelines for the practical management of canine leishmaniosis

The LeishVet group has formed recommendations designed primarily to help the veterinary clinician in the management of canine leishmaniosis. The complexity of this zoonotic infection and the wide range of its clinical manifestations, from inapparent infection to severe disease, make the management of canine leishmaniosis challenging. The recommendations were constructed by combining a comprehensive review of evidence-based studies, extensive clinical experience and critical consensus opinion discussions. The guidelines presented here in a short version with graphical topic displays suggest standardized and rational approaches to the diagnosis, treatment, follow-up, control and prevention of canine leishmaniosis. A staging system that divides the disease into four stages is aimed at assisting the clinician in determining the appropriate therapy, forecasting prognosis, and implementing follow-up steps required for the management of the leishmaniosis patient

One Health: The global challenge of epidemic and endemic leishmaniasis

'One Health' proposes the unification of medical and veterinary sciences with the establishment of collaborative ventures in clinical care, surveillance and control of cross-species disease, education, and research into disease pathogenesis, diagnosis, therapy and vaccination. The concept encompasses the human population, domestic animals and wildlife, and the impact that environmental changes ('environmental health') such as global warming will have on these populations. Visceral leishmaniasis is a perfect example of a small companion animal disease for which prevention and control might abolish or decrease the suffering of canine and human patients, and which aligns well with the One Health approach. In this review we discuss how surveillance for leishmaniases is undertaken globally through the control of anthroponootic visceral leishmaniasis (AVL) and zoonotic visceral leishmaniasis (ZVL). The ZVL epidemic has been managed to date by the culling of infected dogs, treatment of human cases and control of the sandfly vector by insecticidal treatment of human homes and the canine reservoir. Recently, preventive vaccination of dogs in Brazil has led to reduction in the incidence of the canine and human disease. Vaccination permits greater dog owner compliance with control measures than a culling programme. Another advance in disease control in Africa is provided by a surveillance programme that combines remote satellite sensing, ecological modelling, vector surveillance and geo-spatial mapping of the distribution of vectors and of the animal-to-animal or animal-to-human pathogen transmission. This coordinated programme generates advisory notices and alerts on emerging infectious disease outbreaks that may impede or avoid the spreading of visceral leishmaniasis to new areas of the planet as a consequence of global warming

The immunopathology of canine vector-borne diseases

The canine vector-borne infectious diseases (CVBDs) are an emerging problem in veterinary medicine and the zoonotic potential of many of these agents is a significant consideration for human health. The successful diagnosis, treatment and prevention of these infections is dependent upon firm understanding of the underlying immunopathology of the diseases in which there are unique tripartite interactions between the microorganism, the vector and the host immune system. Although significant advances have been made in the areas of molecular speciation and the epidemiology of these infections and their vectors, basic knowledge of the pathology and immunology of the diseases has lagged behind. This review summarizes recent studies of the pathology and host immune response in the major CVBDs (leishmaniosis, babesiosis, ehrlichiosis, hepatozoonosis, anaplasmosis, bartonellosis and borreliosis). The ultimate application of such immunological investigation is the development of effective vaccines. The current commercially available vaccines for canine leishmaniosis, babesiosis and borreliosis are reviewed

Tegumentary leishmaniasis and coinfections other than HIV

<div><p>Background</p><p>Tegumentary leishmaniasis (TL) is a disease of skin and/or mucosal tissues caused by <i>Leishmania</i> parasites. TL patients may concurrently carry other pathogens, which may influence the clinical outcome of TL.</p><p>Methodology and principal findings</p><p>This review focuses on the frequency of TL coinfections in human populations, interactions between <i>Leishmania</i> and other pathogens in animal models and human subjects, and implications of TL coinfections for clinical practice. For the purpose of this review, TL is defined as all forms of cutaneous (localised, disseminated, or diffuse) and mucocutaneous leishmaniasis. Human immunodeficiency virus (HIV) coinfection, superinfection with skin bacteria, and skin manifestations of visceral leishmaniasis are not included. We searched MEDLINE and other databases and included 73 records: 21 experimental studies in animals and 52 studies about human subjects (mainly cross-sectional and case studies). Several reports describe the frequency of <i>Trypanosoma cruzi</i> coinfection in TL patients in Argentina (about 41%) and the frequency of helminthiasis in TL patients in Brazil (15% to 88%). Different hypotheses have been explored about mechanisms of interaction between different microorganisms, but no clear answers emerge. Such interactions may involve innate immunity coupled with regulatory networks that affect quality and quantity of acquired immune responses. Diagnostic problems may occur when concurrent infections cause similar lesions (e.g., TL and leprosy), when different pathogens are present in the same lesions (e.g., <i>Leishmania</i> and <i>Sporothrix schenckii</i>), or when similarities between phylogenetically close pathogens affect accuracy of diagnostic tests (e.g., serology for leishmaniasis and Chagas disease). Some coinfections (e.g., helminthiasis) appear to reduce the effectiveness of antileishmanial treatment, and drug combinations may cause cumulative adverse effects.</p><p>Conclusions and significance</p><p>In patients with TL, coinfection is frequent, it can lead to diagnostic errors and delays, and it can influence the effectiveness and safety of treatment. More research is needed to unravel how coinfections interfere with the pathogenesis of TL.</p></div