Biomarkers of Safety and Immune Protection for Genetically Modified Live Attenuated Leishmania Vaccines Against Visceral Leishmaniasis – Discovery and Implications

PubMed Central

Gannavaram, Sreenivas; Dey, Ranadhir; Avishek, Kumar; Selvapandiyan, Angamuthu; Salotra, Poonam; Nakhasi, Hira L.

2014-01-01

Despite intense efforts there is no safe and efficacious vaccine against visceral leishmaniasis, which is fatal and endemic in many tropical countries. A major shortcoming in the vaccine development against blood-borne parasitic agents such as Leishmania is the inadequate predictive power of the early immune responses mounted in the host against the experimental vaccines. Often immune correlates derived from in-bred animal models do not yield immune markers of protection that can be readily extrapolated to humans. The limited efficacy of vaccines based on DNA, subunit, heat killed parasites has led to the realization that acquisition of durable immunity against the protozoan parasites requires a controlled infection with a live attenuated organism. Recent success of irradiated malaria parasites as a vaccine candidate further strengthens this approach to vaccination. We developed several gene deletion mutants in Leishmania donovani as potential live attenuated vaccines and reported extensively on the immunogenicity of LdCentrin1 deleted mutant in mice, hamsters, and dogs. Additional limited studies using genetically modified live attenuated Leishmania parasites as vaccine candidates have been reported. However, for the live attenuated parasite vaccines, the primary barrier against widespread use remains the absence of clear biomarkers associated with protection and safety. Recent studies in evaluation of vaccines, e.g., influenza and yellow fever vaccines, using systems biology tools demonstrated the power of such strategies in understanding the immunological mechanisms that underpin a protective phenotype. Applying similar tools in isolated human tissues such as PBMCs from healthy individuals infected with live attenuated parasites such as LdCen−/− in vitro followed by human microarray hybridization experiments will enable us to understand how early vaccine-induced gene expression profiles and the associated immune responses are coordinately regulated in normal

A lentiviral vaccine expressing KMP11-HASPB fusion protein increases immune response to Leishmania major in BALB/C.

PubMed

Mortazavidehkordi, Nahid; Fallah, Ali; Abdollahi, Abbas; Kia, Vahid; Khanahmad, Hossein; Najafabadi, Zahra Ghayour; Hashemi, Nooshin; Estiri, Bahareh; Roudbari, Zahra; Najafi, Ali; Farjadfar, Akbar; Hejazi, Seyed Hossein

2018-05-29

Hydrophilic acylated surface protein B (HASPB) is an immunogenic Leishmania-specific protein that antibodies are produced against it in the sera of Leishmania-infected individuals. Kinetoplastid membrane protein 11 (KMP11) is another Leishmania antigen and considered as the suitable candidate for vaccine development Leishmaniasis. It is a highly conserved surface protein expressed in both promastigotes and amastigotes. In this study, KMP11 and HASPB coding sequences were cloned into a pCDH-cGFP lentiviral vector as a fusion protein to be used as a DNA vaccine against L. major. The KMP11-HASPB fusion protein was successfully expressed as evidenced by RT-PCR and Western blot assays. The effect of the vaccine was determined by evaluating the level of IFN-γ, IL-10, IgG1, and IgG2a performed using ELISA as well as determining the parasite load after challenge with L. major in vaccinated mice. The results revealed that IFN-γ, IL-10, IgG1, and IgG2a significantly increased after vaccination using KMP11-HASPB-expressing lentiviruses in BALB/c mice. It is noteworthy that the level of IFN-γ and IgG2a was higher than that of IL-10 and IgG1, respectively, which indicates the activation Th1 cells, macrophages, and cellular immunity. Moreover, the parasite load in the spleen and lymph node of vaccinated mice after challenge was significantly lower than that of controls.

Cross-protective efficacy of Leishmania infantum LiHyD protein against tegumentary leishmaniasis caused by Leishmania major and Leishmania braziliensis species.

PubMed

Lage, Daniela Pagliara; Martins, Vívian Tamietti; Duarte, Mariana Costa; Costa, Lourena Emanuele; Tavares, Grasiele de Sousa Vieira; Ramos, Fernanda Fonseca; Chávez-Fumagalli, Miguel Angel; Menezes-Souza, Daniel; Roatt, Bruno Mendes; Tavares, Carlos Alberto Pereira; Coelho, Eduardo Antonio Ferraz

2016-06-01

Vaccination can be considered the most cost-effective strategy to control neglected diseases, but nowadays there is not an effective vaccine available against leishmaniasis. In the present study, a vaccine based on the combination of the Leishmania-specific hypothetical protein (LiHyD) with saponin was tested in BALB/c mice against infection caused by Leishmania major and Leishmania braziliensis species. This antigen was firstly identified in Leishmania infantum and showed to be protective against infection of BALB/c mice using this parasite species. The immunogenicity of rLiHyD/saponin vaccine was evaluated, and the results showed that immunized mice produced high levels of IFN-γ, IL-12 and GM-CSF after in vitro stimulation with rLiHyD, as well as by using L. major or L. braziliensis protein extracts. After challenge, vaccinated animals showed significant reductions in the infected footpad swellings, as well as in the parasite burden in the infection site, liver, spleen, and infected paws draining lymph nodes, when compared to those that were inoculated with the vaccine diluent (saline) or immunized with saponin. The immunization of rLiHyD without adjuvant was not protective against both challenges. The partial protection obtained by the rLiHyD/saponin vaccine was associated with a parasite-specific IL-12-dependent IFN-γ secretion, which was produced mainly by CD4(+) T cells. In these animals, a decrease in the parasite-mediated IL-4 and IL-10 responses, associated with the presence of high levels of LiHyD- and parasite-specific IgG2a isotype antibodies, were also observed. The present study showed that a hypothetical protein that was firstly identified in L. infantum, when combined to a Th1 adjuvant, was able to confer a cross-protection against highly infective stationary-phase promastigotes of two Leishmania species causing tegumentary leishmaniasis. Copyright © 2016 Elsevier B.V. All rights reserved.

Therapeutic vaccine of killed Leishmania amazonensis plus saponin reduced parasite burden in dogs naturally infected with Leishmania infantum.

PubMed

Viana, Kelvinson Fernandes; Lacerda, Giulia; Teixeira, Natália Soares; Rodrigues Cangussu, Alex Sander; Sousa Aguiar, Raimundo Wagner; Giunchetti, Rodolfo Cordeiro

2018-04-30

A key goal in the control of canine visceral leishmaniosis (CVL) has been the development of vaccines with a highly protective capability to interrupt the parasite transmission cycle. However, in addition to promising vaccine searches, researchers have sought to develop new drugs capable of eliminating parasites in humans and dogs. With that in mind, this study analyzed an immunotherapeutic approach in dogs naturally infected with Leishmania infantum. Fourteen dogs were divided into two groups and received a protocol of immunotherapeutic treatment with five doses of total antigens of Leishmania amazonensis or total antigens of L. amazonensis plus saponin (LaSap). All the animals were evaluated before and 90 and 180 days after treatment, hematology, liver and renal biochemical analyzes, serology, lymphoproliferation, and parasite load by qPCR. The results of immunotherapy with the LaSap vaccine were promising since it was able to preserve hematological and biochemical parameters, as well as improve the clinical status, reduce serum levels of IgG, induce a lymphoproliferative capacity against soluble antigens of L. infantum, and provide a marked reduction in the parasite load after LaSap immunotherapeutic treatment. The immunotherapy data demonstrated that LaSap offered the best formulation to induce clinical cure associated with a parasite load reduction in the skin. However, after 180 days of treatment, the animals again showed a slight increase in parasitism, indicating that immunotherapy does not promote sterilizing cure and a new immunotherapeutic intervention would be necessary to maintain low parasitism in dogs. Copyright © 2018 Elsevier B.V. All rights reserved.

Impact of LbSapSal Vaccine in Canine Immunological and Parasitological Features before and after Leishmania chagasi-Challenge

PubMed Central

Resende, Lucilene Aparecida; Aguiar-Soares, Rodrigo Dian de Oliveira; Gama-Ker, Henrique; Roatt, Bruno Mendes; de Mendonça, Ludmila Zanandreis; Alves, Marina Luiza Rodrigues; da Silveira-Lemos, Denise; Corrêa-Oliveira, Rodrigo; Martins-Filho, Olindo Assis; Araújo, Márcio Sobreira Silva; Fujiwara, Ricardo Toshio; Gontijo, Nelder Figueiredo; Reis, Alexandre Barbosa; Giunchetti, Rodolfo Cordeiro

2016-01-01

Dogs represent the most important domestic reservoir of L. chagasi (syn. L. infantum). A vaccine against canine visceral leishmaniasis (CVL) would be an important tool for decreasing the anxiety related to possible L. chagasi infection and for controlling human visceral leishmaniasis (VL). Because the sand fly salivary proteins are potent immunogens obligatorily co-deposited during transmission of Leishmania parasites, their inclusion in an anti-Leishmania vaccine has been investigated in past decades. We investigated the immunogenicity of the “LbSapSal” vaccine (L. braziliensis antigens, saponin as adjuvant, and Lutzomyia longipalpis salivary gland extract) in dogs at baseline (T0), during the post-vaccination protocol (T3rd) and after early (T90) and late (T885) times following L. chagasi-challenge. Our major data indicated that immunization with “LbSapSal” is able to induce biomarkers characterized by enhanced amounts of type I (tumor necrosis factor [TNF]-α, interleukin [IL]-12, interferon [IFN]-γ) cytokines and reduction in type II cytokines (IL-4 and TGF-β), even after experimental challenge. The establishment of a prominent pro-inflammatory immune response after “LbSapSal” immunization supported the increased levels of nitric oxide production, favoring a reduction in spleen parasitism (78.9%) and indicating long-lasting protection against L. chagasi infection. In conclusion, these results confirmed the hypothesis that the “LbSapSal” vaccination is a potential tool to control the Leishmania chagasi infection. PMID:27556586

Impact of LbSapSal Vaccine in Canine Immunological and Parasitological Features before and after Leishmania chagasi-Challenge.

PubMed

Resende, Lucilene Aparecida; Aguiar-Soares, Rodrigo Dian de Oliveira; Gama-Ker, Henrique; Roatt, Bruno Mendes; Mendonça, Ludmila Zanandreis de; Alves, Marina Luiza Rodrigues; Silveira-Lemos, Denise da; Corrêa-Oliveira, Rodrigo; Martins-Filho, Olindo Assis; Araújo, Márcio Sobreira Silva; Fujiwara, Ricardo Toshio; Gontijo, Nelder Figueiredo; Reis, Alexandre Barbosa; Giunchetti, Rodolfo Cordeiro

2016-01-01

Dogs represent the most important domestic reservoir of L. chagasi (syn. L. infantum). A vaccine against canine visceral leishmaniasis (CVL) would be an important tool for decreasing the anxiety related to possible L. chagasi infection and for controlling human visceral leishmaniasis (VL). Because the sand fly salivary proteins are potent immunogens obligatorily co-deposited during transmission of Leishmania parasites, their inclusion in an anti-Leishmania vaccine has been investigated in past decades. We investigated the immunogenicity of the "LbSapSal" vaccine (L. braziliensis antigens, saponin as adjuvant, and Lutzomyia longipalpis salivary gland extract) in dogs at baseline (T0), during the post-vaccination protocol (T3rd) and after early (T90) and late (T885) times following L. chagasi-challenge. Our major data indicated that immunization with "LbSapSal" is able to induce biomarkers characterized by enhanced amounts of type I (tumor necrosis factor [TNF]-α, interleukin [IL]-12, interferon [IFN]-γ) cytokines and reduction in type II cytokines (IL-4 and TGF-β), even after experimental challenge. The establishment of a prominent pro-inflammatory immune response after "LbSapSal" immunization supported the increased levels of nitric oxide production, favoring a reduction in spleen parasitism (78.9%) and indicating long-lasting protection against L. chagasi infection. In conclusion, these results confirmed the hypothesis that the "LbSapSal" vaccination is a potential tool to control the Leishmania chagasi infection.

Gentamicin-Attenuated Leishmania infantum Vaccine: Protection of Dogs against Canine Visceral Leishmaniosis in Endemic Area of Southeast of Iran

PubMed Central

Daneshvar, Hamid; Namazi, Mohammad Javad; Kamiabi, Hossein; Burchmore, Richard; Cleaveland, Sarah; Phillips, Stephen

2014-01-01

An attenuated line of Leishmania infantum (L. infantum H-line) has been established by culturing promastigotes in vitro under gentamicin pressure. A vaccine trial was conducted using 103 naive dogs from a leishmaniosis non-endemic area (55 vaccinated and 48 unvaccinated) brought into an endemic area of southeast Iran. No local and/or general indications of disease were observed in the vaccinated dogs immediately after vaccination. The efficacy of the vaccine was evaluated after 24 months (4 sandfly transmission seasons) by serological, parasitological analyses and clinical examination. In western blot analysis of antibodies to L. infantum antigens, sera from 10 out of 31 (32.2%) unvaccinated dogs, but none of the sera from vaccinated dogs which were seropositive at >100, recognized the 21 kDa antigen of L. infantum wild-type (WT). Nine out of 31 (29%) unvaccinated dogs, but none of vaccinated dogs, were positive for the presence of Leishmania DNA. One out of 46 (2.2%) vaccinated dogs and 9 out of 31 (29%) unvaccinated dogs developed clinical signs of disease. These results suggest that gentamicin-attenuated L. infantum induced a significant and strong protective effect against canine visceral leishmaniosis in the endemic area. PMID:24743691

Failure of a killed Leishmania amazonensis vaccine against American cutaneous leishmaniasis in Colombia.

PubMed

Vélez, Iván D; Gilchrist, Katherine; Arbelaez, María P; Rojas, Carlos A; Puerta, Juan A; Antunes, Carlos M F; Zicker, Fabio; Modabber, Farrokh

2005-08-01

We report the results of a double-blind, randomized, placebo-controlled clinical trial of a killed whole-cell Leishmania amazonensis candidate vaccine against American cutaneous leishmaniasis (CL) in Colombia. The trial subjects were 2597 healthy volunteers with negative leishmanin skin test (LST) selected from rural Colombian soldiers who were going to patrol endemic areas. They were randomized to receive either three doses of vaccine (n=1295) or placebo (n=1302) given at 20-day intervals. An active and passive case detection system was established to follow-up volunteers for 1 year after vaccination. Safety and efficacy of the vaccine were determined by comparing local and systemic adverse reactions after each dose and the incidence of parasitologically confirmed CL. In the vaccine and placebo groups 101 (7.7%) and 88 (6.8%) volunteers developed CL respectively. The vaccine was shown to be safe but offered no protection against CL caused by L. panamensis in the proposed vaccination schedule.

Killed but metabolically active Leishmania infantum as a novel whole-cell vaccine for visceral leishmaniasis.

PubMed

Bruhn, Kevin W; Birnbaum, Ron; Haskell, Jacquelyn; Vanchinathan, Veena; Greger, Stephanie; Narayan, Rupa; Chang, Pei-Lin; Tran, Thu Anh; Hickerson, Suzanne M; Beverley, Stephen M; Wilson, Mary E; Craft, Noah

2012-04-01

There are currently no effective vaccines for visceral leishmaniasis, the second most deadly parasitic infection in the world. Here, we describe a novel whole-cell vaccine approach using Leishmania infantum chagasi promastigotes treated with the psoralen compound amotosalen (S-59) and low doses of UV A radiation. This treatment generates permanent, covalent DNA cross-links within parasites and results in Leishmania organisms termed killed but metabolically active (KBMA). In this report, we characterize the in vitro growth characteristics of both KBMA L. major and KBMA L. infantum chagasi. Concentrations of S-59 that generate optimally attenuated parasites were identified. Like live L. infantum chagasi, KBMA L. infantum chagasi parasites were able to initially enter liver cells in vivo after intravenous infection. However, whereas live L. infantum chagasi infection leads to hepatosplenomegaly in mice after 6 months, KBMA L. infantum chagasi parasites were undetectable in the organs of mice at this time point. In vitro, KBMA L. infantum chagasi retained the ability to enter macrophages and induce nitric oxide production. These characteristics of KBMA L. infantum chagasi correlated with the ability to prophylactically protect mice via subcutaneous vaccination at levels similar to vaccination with live, virulent organisms. Splenocytes from mice vaccinated with either live L. infantum chagasi or KBMA L. infantum chagasi displayed similar cytokine patterns in vitro. These results suggest that KBMA technology is a potentially safe and effective novel vaccine strategy against the intracellular protozoan L. infantum chagasi. This approach may represent a new method for whole-cell vaccination against other complex intracellular pathogens.

Comparative Evaluation of Two Vaccine Candidates against Experimental Leishmaniasis Due to Leishmania major Infection in Four Inbred Mouse Strains▿

PubMed Central

Benhnini, Fouad; Chenik, Mehdi; Laouini, Dhafer; Louzir, Hechmi; Cazenave, Pierre André; Dellagi, Koussay

2009-01-01

Experimental leishmaniasis in BALB/c and C57BL/6 mice are the most investigated murine models that were used for the preclinical evaluation of Leishmania vaccine candidates. We have previously described two new inbred mouse strains named PWK and MAI issued from feral founders that also support the development of experimental leishmaniasis due to L. major. In this study, we sought to determine whether different mouse inbred strains generate concordant or discordant results when used to evaluate the potential of Leishmania proteins to protect against experimental leishmaniasis. To this end, two Leishmania proteins, namely, LACK (for Leishmania homolog of receptor for activated C kinase) and LmPDI (for L. major protein disulfide isomerase) were compared for their capacity to protect against experimental leishmaniasis in PWK, MAI, BALB/c, and C57BL/6 inbred mouse strains. Our data show that the capacity of Leishmania proteins to confer protection depends on the mouse strain used, stressing the important role played by the genetic background in shaping the immune response against the pathogen. These results may have important implications for the preclinical evaluation of candidate Leishmania vaccines: rather than using a single mouse strain, a panel of different inbred strains of various genetic backgrounds should be tested in parallel. The antigen that confers protection in the larger range of inbred strains may have better chances to be also protective in outbred human populations and should be selected for clinical trials. PMID:19726616

Comparative evaluation of two vaccine candidates against experimental leishmaniasis due to Leishmania major infection in four inbred mouse strains.

PubMed

Benhnini, Fouad; Chenik, Mehdi; Laouini, Dhafer; Louzir, Hechmi; Cazenave, Pierre André; Dellagi, Koussay

2009-11-01

Experimental leishmaniasis in BALB/c and C57BL/6 mice are the most investigated murine models that were used for the preclinical evaluation of Leishmania vaccine candidates. We have previously described two new inbred mouse strains named PWK and MAI issued from feral founders that also support the development of experimental leishmaniasis due to L. major. In this study, we sought to determine whether different mouse inbred strains generate concordant or discordant results when used to evaluate the potential of Leishmania proteins to protect against experimental leishmaniasis. To this end, two Leishmania proteins, namely, LACK (for Leishmania homolog of receptor for activated C kinase) and LmPDI (for L. major protein disulfide isomerase) were compared for their capacity to protect against experimental leishmaniasis in PWK, MAI, BALB/c, and C57BL/6 inbred mouse strains. Our data show that the capacity of Leishmania proteins to confer protection depends on the mouse strain used, stressing the important role played by the genetic background in shaping the immune response against the pathogen. These results may have important implications for the preclinical evaluation of candidate Leishmania vaccines: rather than using a single mouse strain, a panel of different inbred strains of various genetic backgrounds should be tested in parallel. The antigen that confers protection in the larger range of inbred strains may have better chances to be also protective in outbred human populations and should be selected for clinical trials.

Cross-protective effect of a combined L5 plus L3 Leishmania major ribosomal protein based vaccine combined with a Th1 adjuvant in murine cutaneous and visceral leishmaniasis

PubMed Central

2014-01-01

Background Two Leishmania major ribosomal proteins L3 (LmL3) and L5 (LmL5) have been described as protective molecules against cutaneous leishmaniasis due to infection with L. major and Leishmania braziliensis in BALB/c mice when immunized with a Th1 adjuvant (non-methylated CpG-oligodeoxynucleotides; CpG-ODN). In the present study we analyzed the cross-protective efficacy of an LmL3-LmL5-CpG ODN combined vaccine against infection with Leishmania amazonensis and Leishmania chagasi (syn. Leishmania infantum) the etiologic agents of different clinical forms of human leishmaniasis in South America. Methods The combined vaccine was administered subcutaneously to BALB/c mice. After immunization the cellular and humoral responses elicited were analyzed. Mice were independently challenged with L. amazonensis and L. chagasi. The size of the cutaneous lesions caused by the infection with the first species, the parasite loads and the immune response in both infection models were analyzed nine weeks after challenge. Results Mice vaccinated with the combined vaccine showed a Th1-like response against LmL3 and LmL5. Vaccinated mice were able to delay lesion development due to L. amazonensis infection and to control parasite loads in the site of infection. A reduction of the parasite burden in the lymph nodes draining the site of infection and in the liver and spleen was observed in the vaccinated mice after a subcutaneous infection with L. chagasi. In both models of infection, protection was correlated to parasite antigen-specific production of IFN-γ and down-regulation of parasite-mediated IL-4 and IL-10 responses. Conclusions The data presented here demonstrate the potential use of L. major L3 and L5 recombinant ribosomal proteins for the development of vaccines against various Leishmania species. PMID:24382098

Cross-protective effect of a combined L5 plus L3 Leishmania major ribosomal protein based vaccine combined with a Th1 adjuvant in murine cutaneous and visceral leishmaniasis.

PubMed

Ramirez, Laura; Corvo, Laura; Duarte, Mariana C; Chávez-Fumagalli, Miguel A; Valadares, Diogo G; Santos, Diego M; de Oliveira, Camila I; Escutia, Marta R; Alonso, Carlos; Bonay, Pedro; Tavares, Carlos A P; Coelho, Eduardo A F; Soto, Manuel

2014-01-02

Two Leishmania major ribosomal proteins L3 (LmL3) and L5 (LmL5) have been described as protective molecules against cutaneous leishmaniasis due to infection with L. major and Leishmania braziliensis in BALB/c mice when immunized with a Th1 adjuvant (non-methylated CpG-oligodeoxynucleotides; CpG-ODN). In the present study we analyzed the cross-protective efficacy of an LmL3-LmL5-CpG ODN combined vaccine against infection with Leishmania amazonensis and Leishmania chagasi (syn. Leishmania infantum) the etiologic agents of different clinical forms of human leishmaniasis in South America. The combined vaccine was administered subcutaneously to BALB/c mice. After immunization the cellular and humoral responses elicited were analyzed. Mice were independently challenged with L. amazonensis and L. chagasi. The size of the cutaneous lesions caused by the infection with the first species, the parasite loads and the immune response in both infection models were analyzed nine weeks after challenge. Mice vaccinated with the combined vaccine showed a Th1-like response against LmL3 and LmL5. Vaccinated mice were able to delay lesion development due to L. amazonensis infection and to control parasite loads in the site of infection. A reduction of the parasite burden in the lymph nodes draining the site of infection and in the liver and spleen was observed in the vaccinated mice after a subcutaneous infection with L. chagasi. In both models of infection, protection was correlated to parasite antigen-specific production of IFN-γ and down-regulation of parasite-mediated IL-4 and IL-10 responses. The data presented here demonstrate the potential use of L. major L3 and L5 recombinant ribosomal proteins for the development of vaccines against various Leishmania species.

LBSapSal-vaccinated dogs exhibit increased circulating T-lymphocyte subsets (CD4+ and CD8+) as well as a reduction of parasitism after challenge with Leishmania infantum plus salivary gland of Lutzomyia longipalpis

PubMed Central

2014-01-01

Background The development of a protective vaccine against canine visceral leishmaniasis (CVL) is an alternative approach for interrupting the domestic cycle of Leishmania infantum. Given the importance of sand fly salivary proteins as potent immunogens obligatorily co-deposited during transmission of Leishmania parasites, their inclusion in an anti-Leishmania vaccine has been investigated in the last few decades. In this context, we previously immunized dogs with a vaccine composed of L. braziliensis antigens plus saponin as the adjuvant and sand fly salivary gland extract (LBSapSal vaccine). This vaccine elicited an increase in both anti-saliva and anti-Leishmania IgG isotypes, higher counts of specific circulating CD8+ T cells, and high NO production. Methods We investigated the immunogenicity and protective effect of LBSapSal vaccination after intradermal challenge with 1 × 107 late-log-phase L. infantum promastigotes in the presence of sand fly saliva of Lutzomyia longipalpis. The dogs were followed for up to 885 days after challenge. Results The LBSapSal vaccine presents extensive antigenic diversity with persistent humoral and cellular immune responses, indicating resistance against CVL is triggered by high levels of total IgG and its subtypes (IgG1 and IgG2); expansion of circulating CD5+, CD4+, and CD8+ T lymphocytes and is Leishmania-specific; and reduction of splenic parasite load. Conclusions These results encourage further study of vaccine strategies addressing Leishmania antigens in combination with proteins present in the saliva of the vector. PMID:24507702

Cross-protective efficacy from a immunogen firstly identified in Leishmania infantum against tegumentary leishmaniasis.

PubMed

Martins, V T; Lage, D P; Duarte, M C; Costa, L E; Chávez-Fumagalli, M A; Roatt, B M; Menezes-Souza, D; Tavares, C A P; Coelho, E A F

2016-02-01

Experimental vaccine candidates have been evaluated to prevent leishmaniasis, but no commercial vaccine has been proved to be effective against more than one parasite species. LiHyT is a Leishmania-specific protein that was firstly identified as protective against Leishmania infantum. In this study, LiHyT was evaluated as a vaccine to against two Leishmania species causing tegumentary leishmaniasis (TL): Leishmania major and Leishmania braziliensis. BALB/c mice were immunized with rLiHyT plus saponin and lately challenged with promastigotes of the two parasite species. The immune response generated was evaluated before and 10 weeks after infection, as well as the parasite burden at this time after infection. The vaccination induced a Th1 response, which was characterized by the production of IFN-γ, IL-12 and GM-CSF, as well as by high levels of IgG2a antibodies, after in vitro stimulation using both the protein and parasite extracts. After challenge, vaccinated mice showed significant reductions in their infected footpads, as well as in the parasite burden in the tissue and organs evaluated, when compared to the control groups. The anti-Leishmania Th1 response was maintained after infection, being the IFN-γ production based mainly on CD4(+) T cells. We described one conserved Leishmania-specific protein that could compose a pan-Leishmania vaccine. © 2016 John Wiley & Sons Ltd.

Comparative assessment of a DNA and protein Leishmania donovani gamma glutamyl cysteine synthetase vaccine to cross-protect against murine cutaneous leishmaniasis caused by L. major or L. mexicana infection.

PubMed

Campbell, S A; Alawa, J; Doro, B; Henriquez, F L; Roberts, C W; Nok, A; Alawa, C B I; Alsaadi, M; Mullen, A B; Carter, K C

2012-02-08

Leishmaniasis is a major health problem and it is estimated that 12 million people are currently infected. A vaccine which could cross-protect people against different Leishmania spp. would facilitate control of this disease as more than one species of Leishmania may be present. In this study the ability of a DNA vaccine, using the full gene sequence for L. donovani gamma glutamyl cysteine synthetase (γGCS) incorporated in the pVAX vector (pVAXγGCS), and a protein vaccine, using the corresponding recombinant L. donovani γGCS protein (LdγGCS), to protect against L. major or L. mexicana infection was evaluated. DNA vaccination gave transient protection against L. major and no protection against L. mexicana despite significantly enhancing specific antibody titres in vaccinated infected mice compared to infected controls. Vaccination with the LdγGCS protected against both species but only if the protein was incorporated into non-ionic surfactant vesicles for L. mexicana. The results of this study indicate that a L. donovani γGCS vaccine could be used to vaccinate against more than one Leishmania species but only if the recombinant protein is used. Copyright © 2012 Elsevier Ltd. All rights reserved.

Leishmania mexicana Gp63 cDNA Using Gene Gun Induced Higher Immunity to L. mexicana Infection Compared to Soluble Leishmania Antigen in BALB/C

PubMed Central

Rezvan, H; Rees, R; Ali, SA

2011-01-01

Background Leishmaniasis is a worldwide disease prevalent in tropical and sub tropical countries. Many attempts have been made and different strategies have been approached to develop a potent vaccine against Leishmania. DNA immunisation is a method, which is shown to be effective in Leishmania vaccination. Leishmania Soluble Antigen (SLA) has also recently been used Leishmania vaccination. Methods The immunity generated by SLA and L. mexicana gp63 cDNA was compared in groups of 6 mice, which were statistically analysed by student t- test with the P-value of 0.05. SLA was administered by two different methods; intramuscular injection and injection of dendritic cells (DCs) loaded with SLA. L. mexicana gp63 cDNA was administered by the gene gun. Results Immunisation of BALB/c mice with L. mexicana gp63 resulted in high levels of Th1-type immune response and cytotoxic T lymphocytes (CTL) activity, which were accompanied with protection induced by the immunisation against L. mexicana infection. In contrast, administration of SLA, produced a mixed Th1/Th2-type immune responses as well as a high level of CTL activity but did not protect mice from the infection. Conclusion The results indicate higher protection by DNA immunisation using L. mexicana gp63 cDNA compared to SLA, which is accompanied by a high level of Th1 immune response. However, the CTL activity does not necessarily correlate with the protection induced by the vaccine. Also, gene gun immunisation is a potential approach in Leishmania vaccination. These findings would be helpful in opening new windows in Leishmania vaccine research. PMID:22347315

Vaccination with Leishmania histone H1-pulsed dendritic cells confers protection in murine visceral leishmaniasis.

PubMed

Agallou, Maria; Smirlis, Despina; Soteriadou, Ketty P; Karagouni, Evdokia

2012-07-20

Visceral leishmaniasis is the most severe form of leishmaniases affecting millions of people worldwide often resulting in death despite optimal therapy. Thus, there is an urgent need for the development of effective anti-infective vaccine(s). In the present study, we evaluated the prophylactic value of bone marrow-derived dendritic cells (BM-DCs) pulsed with the Leishmania (L.) infantum histone H1. We developed fully mature BM-DCs characterized by enhanced capacity of IL-12 production after ex vivo pulsing with GST-LeishH1. Intravenous administration of these BM-DCs in naive BALB/c mice resulted in antigen-specific spleenocyte proliferation and IgG1 isotype antibody production and conferred protection against experimental challenge with L. infantum independently of CpG oligonucleotides (ODNs) co-administration. Protection was associated with a pronounced enhancement of parasite-specific IFNγ-producing cells and reduction of cells producing IL-10, whereas IL-4 production was comparable in protected and non-protected mice. The polarization of immune responses to Th1 type was further confirmed by the elevation of parasite-specific IgG2a/IgG1 ratio in protected mice. The above data indicate the immunostimulatory capacity of Leishmania histone H1 and further support its exploitation as a candidate protein for vaccine development against leishmaniasis. Copyright © 2012 Elsevier Ltd. All rights reserved.

Preparation of live attenuated leishmania parasites by using laser technology

NASA Astrophysics Data System (ADS)

Hussain, Nabiha; Alkhouri, Hassan; Haddad, Shaden

2018-05-01

Leishmaniasis is a parasitic disease of humans, affecting the skin, mucosal and/or internal organs, caused by flagellate protozoa Leishmania of the Trypanosomatidae family. Leishmania would be one for which a vaccine could be developed with relative ease. Many studies mount an effective response that resolves the infection and confers solid immunity to reinfection and suggesting that infection may be a prerequisite for immunological memory. Genetically altered live attenuated parasites with controlled infectivity could achieve such immunological memory. Recent concepts include use of genetically modified live-attenuated Leishmania parasites, and proteomics approach for the search of a cross-protective leishmanial vaccine that would ideally protect against both cutaneous and visceral forms of the disease. No licensed vaccine is available till date against any form of leishmaniasis. The present study evaluated role of laser technology in development of a safe live Leishmania vaccine, a vaccine is a biological preparation that improves immunity to a particular disease, and is often made from weakened or killed forms of LPs. The parasite culture was expanded in RPMI 1640 medium with 10% fetal calf serum (FCS) and grown until stationary phase for experiments. 80 samples of leishmania promastigotes (Culture media of LPs) were exposed to Nd:YAG laser (wavelength 1064 nm, single spot or double) with different outputs powers (7w, 100 Hz, 99.03w/cm2, 0.99 J/cm2 and 8 w, 100 Hz, 113.18w/cm2 1.13J/cm2)) for suitable exposer times. The effect of semiconductor laser (wavelength 810 nm, 7w, 2000 Hz, 99.03w/cm2, 0.05 J/cm2) or (7 w, 500 Hz, 99.03 w/cm2, 0.2J/cm2) single spot or double with long exposure times. The viability of Leishmania parasites was measured using XTT method; viable parasites were decreased with long exposure times. XTT test referred both these wavelengths were effective in killing percentage of Leishmania promastigotes, the remaining were devoid flagellum that

Immunization with antigenic extracts of Leishmania associated with Montanide ISA 763 adjuvant induces partial protection in BALB/c mice against Leishmania (Leishmania) amazonensis infection.

PubMed

Cargnelutti, Diego Esteban; Salomón, María Cristina; Celedon, Verónica; García Bustos, María Fernanda; Morea, Gastón; Cuello-Carrión, Fernando Darío; Scodeller, Eduardo Alberto

2016-02-01

A proper adjuvant has a relevant role in vaccine formulations to generate an effective immune response. In this study, total Leishmania antigen (TLA) formulated with Montanide ISA 763 or R848 as adjuvants were evaluated as a first generation Leishmania vaccine in a murine model. Immunization protocols were tested in BALB/c mice with a subcutaneous prime/boost regimen with an interval of 3 weeks. Mice immunized with unadjuvanted TLA and phosphate-buffered saline (PBS) served as control groups. On Day 21 and Day 36 of the protocol, we evaluated the humoral immune response induced by each formulation. Fifteen days after the boost, the immunized mice were challenged with 1 × 10(5) promastigotes of Leishmania (Leishmania) amazonensis in the right footpad (RFP). The progress of the infection was followed for 10 weeks; at the end of this period, histopathological studies were performed in the RFP. Vaccines formulated with Montanide ISA 763 generated an increase in the production of immunoglobulin G (IgG; p < 0.05) compared with the control group. There were no statistically significant differences in IgG1 production between the study groups. However, immunization with TLA-Montanide ISA 763 resulted in an increase in IgG2a compared to the unadjuvanted control (p < 0.001). Also noteworthy was the fact that a significant reduction in swelling and histopathological damage of the RFP was recorded with the Montanide ISA 763 formulation. We conclude that the immunization of BALB/c mice with a vaccine formulated with TLA and Montanide ISA 763 generated a protective immune response against L. (L.) amazonensis, characterized by an intense production of IgG2a. Copyright © 2014. Published by Elsevier B.V.

Efficacy of Recombinant Canine Distemper Virus Expressing Leishmania Antigen against Leishmania Challenge in Dogs.

PubMed

Miura, Ryuichi; Kooriyama, Takanori; Yoneda, Misako; Takenaka, Akiko; Doki, Miho; Goto, Yasuyuki; Sanjoba, Chizu; Endo, Yasuyuki; Fujiyuki, Tomoko; Sugai, Akihiro; Tsukiyama-Kohara, Kyoko; Matsumoto, Yoshitsugu; Sato, Hiroki; Kai, Chieko

2015-01-01

Canine distemper virus (CDV) vaccination confers long-term protection against CDV reinfection. To investigate the utility of CDV as a polyvalent vaccine vector for Leishmania, we generated recombinant CDVs, based on an avirulent Yanaka strain, that expressed Leishmania antigens: LACK, TSA, or LmSTI1 (rCDV-LACK, rCDV-TSA, and rCDV-LmSTI1, respectively). Dogs immunized with rCDV-LACK were protected against challenge with lethal doses of virulent CDV, in the same way as the parental Yanaka strain. To evaluate the protective effects of the recombinant CDVs against cutaneous leishmaniasis in dogs, dogs were immunized with one recombinant CDV or a cocktail of three recombinant CDVs, before intradermal challenge (in the ears) with infective-stage promastigotes of Leishmania major. Unvaccinated dogs showed increased nodules with ulcer formation after 3 weeks, whereas dogs immunized with rCDV-LACK showed markedly smaller nodules without ulceration. Although the rCDV-TSA- and rCDV-LmSTI1-immunized dogs showed little protection against L. major, the cocktail of three recombinant CDVs more effectively suppressed the progression of nodule formation than immunization with rCDV-LACK alone. These results indicate that recombinant CDV is suitable for use as a polyvalent live attenuated vaccine for protection against both CDV and L. major infections in dogs.

Total Leishmania antigens with Poly(I:C) induce Th1 protective response.

PubMed

Sanchez, M V; Eliçabe, R J; Di Genaro, M S; Germanó, M J; Gea, S; García Bustos, M F; Salomón, M C; Scodeller, E A; Cargnelutti, D E

2017-11-01

Our proposal was to develop a vaccine based on total Leishmania antigens (TLA) adjuvanted with polyinosinic-polycytidylic acid [Poly(I:C)] able to induce a Th1 response which can provide protection against Leishmania infection. Mice were vaccinated with two doses of TLA-Poly(I:C) administered by subcutaneous route at 3-week interval. Humoral and cellular immune responses induced by the immunization were measured. The protective efficacy of the vaccine was evaluated by challenging mice with infective promastigotes of Leishmania (Leishmania) amazonensis into the footpad. Mice vaccinated with TLA-Poly(I:C) showed a high anti-Leishmania IgG titre, as well as increased IgG1 and IgG2a subclass titres compared with mice vaccinated with the TLA alone. The high IgG2a indicated a Th1 bias response induced by the TLA-Poly(I:C) immunization. Accordingly, the cellular immune response elicited by the formulation was characterized by an increased production of IFN-γ and no significant production of IL-4. The TLA-Poly(I:C) immunization elicited good protection, which was associated with decreased footpad swelling, a lower parasite load and a reduced histopathological alteration in the footpad. Our findings demonstrate a promising vaccine against cutaneous leishmaniasis that is relatively economic and easy to develop and which should be taken into account for preventing leishmaniasis in developing countries. © 2017 John Wiley & Sons Ltd.

Recombinant NAD-dependent SIR-2 protein of Leishmania donovani: immunobiochemical characterization as a potential vaccine against visceral leishmaniasis.

PubMed

Baharia, Rajendra K; Tandon, Rati; Sharma, Tanuj; Suthar, Manish K; Das, Sanchita; Siddiqi, Mohammad Imran; Saxena, Jitendra Kumar; Sundar, Shaym; Sunder, Shyam; Dube, Anuradha

2015-03-01

The development of a vaccine conferring long-lasting immunity remains a challenge against visceral leishmaniasis (VL). Immunoproteomic characterization of Leishmania donovani proteins led to the identification of a novel protein NAD+-dependent Silent Information regulatory-2 (SIR2 family or sirtuin) protein (LdSir2RP) as one of the potent immunostimulatory proteins. Proteins of the SIR2 family are characterized by a conserved catalytic domain that exerts unique NAD-dependent deacetylase activity. In the present study, an immunobiochemical characterization of LdSir2RP and further evaluation of its immunogenicity and prophylactic potential was done to assess for its possible involvement as a vaccine candidate against leishmaniasis. LdSir2RP was successfully cloned, expressed and purified. The gene was present as a monomeric protein of ~45 kDa and further established by the crosslinking experiment. rLdSir2RP shown cytosolic localization in L. donovani and demonstrating NAD+-dependent deacetylase activity. Bioinformatic analysis also confirmed that LdSir2RP protein has NAD binding domain. The rLdSir2RP was further assessed for its cellular response by lymphoproliferative assay and cytokine ELISA in cured Leishmania patients and hamsters (Mesocricetus auratus) in comparison to soluble Leishmania antigen and it was observed to stimulate the production of IFN-γ, IL-12 and TNF-α significantly but not the IL-4 and IL-10. The naïve hamsters when vaccinated with rLdSir2RP alongwith BCG resisted the L. donovani challenge to the tune of ~75% and generated strong IL-12 and IFN-γ mediated Th1 type immune response thereof. The efficacy was further supported by remarkable increase in IgG2 antibody level which is indicative of Th1 type of protective response. Further, with a possible implication in vaccine design against VL, identification of potential T-cell epitopes of rLdSir2RP was done using computational approach. The immunobiochemical characterization strongly suggest the

Efficacy of Recombinant Canine Distemper Virus Expressing Leishmania Antigen against Leishmania Challenge in Dogs

PubMed Central

Yoneda, Misako; Takenaka, Akiko; Doki, Miho; Goto, Yasuyuki; Sanjoba, Chizu; Endo, Yasuyuki; Fujiyuki, Tomoko; Sugai, Akihiro; Tsukiyama-Kohara, Kyoko; Matsumoto, Yoshitsugu; Sato, Hiroki; Kai, Chieko

2015-01-01

Canine distemper virus (CDV) vaccination confers long-term protection against CDV reinfection. To investigate the utility of CDV as a polyvalent vaccine vector for Leishmania, we generated recombinant CDVs, based on an avirulent Yanaka strain, that expressed Leishmania antigens: LACK, TSA, or LmSTI1 (rCDV–LACK, rCDV–TSA, and rCDV–LmSTI1, respectively). Dogs immunized with rCDV-LACK were protected against challenge with lethal doses of virulent CDV, in the same way as the parental Yanaka strain. To evaluate the protective effects of the recombinant CDVs against cutaneous leishmaniasis in dogs, dogs were immunized with one recombinant CDV or a cocktail of three recombinant CDVs, before intradermal challenge (in the ears) with infective-stage promastigotes of Leishmania major. Unvaccinated dogs showed increased nodules with ulcer formation after 3 weeks, whereas dogs immunized with rCDV–LACK showed markedly smaller nodules without ulceration. Although the rCDV–TSA- and rCDV–LmSTI1-immunized dogs showed little protection against L. major, the cocktail of three recombinant CDVs more effectively suppressed the progression of nodule formation than immunization with rCDV–LACK alone. These results indicate that recombinant CDV is suitable for use as a polyvalent live attenuated vaccine for protection against both CDV and L. major infections in dogs. PMID:26162094

Aminophthalocyanine-Mediated Photodynamic Inactivation of Leishmania tropica

PubMed Central

Al-Qahtani, Ahmed; Alkahtani, Saad; Kolli, Bala; Tripathi, Pankaj; Dutta, Sujoy; Al-Kahtane, Abdullah A.; Jiang, Xiong-Jie; Ng, Dennis K. P.

2016-01-01

Photodynamic inactivation of Leishmania spp. requires the cellular uptake of photosensitizers, e.g., endocytosis of silicon(IV)-phthalocyanines (PC) axially substituted with bulky ligands. We report here that when substituted with amino-containing ligands, the PCs (PC1 and PC2) were endocytosed and displayed improved potency against Leishmania tropica promastigotes and axenic amastigotes in vitro. The uptake of these PCs by both Leishmania stages followed saturation kinetics, as expected. Sensitive assays were developed for assessing the photodynamic inactivation of Leishmania spp. by rendering them fluorescent in two ways: transfecting promastigotes to express green fluorescent protein (GFP) and loading them with carboxyfluorescein succinimidyl ester (CFSE). PC-sensitized Leishmania tropica strains were seen microscopically to lose their motility, structural integrity, and GFP/CFSE fluorescence after exposure to red light (wavelength, ∼650 nm) at a fluence of 1 to 2 J cm−2. Quantitative fluorescence assays based on the loss of GFP/CFSE from live Leishmania tropica showed that PC1 and PC2 dose dependently sensitized both stages for photoinactivation, consistent with the results of a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell viability assay. Leishmania tropica strains are >100 times more sensitive than their host cells or macrophages to PC1- and PC2-mediated photoinactivation, judging from the estimated 50% effective concentrations (EC50s) of these cells. Axial substitution of the PC with amino groups instead of other ligands appears to increase its leishmanial photolytic activity by up to 40-fold. PC1 and PC2 are thus potentially useful for photodynamic therapy of leishmaniasis and for oxidative photoinactivation of Leishmania spp. for use as vaccines or vaccine carriers. PMID:26824938

Identification of Leishmania infantum chagasi proteins in urine of patients with visceral leishmaniasis: a promising antigen discovery approach of vaccine candidates.

PubMed

Kashino, S S; Abeijon, C; Qin, L; Kanunfre, K A; Kubrusly, F S; Silva, F O; Costa, D L; Campos, D; Costa, C H N; Raw, I; Campos-Neto, A

2012-07-01

Visceral leishmaniasis (VL) is a serious lethal parasitic disease caused by Leishmania donovani in Asia and by Leishmania infantum chagasi in southern Europe and South America. VL is endemic in 47 countries with an annual incidence estimated to be 500,000 cases. This high incidence is due in part to the lack of an efficacious vaccine. Here, we introduce an innovative approach to directly identify parasite vaccine candidate antigens that are abundantly produced in vivo in humans with VL. We combined RP-HPLC and mass spectrometry and categorized three L. infantum chagasi proteins, presumably produced in spleen, liver and bone marrow lesions and excreted in the patients' urine. Specifically, these proteins were the following: Li-isd1 (XP_001467866.1), Li-txn1 (XP_001466642.1) and Li-ntf2 (XP_001463738.1). Initial vaccine validation studies were performed with the rLi-ntf2 protein produced in Escherichia coli mixed with the adjuvant BpMPLA-SE. This formulation stimulated potent Th1 response in BALB/c mice. Compared to control animals, mice immunized with Li-ntf2+ BpMPLA-SE had a marked parasite burden reduction in spleens at 40 days post-challenge with virulent L. infantum chagasi. These results strongly support the proposed antigen discovery strategy of vaccine candidates to VL and opens novel possibilities for vaccine development to other serious infectious diseases. © 2012 Blackwell Publishing Ltd.

Innate Immunity against Leishmania Infections

PubMed Central

Gurung, Prajwal; Kanneganti, Thirumala-Devi

2015-01-01

Leishmaniasis is a major health problem that affects more than 300 million people throughout the world. The morbidity associated with the disease causes serious economic burden in Leishmania endemic regions. Despite the morbidity and economic burden associated with Leishmaniasis, this disease rarely gets noticed and is still categorized under neglected tropical diseases. The lack of research combined with the ability of Leishmania to evade immune recognition has rendered our efforts to design therapeutic treatments or vaccines challenging. Herein, we review the literature on Leishmania from innate immune perspective and discuss potential problems as well as solutions and future directions that could aid in identifying novel therapeutic targets to eliminate this parasite. PMID:26249747

DNA vaccination with a plasmid encoding LACK-TSA fusion against Leishmania major infection in BALB/c mice.

PubMed

Maspi, N; Ghaffarifar, F; Sharifi, Z; Dalimi, A; Khademi, S Z

2017-12-01

Vaccination would be the most important strategy for the prevention and elimination of leishmaniasis. The aim of the present study was to compare the immune responses induced following DNA vaccination with LACK (Leishmania analogue of the receptor kinase C), TSA (Thiol-specific-antioxidant) genes alone or LACK-TSA fusion against cutaneous leishmaniasis (CL). Cellular and humoral immune responses were evaluated before and after challenge with Leishmania major (L. major). In addition, the mean lesion size was also measured from 3th week post-infection. All immunized mice showed a partial immunity characterized by higher interferon (IFN)-γ and Immunoglobulin G (IgG2a) levels compared to control groups (p<0.05). IFN-γ/ Interleukin (IL)-4 and IgG2a/IgG1 ratios demonstrated the highest IFN-γ and IgG2a levels in the group receiving LACK-TSA fusion. Mean lesion sizes reduced significantly in all immunized mice compared with control groups at 7th week post-infection (p<0.05). In addition, there was a significant reduction in mean lesion size of LACK-TSA and TSA groups than LACK group after challenge (p<0.05). In the present study, DNA immunization promoted Th1 immune response and confirmed the previous observations on immunogenicity of LACK and TSA antigens against CL. Furthermore, this study demonstrated that a bivalent vaccine can induce stronger immune responses and protection against infectious challenge with L. major.

Monocyte Chemotactic Protein 1 in Plasma from Soluble Leishmania Antigen-Stimulated Whole Blood as a Potential Biomarker of the Cellular Immune Response to Leishmania infantum

PubMed Central

Ibarra-Meneses, Ana V.; Sanchez, Carmen; Alvar, Jorge; Moreno, Javier; Carrillo, Eugenia

2017-01-01

New biomarkers are needed to identify asymptomatic Leishmania infection as well as immunity following vaccination or treatment. With the aim of finding a robust biomarker to assess an effective cellular immune response, monocyte chemotactic protein 1 (MCP-1) was examined in plasma from soluble Leishmania antigen (SLA)-stimulated whole blood collected from subjects living in a Leishmania infantum-endemic area. MCP-1, expressed 110 times more strongly than IL-2, identified 87.5% of asymptomatic subjects and verified some asymptomatic subjects close to the cutoff. MCP-1 was also significantly elevated in all patients cured of visceral leishmaniasis (VL), unlike IL-2, indicating the specific memory response generated against Leishmania. These results show MCP-1 to be a robust candidate biomarker of immunity that could be used as a marker of cure and to both select and follow the population in vaccine phase I–III human clinical trials with developed rapid, easy-to-use field tools. PMID:29033933

A vaccine composed of a hypothetical protein and the eukaryotic initiation factor 5a from Leishmania braziliensis cross-protection against Leishmania amazonensis infection.

PubMed

Duarte, Mariana Costa; Lage, Daniela Pagliara; Martins, Vívian Tamietti; Costa, Lourena Emanuele; Carvalho, Ana Maria Ravena Severino; Ludolf, Fernanda; Santos, Thaís Teodoro de Oliveira; Vale, Danniele Luciana; Roatt, Bruno Mendes; Menezes-Souza, Daniel; Fernandes, Ana Paula; Tavares, Carlos Alberto Pereira; Coelho, Eduardo Antonio Ferraz

2017-02-01

In the present study, two proteins cloned from Leishmania braziliensis species, a hypothetical protein (LbHyp) and the eukaryotic initiation factor 5a (EiF5a), were evaluated to protect BALB/c mice against L. amazonensis infection. The animals were immunized with the antigens, either separately or in combination, using saponin as an immune adjuvant in both cases. Spleen cells from vaccinated and later infected mice produced significantly higher levels of protein and parasite-specific IFN-γ, IL-12, and GM-CSF, in addition to low levels of IL-4 and IL-10. Evaluating the parasite load by means of a limiting dilution technique and quantitative Real-Time PCR, vaccinated animals presented significant reductions in the parasite load in both infected tissues and organs, as well as lower footpad swelling, when compared to the control (saline and saponin) groups. The best results regarding the protection of the animals were achieved when the combined vaccine was administered into the animals. Protection was associated with an IFN-γ production against parasite antigens, which was mediated by both CD4 + and CD8 + T cells and correlated with antileishmanial nitrite production. In conclusion, data from the present study show that this polyprotein vaccine, which combines two L. braziliensis proteins, can induce protection against L. amazonensis infection. Copyright © 2016 Elsevier GmbH. All rights reserved.

Therapeutic vaccines for leishmaniasis.

PubMed

Khamesipour, Ali

2014-11-01

Numerous therapeutic strategies are used to treat leishmaniasis. The treatment of cutaneous leishmaniasis (CL) is solely depends on antimonate derivatives with safety issues and questionable efficacy and there is no fully effective modality to treat CL caused by Leishmania tropica and Leishmania braziliensis. There is no prophylactic vaccine available against any form of leishmaniasis. Immunotherapy for CL has a long history; immunotherapy trials of first and second generation vaccines showed promising results. The current article briefly covers the prophylactic vaccines and explains different immunotherapy strategies that have been used to treat leishmaniasis. This paper does not include experimental vaccines and only lays emphasis on human trials and those vaccines which reached human trials. Immunotherapy is currently used to successfully treat several disorders; Low cost, limited side effects and no possibility to develop resistance make immunotherapy a valuable choice especially for infectious disease with chemotherapy problems. Efforts are needed to explore the immunological surrogate marker(s) of cure and protection in leishmaniasis and overcome the difficulties in standardization of crude Leishmania vaccines. One of the reasons for anti-leishmaniasis vaccine failure is lack of an appropriate adjuvant. So far, not enough attention has been paid to develop vaccines for immunotherapy of leishmaniasis.

Vaccines for leishmaniasis: from proteome to vaccine candidates.

PubMed

Schroeder, Juliane; Aebischer, Toni

2011-01-01

Leishmania spp. cause a wide spectrum of tropical diseases which are threatening an estimated 350 million people around the globe. While in most cases non-fatal, the disease is associated with high morbidity, social stigmata and poverty. However, the most severe form visceral leishmaniasis can be fatal if left untreated. Chemotherapeutics are available but show high toxicity, costs and are prone to resistance development due to prolonged treatment periods. Healing is associated with a life-long resistance to re-infection and this argues for the feasibility of vaccination. However, despite much effort, no such vaccine has become available yet. Here, the status of vaccine development in this field is briefly summarized before the focus is set on the promise of reverse vaccinology for anti-Leishmania vaccine development in the post-genomic era. We report on our own experience with this approach using an instructive example of successful candidate vaccine antigen identification.

Detection of Leishmania RNA Virus in Leishmania Parasites

PubMed Central

Desponds, Chantal; Kuhlmann, F. Matthew; Robinson, John; Hartley, Mary-Anne; Prevel, Florence; Castiglioni, Patrik; Pratlong, Francine; Bastien, Patrick; Müller, Norbert; Parmentier, Laurent; Saravia, Nancy Gore; Beverley, Stephen M.; Fasel, Nicolas

2013-01-01

Background Patients suffering from cutaneous leishmaniasis (CL) caused by New World Leishmania (Viannia) species are at high risk of developing mucosal (ML) or disseminated cutaneous leishmaniasis (DCL). After the formation of a primary skin lesion at the site of the bite by a Leishmania-infected sand fly, the infection can disseminate to form secondary lesions. This metastatic phenotype causes significant morbidity and is often associated with a hyper-inflammatory immune response leading to the destruction of nasopharyngeal tissues in ML, and appearance of nodules or numerous ulcerated skin lesions in DCL. Recently, we connected this aggressive phenotype to the presence of Leishmania RNA virus (LRV) in strains of L. guyanensis, showing that LRV is responsible for elevated parasitaemia, destructive hyper-inflammation and an overall exacerbation of the disease. Further studies of this relationship and the distribution of LRVs in other Leishmania strains and species would benefit from improved methods of viral detection and quantitation, especially ones not dependent on prior knowledge of the viral sequence as LRVs show significant evolutionary divergence. Methodology/Principal Findings This study reports various techniques, among which, the use of an anti-dsRNA monoclonal antibody (J2) stands out for its specific and quantitative recognition of dsRNA in a sequence-independent fashion. Applications of J2 include immunofluorescence, ELISA and dot blot: techniques complementing an arsenal of other detection tools, such as nucleic acid purification and quantitative real-time-PCR. We evaluate each method as well as demonstrate a successful LRV detection by the J2 antibody in several parasite strains, a freshly isolated patient sample and lesion biopsies of infected mice. Conclusions/Significance We propose that refinements of these methods could be transferred to the field for use as a diagnostic tool in detecting the presence of LRV, and potentially assessing the LRV

Mechanisms of immunity to Leishmania major infection in mice: the contribution of DNA vaccines coding for two novel sets of histones (H2A-H2B or H3-H4).

PubMed

Carrión, Javier

2011-09-01

The immune phenotype conferred by two different sets of histone genes (H2A-H2B or H3-H4) was assessed. BALB/c mice vaccinated with pcDNA3H2AH2B succumbed to progressive cutaneous leishmaniosis (CL), whereas vaccination with pcDNA3H3H4 resulted in partial resistance to Leishmania major challenge associated with the development of mixed T helper 1 (Th1)/Th2-type response and a reduction in parasite-specific Treg cells number at the site of infection. Therefore, the presence of histones H3 and H4 may be considered essential in the development of vaccine strategies against CL based on the Leishmania histones. Copyright © 2011 Elsevier Ltd. All rights reserved.

Status of vaccine research and development of vaccines for leishmaniasis.

PubMed

Gillespie, Portia M; Beaumier, Coreen M; Strych, Ulrich; Hayward, Tara; Hotez, Peter J; Bottazzi, Maria Elena

2016-06-03

A number of leishmaniasis vaccine candidates are at various stages of pre-clinical and clinical development. Leishmaniasis is a vector-borne neglected tropical disease (NTD) caused by a protozoan parasite of the genus Leishmania and transmitted to humans by the bite of a sand fly. Visceral leishmaniasis (VL, kala-azar) is a high mortality NTD found mostly in South Asia and East Africa, while cutaneous leishmaniasis (CL) is a disfiguring NTD highly endemic in the Middle East, Central Asia, North Africa, and the Americas. Estimates attribute 50,000 annual deaths and 3.3 million disability-adjusted life years to leishmaniasis. There are only a few approved drug treatments, no prophylactic drug and no vaccine. Ideally, an effective vaccine against leishmaniasis will elicit long-lasting immunity and protect broadly against VL and CL. Vaccines such as Leish-F1, F2 and F3, developed at IDRI and designed based on selected Leishmania antigen epitopes, have been in clinical trials. Other groups, including the Sabin Vaccine Institute in collaboration with the National Institutes of Health are investigating recombinant Leishmania antigens in combination with selected sand fly salivary gland antigens in order to augment host immunity. To date, both VL and CL vaccines have been shown to be cost-effective in economic modeling studies. Copyright © 2016 World Health Organization. Published by Elsevier Ltd.. All rights reserved.

Identifying vaccine targets for anti-leishmanial vaccine development

PubMed Central

Sundar, Shyam; Singh, Bhawana

2014-01-01

Leishmaniasis is a neglected tropical disease spread by an arthropod vector. It remains a significant health problem with an incidence of 0.2–0.4 million VL and 0.7–1.2 million CL cases each year. There are limitations associated with the current therapeutic regimens for leishmaniasis and the fact that after recovery from infection the host becomes immune to subsequent infection therefore, these factors forces the feasibility of a vaccine for leishmaniasis. Publication of the genome sequence of Leishmania has paved a new way to understand the pathogenesis and host immunological status therefore providing a deep insight in the field of vaccine research. This review is an effort to study the antigenic targets in Leishmania to develop anti-leishmanial vaccine. PMID:24606556

Aberrant IL-4 production by SOCS3-over-expressing T cells during infection with Leishmania major exacerbates disease manifestations.

PubMed

Nakaya, Mako; Hamano, Shinjiro; Kawasumi, Miyuri; Yoshida, Hiroki; Yoshimura, Akihiko; Kobayashi, Takashi

2011-03-01

Suppressor of cytokine signaling (SOCS) 3 is a major negative feedback regulator of signal transducer and activator of transcription 3-activating cytokines. Studies using T-cell-specific SOCS3-deficient mice indicate that the absence of SOCS3 in T cells results in exacerbation of disease progression after infection by Leishmania major due to skewing of the T(h)3 cell phenotype accompanied by hyper-production of IL-10 and transforming growth factor β (TGF-β). Here we show that transgenic mice over-expressing the SOCS3 gene in T cells (Lck-SOCS3 Tg mice) are also susceptible to infection by L. major. Forced expression of SOCS3 in T cells did not affect the production of the anti-inflammatory cytokines IL-10 and TGF-β or that of the protective T(h)1 type cytokine IFN-γ, which is required for parasite clearance. CD4(+) T cells isolated from infected-Lck-SOCS3 Tg mice produced much higher levels of IL-4 when they were re-stimulated with L. major antigen in vitro. Exacerbation of disease progression in Lck-SOCS3 Tg mice was completely reversed by administration of a neutralizing antibody against IL-4. These data suggest that tight regulation of SOCS3 expression in T(h) cells is crucial for disease control during infection by L. major.

SIR2-deficient Leishmania infantum induces a defined IFN-gamma/IL-10 pattern that correlates with protection.

PubMed

Silvestre, Ricardo; Cordeiro-Da-Silva, Anabela; Santarém, Nuno; Vergnes, Baptiste; Sereno, Denis; Ouaissi, Ali

2007-09-01

The ability to manipulate the Leishmania genome to create genetically modified parasites by introducing or eliminating genes is considered a powerful alternative for developing a new generation vaccine against leishmaniasis. Previously, we showed that the deletion of one allele of the Leishmania infantum silent information regulatory 2 (LiSIR2) locus was sufficient to dramatically affect amastigote axenic proliferation. Furthermore, LiSIR2 single knockout (LiSIR2(+/-)) amastigotes were unable to replicate in vitro inside macrophages. Because this L. infantum mutant persisted in BALB/c mice for up to 6 wk but failed to establish an infection, we tested its ability to provide protection toward a virulent L. infantum challenge. Strikingly, vaccination with a single i.p. injection of LiSIR2(+/-) single knockout elicits complete protection. Thus, vaccinated BALB/c mice showed a reversal of T cell anergy with specific anti-Leishmania cytotoxic activity and high levels of NO production. Moreover, vaccinated mice simultaneously generated specific anti-Leishmania IgG Ab subclasses suggestive of both type 1 and type 2 responses. A strong correlation was found between the elimination of the parasites and an increased Leishmania-specific IFN-gamma/IL-10 ratio. Therefore, we propose that the polarization to a high IFN-gamma/low IL-10 ratio after challenge is a clear indicator of vaccine success. Furthermore these mutants, which presented attenuated virulence, represent a good model to understand the correlatives of protection in visceral leishmaniasis.

Skin-resident memory CD4+ T cells enhance protection against Leishmania major infection.

PubMed

Glennie, Nelson D; Yeramilli, Venkata A; Beiting, Daniel P; Volk, Susan W; Weaver, Casey T; Scott, Phillip

2015-08-24

Leishmaniasis causes a significant disease burden worldwide. Although Leishmania-infected patients become refractory to reinfection after disease resolution, effective immune protection has not yet been achieved by human vaccines. Although circulating Leishmania-specific T cells are known to play a critical role in immunity, the role of memory T cells present in peripheral tissues has not been explored. Here, we identify a population of skin-resident Leishmania-specific memory CD4+ T cells. These cells produce IFN-γ and remain resident in the skin when transplanted by skin graft onto naive mice. They function to recruit circulating T cells to the skin in a CXCR3-dependent manner, resulting in better control of the parasites. Our findings are the first to demonstrate that CD4+ TRM cells form in response to a parasitic infection, and indicate that optimal protective immunity to Leishmania, and thus the success of a vaccine, may depend on generating both circulating and skin-resident memory T cells. © 2015 Glennie et al.

Skin-resident memory CD4+ T cells enhance protection against Leishmania major infection

PubMed Central

Glennie, Nelson D.; Yeramilli, Venkata A.; Beiting, Daniel P.; Volk, Susan W.; Weaver, Casey T.

2015-01-01

Leishmaniasis causes a significant disease burden worldwide. Although Leishmania-infected patients become refractory to reinfection after disease resolution, effective immune protection has not yet been achieved by human vaccines. Although circulating Leishmania-specific T cells are known to play a critical role in immunity, the role of memory T cells present in peripheral tissues has not been explored. Here, we identify a population of skin-resident Leishmania-specific memory CD4+ T cells. These cells produce IFN-γ and remain resident in the skin when transplanted by skin graft onto naive mice. They function to recruit circulating T cells to the skin in a CXCR3-dependent manner, resulting in better control of the parasites. Our findings are the first to demonstrate that CD4+ TRM cells form in response to a parasitic infection, and indicate that optimal protective immunity to Leishmania, and thus the success of a vaccine, may depend on generating both circulating and skin-resident memory T cells. PMID:26216123

Computational prediction of protein-protein interactions in Leishmania predicted proteomes.

PubMed

Rezende, Antonio M; Folador, Edson L; Resende, Daniela de M; Ruiz, Jeronimo C

2012-01-01

The Trypanosomatids parasites Leishmania braziliensis, Leishmania major and Leishmania infantum are important human pathogens. Despite of years of study and genome availability, effective vaccine has not been developed yet, and the chemotherapy is highly toxic. Therefore, it is clear just interdisciplinary integrated studies will have success in trying to search new targets for developing of vaccines and drugs. An essential part of this rationale is related to protein-protein interaction network (PPI) study which can provide a better understanding of complex protein interactions in biological system. Thus, we modeled PPIs for Trypanosomatids through computational methods using sequence comparison against public database of protein or domain interaction for interaction prediction (Interolog Mapping) and developed a dedicated combined system score to address the predictions robustness. The confidence evaluation of network prediction approach was addressed using gold standard positive and negative datasets and the AUC value obtained was 0.94. As result, 39,420, 43,531 and 45,235 interactions were predicted for L. braziliensis, L. major and L. infantum respectively. For each predicted network the top 20 proteins were ranked by MCC topological index. In addition, information related with immunological potential, degree of protein sequence conservation among orthologs and degree of identity compared to proteins of potential parasite hosts was integrated. This information integration provides a better understanding and usefulness of the predicted networks that can be valuable to select new potential biological targets for drug and vaccine development. Network modularity which is a key when one is interested in destabilizing the PPIs for drug or vaccine purposes along with multiple alignments of the predicted PPIs were performed revealing patterns associated with protein turnover. In addition, around 50% of hypothetical protein present in the networks received some degree

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

PubMed Central

Bahi-Jaber, Narges; Petitdidier, Elodie; Markikou-Ouni, Wafa; Aoun, Karim; Moreno, Javier; Carrillo, Eugenia; Salotra, Poonam; Kaushal, Himanshu; Negi, Narender Singh; Arevalo, Jorge; Falconi-Agapito, Francesca; Privat, Angela; Cruz, Maria; Pagniez, Julie; Papierok, Gérard-Marie; Rhouma, Faten Bel Haj; Torres, Pilar; Lemesre, Jean-Loup; Chenik, Mehdi; Meddeb-Garnaoui, Amel

2014-01-01

PSA (Promastigote Surface Antigen) belongs to a family of membrane-bound and secreted proteins present in several Leishmania (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 a L. 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 were subdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantum infection) or non immune/naive individuals (HLR: Healthy Low Responders), depending on whether they produce high or low levels of IFN-γ in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detected in sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-γ, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-α in more. No significant cytokine response was observed in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increase in CD4+ T cells producing IFN-γ after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between the percentage of IFN-γ-producing CD4+ T cells and the released IFN-γ. We showed that the LaPSA-38S protein was able to induce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infection indicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacity of Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infection. PMID:24786587

Immunoinformatics Features Linked to Leishmania Vaccine Development: Data Integration of Experimental and In Silico Studies

PubMed Central

Brito, Rory C. F.; Guimarães, Frederico G.; Velloso, João P. L.; Corrêa-Oliveira, Rodrigo; Ruiz, Jeronimo C.; Reis, Alexandre B.; Resende, Daniela M.

2017-01-01

Leishmaniasis is a wide-spectrum disease caused by parasites from Leishmania genus. There is no human vaccine available and it is considered by many studies as apotential effective tool for disease control. To discover novel antigens, computational programs have been used in reverse vaccinology strategies. In this work, we developed a validation antigen approach that integrates prediction of B and T cell epitopes, analysis of Protein-Protein Interaction (PPI) networks and metabolic pathways. We selected twenty candidate proteins from Leishmania tested in murine model, with experimental outcome published in the literature. The predictions for CD4+ and CD8+ T cell epitopes were correlated with protection in experimental outcomes. We also mapped immunogenic proteins on PPI networks in order to find Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways associated with them. Our results suggest that non-protective antigens have lowest frequency of predicted T CD4+ and T CD8+ epitopes, compared with protective ones. T CD4+ and T CD8+ cells are more related to leishmaniasis protection in experimental outcomes than B cell predicted epitopes. Considering KEGG analysis, the proteins considered protective are connected to nodes with few pathways, including those associated with ribosome biosynthesis and purine metabolism. PMID:28208616

Immunoinformatics Features Linked to Leishmania Vaccine Development: Data Integration of Experimental and In Silico Studies.

PubMed

Brito, Rory C F; Guimarães, Frederico G; Velloso, João P L; Corrêa-Oliveira, Rodrigo; Ruiz, Jeronimo C; Reis, Alexandre B; Resende, Daniela M

2017-02-10

Leishmaniasis is a wide-spectrum disease caused by parasites from Leishmania genus. There is no human vaccine available and it is considered by many studies as apotential effective tool for disease control. To discover novel antigens, computational programs have been used in reverse vaccinology strategies. In this work, we developed a validation antigen approach that integrates prediction of B and T cell epitopes, analysis of Protein-Protein Interaction (PPI) networks and metabolic pathways. We selected twenty candidate proteins from Leishmania tested in murine model, with experimental outcome published in the literature. The predictions for CD4⁺ and CD8⁺ T cell epitopes were correlated with protection in experimental outcomes. We also mapped immunogenic proteins on PPI networks in order to find Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways associated with them. Our results suggest that non-protective antigens have lowest frequency of predicted T CD4⁺ and T CD8⁺ epitopes, compared with protective ones. T CD4⁺ and T CD8⁺ cells are more related to leishmaniasis protection in experimental outcomes than B cell predicted epitopes. Considering KEGG analysis, the proteins considered protective are connected to nodes with few pathways, including those associated with ribosome biosynthesis and purine metabolism.

Prediction and analysis of promiscuous T cell-epitopes derived from the vaccine candidate antigens of Leishmania donovani binding to MHC class-II alleles using in silico approach.

PubMed

Kashyap, Manju; Jaiswal, Varun; Farooq, Umar

2017-09-01

Visceral leishmaniasis is a dreadful infectious disease and caused by the intracellular protozoan parasites, Leishmania donovani and Leishmania infantum. Despite extensive efforts for developing effective prophylactic vaccine, still no vaccine is available against leishmaniasis. However, advancement in immunoinformatics methods generated new dimension in peptide based vaccine development. The present study was aimed to identify T-cell epitopes from the vaccine candidate antigens like Lipophosphogylcan-3(LPG-3) and Nucleoside hydrolase (NH) from the L. donovani using in silico methods. Available best tools were used for the identification of promiscuous peptides for MHC class-II alleles. A total of 34 promiscuous peptides from LPG-3, 3 from NH were identified on the basis of their 100% binding affinity towards all six HLA alleles, taken in this study. These peptides were further checked computationally to know their IFN-γ and IL4 inducing potential and nine peptides were identified. Peptide binding interactions with predominant HLA alleles were done by docking. Out of nine docked promiscuous peptides, only two peptides (QESRILRVIKKKLVR, RILRVIKKKLVRKTL), from LPG-3 and one peptide (FDKFWCLVIDALKRI) from NH showed lowest binding energy with all six alleles. These promiscuous T-cell epitopes were predicted on the basis of their antigenicity, hydrophobicity, potential immune response and docking scores. The immunogenicity of predicted promiscuous peptides might be used for subunit vaccine development with immune-modulating adjuvants. Copyright © 2017 Elsevier B.V. All rights reserved.

Leishmania exosomes and other virulence factors: Impact on innate immune response and macrophage functions.

PubMed

Atayde, Vanessa Diniz; Hassani, Kasra; da Silva Lira Filho, Alonso; Borges, Andrezza Raposo; Adhikari, Anupam; Martel, Caroline; Olivier, Martin

2016-11-01

Leishmania parasites are the causative agents of the leishmaniases, a collection of vector-borne diseases that range from simple cutaneous to fatal visceral forms. Employing potent immune modulation mechanisms, Leishmania is able to render the host macrophage inactive and persist inside its phagolysosome. In the last few years, the role of exosomes in Leishmania-host interactions has been increasingly investigated. For instance, it was reported that Leishmania exosome release is augmented following temperature shift, a condition mimicking parasite's entry into its mammalian host. Leishmania exosomes were found to strongly affect macrophage cell signaling and functions, similarly to whole parasites. Importantly, these vesicles were shown to be pro-inflammatory, capable to recruit neutrophils at their inoculation site exacerbating the pathology. In this review, we provide the most recent insights on the role of exosomes and other virulence factors, especially the surface protease GP63, in Leishmania-host interactions, deepening our knowledge on leishmaniasis and paving the way for the development of new therapeutics. Copyright © 2016 Elsevier Inc. All rights reserved.

Leishmania infantum FML pulsed-dendritic cells induce a protective immune response in murine visceral leishmaniasis.

PubMed

Foroughi-Parvar, Faeze; Hatam, Gholam-Reza; Sarkari, Bahador; Kamali-Sarvestani, Eskandar

2015-01-01

To investigate the efficacy of FML loaded dendritic cells (DCs) in protection against visceral leishmaniasis. Mice were immunized with FML- or soluble Leishmania antigen-loaded DCs as well as FML or soluble Leishmania antigen in saponin and challenged with parasite. The levels of cytokines before and after challenge were detected by ELISA. Parasite burden (total Leishman-Donovan unit) was determined after parasite challenge. FML-saponin induced the highest IFN-γ/IL-4 ratio among vaccinated groups, though this ratio was higher in FML-loaded DCs group subsequent to challenge with Leishmania infantum. Moreover, the greatest reduction in parasite number was detected in mice vaccinated with FML-loaded DCs compared with phosphate-buffered saline-treated mice (p = 0.002). FML-loaded DCs are one of the promising tools for protection against murine visceral leishmaniasis.

New insights into the genetic diversity of Leishmania RNA Virus 1 and its species-specific relationship with Leishmania parasites.

PubMed

Cantanhêde, Lilian Motta; Fernandes, Flavia Gonçalves; Ferreira, Gabriel Eduardo Melim; Porrozzi, Renato; Ferreira, Ricardo de Godoi Mattos; Cupolillo, Elisa

2018-01-01

Cutaneous leishmaniasis is a neglected parasitic disease that manifests in infected individuals under different phenotypes, with a range of factors contributing to its broad clinical spectrum. One factor, Leishmania RNA Virus 1 (LRV1), has been described as an endosymbiont present in different species of Leishmania. LRV1 significantly worsens the lesion, exacerbating the immune response in both experimentally infected animals and infected individuals. Little is known about the composition and genetic diversity of these viruses. Here, we investigated the relationship between the genetic composition of LRV1 detected in strains of Leishmania (Viannia) braziliensis and L. (V.) guyanensis and the interaction between the endosymbiont and the parasitic species, analyzing an approximately 850 base pair region of the viral genome. We also included one LRV1 sequence detected in L. (V.) shawi, representing the first report of LRV1 in a species other than L. braziliensis and L. guyanensis. The results illustrate the genetic diversity of the LRV1 strains analyzed here, with smaller divergences detected among viral sequences from the same parasite species. Phylogenetic analyses showed that the LRV1 sequences are grouped according to the parasite species and possibly according to the population of the parasite in which the virus was detected, corroborating the hypothesis of joint evolution of the viruses with the speciation of Leishmania parasites.

The development and clinical evaluation of second-generation leishmaniasis vaccines.

PubMed

Duthie, Malcolm S; Raman, Vanitha S; Piazza, Franco M; Reed, Steven G

2012-01-05

Infection with Leishmania parasites results in a range of clinical manifestations and outcomes. Control of Leishmania parasite transmission is extremely difficult due to the large number of vectors and potential reservoirs, and none of the current treatments are ideal. Vaccination could be an effective strategy to provide sustained control. In this review, the current global situation with regard to leishmaniasis, the immunology of Leishmania infection and various efforts to identify second generation vaccine candidates are briefly discussed. The variety of clinical trials conducted using the only current second generation vaccine approved for clinical use, LEISH-F1+MPL-SE, are described. Given that epidemiological evidence suggests that reducing the canine reservoir also positively impacts human incidence, efforts at providing a vaccine for leishmaniasis in dogs are highlighted. Finally, potential refinements and surrogate markers that could expedite the introduction of a vaccine that can limit the severity and incidence of leishmaniasis are discussed. Copyright © 2011 Elsevier Ltd. All rights reserved.

The Development and Clinical Evaluation of Second-Generation Leishmaniasis Vaccines

PubMed Central

Duthie, Malcolm S.; Raman, Vanitha S.; Piazza, Franco M.; Reed, Steven G.

2011-01-01

Infection with Leishmania parasites results in a range of clinical manifestations and outcomes. Control of Leishmania parasite transmission is extremely difficult due to the large number of vectors and potential reservoirs, and none of the current treatments are ideal. Vaccination could be an effective strategy to provide sustained control. In this review, the current global situation with regard to leishmaniasis, the immunology of Leishmania infection and various efforts to identify second generation vaccine candidates are briefly discussed. The variety of clinical trials conducted using the only current second generation vaccine approved for clinical use, LEISH-F1 + MPL-SE, are described. Given that epidemiological evidence suggests that reducing the canine reservoir also positively impacts human incidence, efforts at providing a vaccine for leishmaniasis in dogs are highlighted. Finally, potential refinements and surrogate markers that could expedite the introduction of a vaccine that can limit the severity and incidence of leishmaniasis are discussed. PMID:22085553

Comparative Analysis of Cellular Immune Responses in Treated Leishmania Patients and Hamsters against Recombinant Th1 Stimulatory Proteins of Leishmania donovani

PubMed Central

Joshi, Sumit; Yadav, Narendra K.; Rawat, Keerti; Tripathi, Chandra Dev P.; Jaiswal, Anil K.; Khare, Prashant; Tandon, Rati; Baharia, Rajendra K.; Das, Sanchita; Gupta, Reema; Kushawaha, Pramod K.; Sundar, Shyam; Sahasrabuddhe, Amogh A.; Dube, Anuradha

2016-01-01

Our prior studies demonstrated that cellular response of T helper 1 (Th1) type was generated by a soluble antigenic fraction (ranging from 89.9 to 97.1 kDa) of Leishmania donovani promastigote, in treated Leishmania patients as well as hamsters and showed significant prophylactic potential against experimental visceral leishmaniasis (VL). Eighteen Th1 stimulatory proteins were identified through proteomic analysis of this subfraction, out of which 15 were developed as recombinant proteins. In the present work, we have evaluated these 15 recombinant proteins simultaneously for their comparative cellular responses in treated Leishmania patients and hamsters. Six proteins viz. elongation factor-2, enolase, aldolase, triose phosphate isomerase, protein disulfide isomerase, and p45 emerged as most immunogenic as they produced a significant lymphoproliferative response, nitric oxide generation and Th1 cytokine response in PBMCs and lymphocytes of treated Leishmania patients and hamsters respectively. The results suggested that these proteins may be exploited for developing a successful poly-protein and/or poly-epitope vaccine against VL. PMID:27047452

Leishmania Hijacks Myeloid Cells for Immune Escape

PubMed Central

Martínez-López, María; Soto, Manuel; Iborra, Salvador; Sancho, David

2018-01-01

Protozoan parasites of the Leishmania genus are the causative agents of leishmaniasis, a group of neglected tropical diseases whose clinical manifestations vary depending on the infectious Leishmania species but also on host factors. Recognition of the parasite by host myeloid immune cells is a key to trigger an effective Leishmania-specific immunity. However, the parasite is able to persist in host myeloid cells by evading, delaying and manipulating host immunity in order to escape host resistance and ensure its transmission. Neutrophils are first in infiltrating infection sites and could act either favoring or protecting against infection, depending on factors such as the genetic background of the host or the parasite species. Macrophages are the main host cells where the parasites grow and divide. However, macrophages are also the main effector population involved in parasite clearance. Parasite elimination by macrophages requires the priming and development of an effector Th1 adaptive immunity driven by specific subtypes of dendritic cells. Herein, we will provide a comprehensive outline of how myeloid cells regulate innate and adaptive immunity against Leishmania, and the mechanisms used by the parasites to promote their evasion and sabotage. Understanding the interactions between Leishmania and the host myeloid cells may lead to the development of new therapeutic approaches and improved vaccination to leishmaniases, an important worldwide health problem in which current therapeutic or preventive approaches are limited. PMID:29867798

Mimotope-Based Vaccines of Leishmania infantum Antigens and Their Protective Efficacy against Visceral Leishmaniasis

PubMed Central

Costa, Lourena Emanuele; Goulart, Luiz Ricardo; Pereira, Nathália Cristina de Jesus; Lima, Mayara Ingrid Sousa; Duarte, Mariana Costa; Martins, Vivian Tamietti; Lage, Paula Sousa; Menezes-Souza, Daniel; Ribeiro, Tatiana Gomes; Melo, Maria Norma; Fernandes, Ana Paula; Soto, Manuel; Tavares, Carlos Alberto Pereira; Chávez-Fumagalli, Miguel Angel; Coelho, Eduardo Antonio Ferraz

2014-01-01

Background The development of cost-effective prophylactic strategies to prevent leishmaniasis has become a high-priority. The present study has used the phage display technology to identify new immunogens, which were evaluated as vaccines in the murine model of visceral leishmaniasis (VL). Epitope-based immunogens, represented by phage-fused peptides that mimic Leishmania infantum antigens, were selected according to their affinity to antibodies from asymptomatic and symptomatic VL dogs' sera. Methodology/Main Findings Twenty phage clones were selected after three selection cycles, and were evaluated by means of in vitro assays of the immune stimulation of spleen cells derived from naive and chronically infected with L. infantum BALB/c mice. Clones that were able to induce specific Th1 immune response, represented by high levels of IFN-γ and low levels of IL-4 were selected, and based on their selectivity and specificity, two clones, namely B10 and C01, were further employed in the vaccination protocols. BALB/c mice vaccinated with clones plus saponin showed both a high and specific production of IFN-γ, IL-12, and GM-CSF after in vitro stimulation with individual clones or L. infantum extracts. Additionally, these animals, when compared to control groups (saline, saponin, wild-type phage plus saponin, or non-relevant phage clone plus saponin), showed significant reductions in the parasite burden in the liver, spleen, bone marrow, and paws' draining lymph nodes. Protection was associated with an IL-12-dependent production of IFN-γ, mainly by CD8+ T cells, against parasite proteins. These animals also presented decreased parasite-mediated IL-4 and IL-10 responses, and increased levels of parasite-specific IgG2a antibodies. Conclusions/Significance This study describes two phage clones that mimic L. infantum antigens, which were directly used as immunogens in vaccines and presented Th1-type immune responses, and that significantly reduced the parasite burden. This

Mimotope-based vaccines of Leishmania infantum antigens and their protective efficacy against visceral leishmaniasis.

PubMed

Costa, Lourena Emanuele; Goulart, Luiz Ricardo; Pereira, Nathália Cristina de Jesus; Lima, Mayara Ingrid Sousa; Duarte, Mariana Costa; Martins, Vivian Tamietti; Lage, Paula Sousa; Menezes-Souza, Daniel; Ribeiro, Tatiana Gomes; Melo, Maria Norma; Fernandes, Ana Paula; Soto, Manuel; Tavares, Carlos Alberto Pereira; Chávez-Fumagalli, Miguel Angel; Coelho, Eduardo Antonio Ferraz

2014-01-01

The development of cost-effective prophylactic strategies to prevent leishmaniasis has become a high-priority. The present study has used the phage display technology to identify new immunogens, which were evaluated as vaccines in the murine model of visceral leishmaniasis (VL). Epitope-based immunogens, represented by phage-fused peptides that mimic Leishmania infantum antigens, were selected according to their affinity to antibodies from asymptomatic and symptomatic VL dogs' sera. Twenty phage clones were selected after three selection cycles, and were evaluated by means of in vitro assays of the immune stimulation of spleen cells derived from naive and chronically infected with L. infantum BALB/c mice. Clones that were able to induce specific Th1 immune response, represented by high levels of IFN-γ and low levels of IL-4 were selected, and based on their selectivity and specificity, two clones, namely B10 and C01, were further employed in the vaccination protocols. BALB/c mice vaccinated with clones plus saponin showed both a high and specific production of IFN-γ, IL-12, and GM-CSF after in vitro stimulation with individual clones or L. infantum extracts. Additionally, these animals, when compared to control groups (saline, saponin, wild-type phage plus saponin, or non-relevant phage clone plus saponin), showed significant reductions in the parasite burden in the liver, spleen, bone marrow, and paws' draining lymph nodes. Protection was associated with an IL-12-dependent production of IFN-γ, mainly by CD8+ T cells, against parasite proteins. These animals also presented decreased parasite-mediated IL-4 and IL-10 responses, and increased levels of parasite-specific IgG2a antibodies. This study describes two phage clones that mimic L. infantum antigens, which were directly used as immunogens in vaccines and presented Th1-type immune responses, and that significantly reduced the parasite burden. This is the first study that describes phage-displayed peptides as

Evaluation of immune responses and analysis of the effect of vaccination of the Leishmania major recombinant ribosomal proteins L3 or L5 in two different murine models of cutaneous leishmaniasis.

PubMed

Ramírez, Laura; Santos, Diego M; Souza, Ana P; Coelho, Eduardo A F; Barral, Aldina; Alonso, Carlos; Escutia, Marta R; Bonay, Pedro; de Oliveira, Camila I; Soto, Manuel

2013-02-18

Four new antigenic proteins located in Leishmania ribosomes have been characterized: S4, S6, L3 and L5. Recombinant versions of the four ribosomal proteins from Leishmania major were recognized by sera from human and canine patients suffering different clinical forms of leishmaniasis. The prophylactic properties of these proteins were first studied in the experimental model of cutaneous leishmaniasis caused by L. major inoculation into BALB/c mice. The administration of two of them, LmL3 or LmL5 combined with CpG-oligodeoxynucleotides (CpG-ODN) was able to protect BALB/c mice against L. major infection. Vaccinated mice showed smaller lesions and parasite burden compared to mice inoculated with vaccine diluent or vaccine adjuvant. Protection was correlated with an antigen-specific increased production of IFN-γ paralleled by a decrease of the antigen-specific IL-10 mediated response in protected mice relative to non-protected controls. Further, it was demonstrated that BALB/c mice vaccinated with recombinant LmL3 or LmL5 plus CpG-ODN were also protected against the development of cutaneous lesions following inoculation of L. braziliensis. Together, data presented here indicate that LmL3 or LmL5 ribosomal proteins combined with Th1 inducing adjuvants, may be relevant components of a vaccine against cutaneous leishmaniasis caused by distinct species. Copyright © 2013 Elsevier Ltd. All rights reserved.

Cytokine Responses to Novel Antigens in a Peri-Urban Population in Brazil Exposed to Leishmania infantum chagasi

PubMed Central

Stober, Carmel B.; Jeronimo, Selma M. B.; Pontes, Nubia N.; Miller, E. Nancy; Blackwell, Jenefer M.

2012-01-01

Visceral leishmaniasis (VL) is fatal if untreated, and there are no vaccines for this disease. High levels of CD4-derived interferon-γ (IFN-γ) in the presence of low levels of interleukin-10 (IL-10) predicts vaccine success. Tumor necrosis factor-α (TNF-α) is also important in this process. We characterized human immune responses in three groups exposed to Leishmania infantum chagasi in Brazil: 1) drug-cured VL patients (recovered VL); 2) asymptomatic persons with positive Leishmania-specific delayed-type hypersensitivity skin reactions (DTH+); and 3) DTH-negative household contacts. Magnitude of DTH correlated with crude Leishmania antigen–driven IFN-γ, TNF-α, and IL-5, but not IL-10. DTH+ persons showed equivalent levels of IFN-γ, but higher levels of IL-10, to tryparedoxin peroxidase and Leishmania homolog of receptor for activated C kinase compared with recovered VL patients. The IFN-γ:IL-10 and TNF-α:IL-10 ratios were higher in recovered VL patients than in DTH+ persons. Seven of 11 novel candidates (R71, L37, N52, L302.06, M18, J41, and M22) elicited cytokine responses (36–71% of responders) in recovered VL patients and DTH+ persons. This result confirmed their putative status as cross-species vaccine/immunotherapeutic candidates. PMID:22826477

Cytokine responses to novel antigens in a peri-urban population in Brazil exposed to Leishmania infantum chagasi.

PubMed

Stober, Carmel B; Jeronimo, Selma M B; Pontes, Nubia N; Miller, E Nancy; Blackwell, Jenefer M

2012-10-01

Visceral leishmaniasis (VL) is fatal if untreated, and there are no vaccines for this disease. High levels of CD4-derived interferon-γ (IFN-γ) in the presence of low levels of interleukin-10 (IL-10) predicts vaccine success. Tumor necrosis factor-α (TNF-α) is also important in this process. We characterized human immune responses in three groups exposed to Leishmania infantum chagasi in Brazil: 1) drug-cured VL patients (recovered VL); 2) asymptomatic persons with positive Leishmania-specific delayed-type hypersensitivity skin reactions (DTH+); and 3) DTH-negative household contacts. Magnitude of DTH correlated with crude Leishmania antigen-driven IFN-γ, TNF-α, and IL-5, but not IL-10. DTH+ persons showed equivalent levels of IFN-γ, but higher levels of IL-10, to tryparedoxin peroxidase and Leishmania homolog of receptor for activated C kinase compared with recovered VL patients. The IFN-γ:IL-10 and TNF-α:IL-10 ratios were higher in recovered VL patients than in DTH+ persons. Seven of 11 novel candidates (R71, L37, N52, L302.06, M18, J41, and M22) elicited cytokine responses (36-71% of responders) in recovered VL patients and DTH+ persons. This result confirmed their putative status as cross-species vaccine/immunotherapeutic candidates.

Efficacy of intranasal LaAg vaccine against Leishmania amazonensis infection in partially resistant C57Bl/6 mice.

PubMed

Pratti, Juliana Elena Silveira; Ramos, Tadeu Diniz; Pereira, Joyce Carvalho; da Fonseca-Martins, Alessandra Marcia; Maciel-Oliveira, Diogo; Oliveira-Silva, Gabriel; de Mello, Mirian França; Chaves, Suzana Passos; Gomes, Daniel Claudio Oliveira; Diaz, Bruno Lourenço; Rossi-Bergmann, Bartira; de Matos Guedes, Herbert Leonel

2016-10-06

We have previously demonstrated that intranasal vaccination of highly susceptible BALB/c mice with whole Leishmania amazonensis antigens (LaAg) leads to protection against murine cutaneous leishmaniasis. Here, we evaluate the response of partially resistant C57BL/6 mice to vaccination as a more representative experimental model of human cutaneous leishmaniasis. C57BL/6 mice from different animal facilities were infected with L. amazonensis (Josefa strain) to establish the profile of infection. Intranasal vaccination was performed before the infection challenge with two doses of 10 μg of LaAg alone or associated with the adjuvant ADDAVAX® by instillation in the nostrils. The lesion progression was measured with a dial caliper and the parasite load by limited dilution assay in the acute and chronic phases of infection. Cytokines were quantified by ELISA in the homogenates of infected footpads. C57BL/6 mice from different animal facilities presented the same L. amazonensis infection profile, displaying a progressive acute phase followed by a controlled chronic phase. Parasites cultured in M199 and Schneider's media were equally infective. Intranasal vaccination with LaAg led to milder acute and chronic phases of the disease. The mechanism of protection was associated with increased production of IFN-gamma in the infected tissue as measured in the acute phase. Association with the ADDAVAX® adjuvant did not improve the efficacy of intranasal LaAg vaccination. Rather, ADDAVAX® reduced vaccination efficacy. This study demonstrates that the efficacy of adjuvant-free intranasal vaccination with LaAg is extendable to the more resistant C57Bl/6 mouse model of infection with L. amazonensis, and is thus not exclusive to the susceptible BALB/c model. These results imply that mucosal immunomodulation by LaAg leads to peripheral protection irrespective of the genetic background of the host.

Immunization with the hybrid protein vaccine, consisting of Leishmania major cysteine proteinases Type I (CPB) and Type II (CPA), partially protects against leishmaniasis.

PubMed

Zadeh-Vakili, Azita; Taheri, Tahere; Taslimi, Yasaman; Doustdari, Fatemeh; Salmanian, Ali-Hatef; Rafati, Sima

2004-05-07

Cysteine proteinases (CPs) are enzymes that belong to the papain superfamily, which are found in a number of organisms from prokaryotes to mammals. On the parasitic protozoan Leishmania, extensive studies have shown that CPs are involved in parasite survival, replication and the onset of disease, and have, therefore, been considered as attractive drugs and/or vaccine targets for the control of leishmaniasis. We have previously shown that cysteine proteinases, Type I (CPB) and Type II (CPA), in Leishmania major (L. major), delivered as recombinant proteins or in plasmid DNA, induce partial protection against infection with the parasite in BALB/c mice. We had shown that the level of protection was greater if a cocktail of cpa and cpb containing DNA constructs was used. Therefore, to reduce the costs associated with the production of these vaccine candidates, a construct was developed, whereby the cpa and cpb genes were fused together to give rise to a single hybrid protein. The genes were fused in tandem where the C-terminal extension (CTE), encoding region of CPB, was located at the 3' of the fused genes, and ultimately expressed in the bacterial expression construct pET-23a. The expression of the CPA/B hybrid protein (60 kDa) was verified using rabbit anti-CPA and anti-CPB antibodies by SDS-PAGE and immunoblotting. The protective potential of the CPA/B hybrid protein against the infection with Leishmania was then assessed in BALB/c mice. The animals were vaccinated with CPA/B, challenged with live L. major promastigotes, and the degree of protection was examined by measuring footpad lesion sizes. It was found that there was a delay in the expansion of lesions size compared to control groups. Furthermore, an immunological analysis of antibody isotypes, before and after infection, showed high levels of IgG2a compared to IgG1 (more than five-fold) in the CPA/B hybrid protein vaccinated group. In addition, a predominant Th1 immune response characterized by in vitro IFN

Insights into the sand fly saliva: Blood-feeding and immune interactions between sand flies, hosts, and Leishmania.

PubMed

Lestinova, Tereza; Rohousova, Iva; Sima, Michal; de Oliveira, Camila I; Volf, Petr

2017-07-01

Leishmaniases are parasitic diseases present worldwide that are transmitted to the vertebrate host by the bite of an infected sand fly during a blood feeding. Phlebotomine sand flies inoculate into the mammalian host Leishmania parasites embedded in promastigote secretory gel (PSG) with saliva, which is composed of a diverse group of molecules with pharmacological and immunomodulatory properties. In this review, we focus on 3 main aspects of sand fly salivary molecules: (1) structure and composition of salivary glands, including the properties of salivary molecules related to hemostasis and blood feeding, (2) immunomodulatory properties of salivary molecules and the diverse impacts of these molecules on leishmaniasis, ranging from disease exacerbation to vaccine development, and (3) use of salivary molecules for field applications, including monitoring host exposure to sand flies and the risk of Leishmania transmission. Studies showed interesting differences between salivary proteins of Phlebotomus and Lutzomyia species, however, no data were ever published on salivary proteins of Sergentomyia species. In the last 15 years, numerous studies have characterized sand fly salivary proteins and, in parallel, have addressed the impact of such molecules on the biology of the host-sand fly-parasite interaction. The results obtained shall pave the way for the development of field-application tools that could contribute to the management of leishmaniasis in endemic areas.

Not All Antigens Are Created Equally: Progress, Challenges, and Lessons Associated with Developing a Vaccine for Leishmaniasis

PubMed Central

Reed, Steven G.

2017-01-01

ABSTRACT From experimental models and the analyses of patients, it is well documented that antigen-specific T cells are critical for protection against Leishmania infection. Effective vaccines require both targeting to the pathogen and an immune stimulant to induce maturation of appropriate immune responses. While a great number of antigens have been examined as vaccine candidates against various Leishmania species, few have advanced to human or canine clinical trials. With emphasis on antigen expression, in this minireview we discuss some of the vaccine platforms that are currently being explored for the development of Leishmania vaccines. It is clear that the vaccine platform of choice can have a significant impact upon the level of protection induced by particular antigens, and we provide and highlight some examples for which the vaccine system used has impacted the protective efficacy imparted. PMID:28515135

Not All Antigens Are Created Equally: Progress, Challenges, and Lessons Associated with Developing a Vaccine for Leishmaniasis.

PubMed

Duthie, Malcolm S; Reed, Steven G

2017-07-01

From experimental models and the analyses of patients, it is well documented that antigen-specific T cells are critical for protection against Leishmania infection. Effective vaccines require both targeting to the pathogen and an immune stimulant to induce maturation of appropriate immune responses. While a great number of antigens have been examined as vaccine candidates against various Leishmania species, few have advanced to human or canine clinical trials. With emphasis on antigen expression, in this minireview we discuss some of the vaccine platforms that are currently being explored for the development of Leishmania vaccines. It is clear that the vaccine platform of choice can have a significant impact upon the level of protection induced by particular antigens, and we provide and highlight some examples for which the vaccine system used has impacted the protective efficacy imparted. Copyright © 2017 American Society for Microbiology.

Unraveling the genetic diversity and phylogeny of Leishmania RNA virus 1 strains of infected Leishmania isolates circulating in French Guiana.

PubMed

Tirera, Sourakhata; Ginouves, Marine; Donato, Damien; Caballero, Ignacio S; Bouchier, Christiane; Lavergne, Anne; Bourreau, Eliane; Mosnier, Emilie; Vantilcke, Vincent; Couppié, Pierre; Prevot, Ghislaine; Lacoste, Vincent

2017-07-01

Leishmania RNA virus type 1 (LRV1) is an endosymbiont of some Leishmania (Vianna) species in South America. Presence of LRV1 in parasites exacerbates disease severity in animal models and humans, related to a disproportioned innate immune response, and is correlated with drug treatment failures in humans. Although the virus was identified decades ago, its genomic diversity has been overlooked until now. We subjected LRV1 strains from 19 L. (V.) guyanensis and one L. (V.) braziliensis isolates obtained from cutaneous leishmaniasis samples identified throughout French Guiana with next-generation sequencing and de novo sequence assembly. We generated and analyzed 24 unique LRV1 sequences over their full-length coding regions. Multiple alignment of these new sequences revealed variability (0.5%-23.5%) across the entire sequence except for highly conserved motifs within the 5' untranslated region. Phylogenetic analyses showed that viral genomes of L. (V.) guyanensis grouped into five distinct clusters. They further showed a species-dependent clustering between viral genomes of L. (V.) guyanensis and L. (V.) braziliensis, confirming a long-term co-evolutionary history. Noteworthy, we identified cases of multiple LRV1 infections in three of the 20 Leishmania isolates. Here, we present the first-ever estimate of LRV1 genomic diversity that exists in Leishmania (V.) guyanensis parasites. Genetic characterization and phylogenetic analyses of these viruses has shed light on their evolutionary relationships. To our knowledge, this study is also the first to report cases of multiple LRV1 infections in some parasites. Finally, this work has made it possible to develop molecular tools for adequate identification and genotyping of LRV1 strains for diagnostic purposes. Given the suspected worsening role of LRV1 infection in the pathogenesis of human leishmaniasis, these data have a major impact from a clinical viewpoint and for the management of Leishmania-infected patients.

Recovery of antigen-specific T cell responses from dogs infected with Leishmania (L.) infantum by use of vaccine associated TLR-agonist adjuvant.

PubMed

Schaut, Robert G; Grinnage-Pulley, Tara L; Esch, Kevin J; Toepp, Angela J; Duthie, Malcolm S; Howard, Randall F; Reed, Steven G; Petersen, Christine A

2016-10-17

Visceral leishmaniasis (VL), caused by infection with the obligate intracellular protozoan parasite Leishmania infantum, is a fatal disease of dogs and humans. Protection against VL requires a T helper 1 (Th1) skewed CD4 + T response, but despite this knowledge, there are currently no approved-to-market vaccines for humans and only three veterinary-use vaccines globally. As VL progresses from asymptomatic to symptomatic, L. infantum-specific interferon gamma (IFNγ) driven-Th1 responses become dampened and a state of immune exhaustion established. T cell exhaustion and other immunoregulatory processes, starting during asymptomatic disease, are likely to hinder vaccine-induced responses if vaccine is administered to infected, but asymptomatic and seronegative, individuals. In this study we evaluated how immune exhaustion, shown previously by our group to worsen in concert with VL progression, effected the capacity of vaccine candidate antigen/toll-like receptor (TLR) agonist combinations to promote protective CD4 + T cell responses during progressive VL. In conjunction with Th1 responses, we also evaluated concomitant stimulation of immune-balanced IL-10 regulatory cytokine production by these vaccine products in progressive VL canine T cells. Vaccine antigen L111f in combination with TLR agonists significantly recovered CD4 + T cell IFNγ intracellular production in T cells from asymptomatic VL dogs. Vaccine antigen NS with TLR agonists significantly recovered CD4 + T cell production in both endemic control and VL dogs. Combinations of TLR agonists and vaccine antigens overcame L. infantum induced cellular exhaustion, allowing robust Th1 CD4 + T cell responses from symptomatic dogs that previously had dampened responses to antigen alone. Antigen-agonist adjuvants can be utilized to promote more robust vaccine responses from infected hosts in endemic areas where vaccination of asymptomatic, L. infantum-infected animals is likely. Copyright © 2016. Published by

Evaluation of vaccinal effectiveness of preparations containing membrane antigens of Leishmania (L.) amazonensis in experimental cutaneous leishmaniasis model.

PubMed

Ribeiro, João G; Ferreira, Amália S; Macedo, Sharon R A; Rossi, Norton R D L P; da Silva, Mayara C P; Guerra, Rosane N M; de Barros, Neuza B; Nicolete, Roberto

2017-06-01

American tegumentary leishmaniasis (ATL) is considered a neglected disease, for which an effective vaccine or an efficient diagnosis is not yet available and whose chemotherapeutic arsenal is threatened by the emergence of resistance by etiological agents such as Leishmania amazonensis. ATL is endemic in poor countries and has a high incidence in Brazil. Vaccines developed from native parasite fractions have led to the identification of defined antigenic subunits and the development of vaccine adjuvant technology. The purpose of the present study was to develop and compare preparations based on membrane antigens from L. amazonensis, as a biotechnological prototype for the immunoprophylaxis of the disease in a murine experimental model. For this purpose, batches of biodegradable polymeric micro/nanoparticles were produced, characterized and compared with other parasite's antigens in solution. All preparations containing membrane antigens presented low toxicity on murine macrophages. The in vivo evaluation of immunization efficacy was performed against a challenge with L. amazonensis, along with an evaluation of the immune response profile generated in BALB/C mice. The animals were followed for sample processing and quantification of serum-specific cytokines, nitrites and antibodies. The sera of animals immunized with the non-encapsulated antigen formulations showed higher intensities of nitrites and total IgGs. This approach evidenced the importance of the biological studies involving the immune response of the host against the parasite being interconnected and related to the subfractionation of its proteins in the search for more effective vaccine candidates. Copyright © 2017 Elsevier B.V. All rights reserved.

Identification of Tunisian Leishmania spp. by PCR amplification of cysteine proteinase B (cpb) genes and phylogenetic analysis.

PubMed

Chaouch, Melek; Fathallah-Mili, Akila; Driss, Mehdi; Lahmadi, Ramzi; Ayari, Chiraz; Guizani, Ikram; Ben Said, Moncef; Benabderrazak, Souha

2013-03-01

Discrimination of the Old World Leishmania parasites is important for diagnosis and epidemiological studies of leishmaniasis. We have developed PCR assays that allow the discrimination between Leishmania major, Leishmania tropica and Leishmania infantum Tunisian species. The identification was performed by a simple PCR targeting cysteine protease B (cpb) gene copies. These PCR can be a routine molecular biology tools for discrimination of Leishmania spp. from different geographical origins and different clinical forms. Our assays can be an informative source for cpb gene studying concerning drug, diagnostics and vaccine research. The PCR products of the cpb gene and the N-acetylglucosamine-1-phosphate transferase (nagt) Leishmania gene were sequenced and aligned. Phylogenetic trees of Leishmania based cpb and nagt sequences are close in topology and present the classic distribution of Leishmania in the Old World. The phylogenetic analysis has enabled the characterization and identification of different strains, using both multicopy (cpb) and single copy (nagt) genes. Indeed, the cpb phylogenetic analysis allowed us to identify the Tunisian Leishmania killicki species, and a group which gathers the least evolved isolates of the Leishmania donovani complex, that was originated from East Africa. This clustering confirms the African origin for the visceralizing species of the L. donovani complex. Copyright © 2012 Elsevier B.V. All rights reserved.

Using Proteomics to Understand How Leishmania Parasites Survive inside the Host and Establish Infection

PubMed Central

Veras, Patrícia Sampaio Tavares; Bezerra de Menezes, Juliana Perrone

2016-01-01

Leishmania is a protozoan parasite that causes a wide range of different clinical manifestations in mammalian hosts. It is a major public health risk on different continents and represents one of the most important neglected diseases. Due to the high toxicity of the drugs currently used, and in the light of increasing drug resistance, there is a critical need to develop new drugs and vaccines to control Leishmania infection. Over the past few years, proteomics has become an important tool to understand the underlying biology of Leishmania parasites and host interaction. The large-scale study of proteins, both in parasites and within the host in response to infection, can accelerate the discovery of new therapeutic targets. By studying the proteomes of host cells and tissues infected with Leishmania, as well as changes in protein profiles among promastigotes and amastigotes, scientists hope to better understand the biology involved in the parasite survival and the host-parasite interaction. This review demonstrates the feasibility of proteomics as an approach to identify new proteins involved in Leishmania differentiation and intracellular survival. PMID:27548150

Using Proteomics to Understand How Leishmania Parasites Survive inside the Host and Establish Infection.

PubMed

Veras, Patrícia Sampaio Tavares; Bezerra de Menezes, Juliana Perrone

2016-08-19

Leishmania is a protozoan parasite that causes a wide range of different clinical manifestations in mammalian hosts. It is a major public health risk on different continents and represents one of the most important neglected diseases. Due to the high toxicity of the drugs currently used, and in the light of increasing drug resistance, there is a critical need to develop new drugs and vaccines to control Leishmania infection. Over the past few years, proteomics has become an important tool to understand the underlying biology of Leishmania parasites and host interaction. The large-scale study of proteins, both in parasites and within the host in response to infection, can accelerate the discovery of new therapeutic targets. By studying the proteomes of host cells and tissues infected with Leishmania, as well as changes in protein profiles among promastigotes and amastigotes, scientists hope to better understand the biology involved in the parasite survival and the host-parasite interaction. This review demonstrates the feasibility of proteomics as an approach to identify new proteins involved in Leishmania differentiation and intracellular survival.

Nanobiotechnologic approach to a promising vaccine prototype for immunisation against leishmaniasis: a fast and effective method to incorporate GPI-anchored proteins of Leishmania amazonensis into liposomes.

PubMed

Colhone, Marcelle Carolina; Silva-Jardim, Izaltina; Stabeli, Rodrigo Guerino; Ciancaglini, Pietro

2015-01-01

Liposomes are known to be a potent adjuvant for a wide range of antigens, as well as appropriate antigen carriers for antibody generation response in vivo. In addition, liposomes are effective vehicles for peptides and proteins, thus enhancing their immunogenicity. Considering these properties of liposomes and the antigenicity of the Leishmania membrane proteins, we evaluated if liposomes carrying glycosylphosphatidylinositol (GPI)-anchored proteins of Leishmania amazonensis promastigotes could induce protective immunity in BALB/c mice. To assay protective immunity, BALB/c mice were intraperitoneally injected with liposomes, GPI-protein extract (EPSGPI) as well as with the proteoliposomes carrying GPI-proteins. Mice inoculated with EPSGPI and total protein present in constitutive proteoliposomes displayed a post-infection protection of about 70% and 90%, respectively. The liposomes are able to work as adjuvant in the EPSGPI protection. These systems seem to be a promising vaccine prototype for immunisation against leishmaniasis.

HisAK70: progress towards a vaccine against different forms of leishmaniosis.

PubMed

Domínguez-Bernal, Gustavo; Horcajo, Pilar; Orden, José A; Ruiz-Santa-Quiteria, José A; De La Fuente, Ricardo; Ordóñez-Gutiérrez, Lara; Martínez-Rodrigo, Abel; Mas, Alicia; Carrión, Javier

2015-12-09

Leishmania major and Leishmania infantum are among the main species that are responsible for cutaneous leishmaniosis (CL) and visceral leishmaniosis (VL), respectively. The leishmanioses represent the second-largest parasitic killer in the world after malaria. Recently, we succeeded in generating a plasmid DNA (pCMV-HISA70m2A) and demonstrated that immunized mice were protected against L. major challenge. The efficacy of the DNA-vaccine was further enhanced by the inclusion of KMP-11 antigen into the antibiotic-free plasmid pVAX1-asd. Here, we describe the use of a HisAK70 DNA-vaccine encoding seven Leishmania genes (H2A, H2B, H3, H4, A2, KMP11 and HSP70) for vaccination of mice to assess the induction of a resistant phenotype against VL and CL. HisAK70 was successful in vaccinated mice, resulting in a high amount of efficient sterile hepatic granulomas associated with a hepatic parasite burden fully resolved in the VL model; and resulting in 100% inhibition of parasite visceralization in the CL model. The results suggest that immunization with the HisAK70 DNA-vaccine may provide a rapid, suitable, and efficient vaccination strategy to confer cross-protective immunity against VL and CL.

An α-Gal-containing neoglycoprotein-based vaccine partially protects against murine cutaneous leishmaniasis caused by Leishmania major

PubMed Central

Iniguez, Eva; Schocker, Nathaniel S.; Subramaniam, Krishanthi; Portillo, Susana; Montoya, Alba L.; Al-Salem, Waleed S.; Torres, Caresse L.; Rodriguez, Felipe; Moreira, Otacilio C.; Acosta-Serrano, Alvaro; Michael, Katja; Maldonado, Rosa A.

2017-01-01

Background Protozoan parasites from the genus Leishmania cause broad clinical manifestations known as leishmaniases, which affect millions of people worldwide. Cutaneous leishmaniasis (CL), caused by L. major, is one the most common forms of the disease in the Old World. There is no preventive or therapeutic human vaccine available for L. major CL, and existing drug treatments are expensive, have toxic side effects, and resistant parasite strains have been reported. Hence, further therapeutic interventions against the disease are necessary. Terminal, non-reducing, and linear α-galactopyranosyl (α-Gal) epitopes are abundantly found on the plasma membrane glycolipids of L. major known as glycoinositolphospholipids. The absence of these α-Gal epitopes in human cells makes these glycans highly immunogenic and thus potential targets for vaccine development against CL. Methodology/Principal findings Here, we evaluated three neoglycoproteins (NGPs), containing synthetic α-Gal epitopes covalently attached to bovine serum albumin (BSA), as vaccine candidates against L. major, using α1,3-galactosyltransferase-knockout (α1,3GalT-KO) mice. These transgenic mice, similarly to humans, do not express nonreducing, linear α-Gal epitopes in their cells and are, therefore, capable of producing high levels of anti-α-Gal antibodies. We observed that Galα(1,6)Galβ-BSA (NGP5B), but not Galα(1,4)Galβ-BSA (NGP12B) or Galα(1,3)Galα-BSA (NGP17B), was able to significantly reduce the size of footpad lesions by 96% in comparison to control groups. Furthermore, we observed a robust humoral and cellular immune response with production of high levels of protective lytic anti-α-Gal antibodies and induction of Th1 cytokines. Conclusions/Significance We propose that NGP5B is an attractive candidate for the study of potential synthetic α-Gal-neoglycoprotein-based vaccines against L. major infection. PMID:29069089

An α-Gal-containing neoglycoprotein-based vaccine partially protects against murine cutaneous leishmaniasis caused by Leishmania major.

PubMed

Iniguez, Eva; Schocker, Nathaniel S; Subramaniam, Krishanthi; Portillo, Susana; Montoya, Alba L; Al-Salem, Waleed S; Torres, Caresse L; Rodriguez, Felipe; Moreira, Otacilio C; Acosta-Serrano, Alvaro; Michael, Katja; Almeida, Igor C; Maldonado, Rosa A

2017-10-01

Protozoan parasites from the genus Leishmania cause broad clinical manifestations known as leishmaniases, which affect millions of people worldwide. Cutaneous leishmaniasis (CL), caused by L. major, is one the most common forms of the disease in the Old World. There is no preventive or therapeutic human vaccine available for L. major CL, and existing drug treatments are expensive, have toxic side effects, and resistant parasite strains have been reported. Hence, further therapeutic interventions against the disease are necessary. Terminal, non-reducing, and linear α-galactopyranosyl (α-Gal) epitopes are abundantly found on the plasma membrane glycolipids of L. major known as glycoinositolphospholipids. The absence of these α-Gal epitopes in human cells makes these glycans highly immunogenic and thus potential targets for vaccine development against CL. Here, we evaluated three neoglycoproteins (NGPs), containing synthetic α-Gal epitopes covalently attached to bovine serum albumin (BSA), as vaccine candidates against L. major, using α1,3-galactosyltransferase-knockout (α1,3GalT-KO) mice. These transgenic mice, similarly to humans, do not express nonreducing, linear α-Gal epitopes in their cells and are, therefore, capable of producing high levels of anti-α-Gal antibodies. We observed that Galα(1,6)Galβ-BSA (NGP5B), but not Galα(1,4)Galβ-BSA (NGP12B) or Galα(1,3)Galα-BSA (NGP17B), was able to significantly reduce the size of footpad lesions by 96% in comparison to control groups. Furthermore, we observed a robust humoral and cellular immune response with production of high levels of protective lytic anti-α-Gal antibodies and induction of Th1 cytokines. We propose that NGP5B is an attractive candidate for the study of potential synthetic α-Gal-neoglycoprotein-based vaccines against L. major infection.

Randomised vaccine trial of single dose of killed Leishmania major plus BCG against anthroponotic cutaneous leishmaniasis in Bam, Iran.

PubMed

Sharifi, I; FeKri, A R; Aflatonian, M R; Khamesipour, A; Nadim, A; Mousavi, M R; Momeni, A Z; Dowlati, Y; Godal, T; Zicker, F; Smith, P G; Modabber, F

1998-05-23

A vaccine consisting of a single dose of whole-cell autoclave-killed Leishmania major (ALM) mixed with BCG was assessed in comparison with BCG alone against anthroponotic (human to human transmission) cutaneous leishmaniasis in a randomised double-blind trial in Bam, Iran. 3637 schoolchildren, aged 6-15 years, with no history of cutaneous leishmaniasis and no response to a leishmanin skin test, were randomly assigned to receive 1 mg ALM mixed with BCG (n = 1839), or BCG alone (n = 1798). Safety of the vaccine and the incidence of confirmed cases of cutaneous leishmaniasis were followed up for 2 years. Side-effects were those usually associated with BCG vaccination, but tended to persist longer in the ALM + BCG group. After exclusion of four cases occurring within 80 days of vaccination (one in the ALM + BCG group and three in the BCG group), the 2-year incidence of cutaneous leishmaniasis did not differ significantly between vaccine and BCG groups: 2.8% vs 3.3%, respectively (total cases 112). A sex-stratified analysis showed that in boys the vaccine conferred a protective efficacy of 18% and 78% for the first and second years, respectively--a crude 2-year overall protection of 55% (95% CI 19-75%, p < 0.01). In the first 9 months after vaccination, there was a non-significant excess of cases in the ALM + BCG group (25 vs 16), whereas the incidence of cutaneous leishmaniasis thereafter was significantly reduced in the ALM + BCG group (27 vs 44, p < 0.05). A single dose of ALM + BCG was safe and more immunogenic than BCG alone, as measured by leishmanin skin test. The exact reason for the apparent protective effect of the vaccine in boys is unknown, and may be a chance finding. However, since boys are more exposed to the infection, which is indicated by higher disease prevalence in boys in this study population, the preferential protective effect in boys may have resulted from a greater booster effect produced by repeated exposure to infected sandflies. Booster

Pre-clinical antigenicity studies of an innovative multivalent vaccine for human visceral leishmaniasis.

PubMed

Cecílio, Pedro; Pérez-Cabezas, Begoña; Fernández, Laura; Moreno, Javier; Carrillo, Eugenia; Requena, José M; Fichera, Epifanio; Reed, Steven G; Coler, Rhea N; Kamhawi, Shaden; Oliveira, Fabiano; Valenzuela, Jesus G; Gradoni, Luigi; Glueck, Reinhard; Gupta, Gaurav; Cordeiro-da-Silva, Anabela

2017-11-01

The notion that previous infection by Leishmania spp. in endemic areas leads to robust anti-Leishmania immunity, supports vaccination as a potentially effective approach to prevent disease development. Nevertheless, to date there is no vaccine available for human leishmaniasis. We optimized and assessed in vivo the safety and immunogenicity of an innovative vaccine candidate against human visceral leishmaniasis (VL), consisting of Virus-Like Particles (VLP) loaded with three different recombinant proteins (LJL143 from Lutzomyia longipalpis saliva as the vector-derived (VD) component, and KMP11 and LeishF3+, as parasite-derived (PD) antigens) and adjuvanted with GLA-SE, a TLR4 agonist. No apparent adverse reactions were observed during the experimental time-frame, which together with the normal hematological parameters detected seems to point to the safety of the formulation. Furthermore, measurements of antigen-specific cellular and humoral responses, generally higher in immunized versus control groups, confirmed the immunogenicity of the vaccine formulation. Interestingly, the immune responses against the VD protein were reproducibly more robust than those elicited against leishmanial antigens, and were apparently not caused by immunodominance of the VD antigen. Remarkably, priming with the VD protein alone and boosting with the complete vaccine candidate contributed towards an increase of the immune responses to the PD antigens, assessed in the form of increased ex vivo CD4+ and CD8+ T cell proliferation against both the PD antigens and total Leishmania antigen (TLA). Overall, our immunogenicity data indicate that this innovative vaccine formulation represents a promising anti-Leishmania vaccine whose efficacy deserves to be tested in the context of the "natural infection".

Pre-clinical antigenicity studies of an innovative multivalent vaccine for human visceral leishmaniasis

PubMed Central

Cecílio, Pedro; Pérez-Cabezas, Begoña; Fernández, Laura; Moreno, Javier; Carrillo, Eugenia; Requena, José M.; Fichera, Epifanio; Reed, Steven G.; Coler, Rhea N.; Kamhawi, Shaden; Oliveira, Fabiano; Valenzuela, Jesus G.; Gradoni, Luigi; Glueck, Reinhard; Gupta, Gaurav

2017-01-01

The notion that previous infection by Leishmania spp. in endemic areas leads to robust anti-Leishmania immunity, supports vaccination as a potentially effective approach to prevent disease development. Nevertheless, to date there is no vaccine available for human leishmaniasis. We optimized and assessed in vivo the safety and immunogenicity of an innovative vaccine candidate against human visceral leishmaniasis (VL), consisting of Virus-Like Particles (VLP) loaded with three different recombinant proteins (LJL143 from Lutzomyia longipalpis saliva as the vector-derived (VD) component, and KMP11 and LeishF3+, as parasite-derived (PD) antigens) and adjuvanted with GLA-SE, a TLR4 agonist. No apparent adverse reactions were observed during the experimental time-frame, which together with the normal hematological parameters detected seems to point to the safety of the formulation. Furthermore, measurements of antigen-specific cellular and humoral responses, generally higher in immunized versus control groups, confirmed the immunogenicity of the vaccine formulation. Interestingly, the immune responses against the VD protein were reproducibly more robust than those elicited against leishmanial antigens, and were apparently not caused by immunodominance of the VD antigen. Remarkably, priming with the VD protein alone and boosting with the complete vaccine candidate contributed towards an increase of the immune responses to the PD antigens, assessed in the form of increased ex vivo CD4+ and CD8+ T cell proliferation against both the PD antigens and total Leishmania antigen (TLA). Overall, our immunogenicity data indicate that this innovative vaccine formulation represents a promising anti-Leishmania vaccine whose efficacy deserves to be tested in the context of the “natural infection”. PMID:29176865

Efficacy of killed whole-parasite vaccines in the prevention of leishmaniasis: a meta-analysis.

PubMed

Noazin, Sassan; Khamesipour, Ali; Moulton, Lawrence H; Tanner, Marcel; Nasseri, Kiumarss; Modabber, Farrokh; Sharifi, Iraj; Khalil, E A G; Bernal, Ivan Dario Velez; Antunes, Carlos M F; Smith, Peter G

2009-07-30

Despite decades of investigation in countries on three continents, an efficacious vaccine against Leishmania infections has not been developed. Although some indication of protection was observed in some of the controlled trials conducted with "first-generation" whole, inactivated Leishmania parasite vaccines, convincing evidence of protection was lacking. After reviewing all previously published or unpublished randomized, controlled field efficacy clinical trials of prophylactic candidate vaccines, a meta-analysis of qualified trials was conducted to evaluate whether there was some evidence of protection revealed by considering the results of all trials together. The findings indicate that the whole-parasite vaccine candidates tested do not confer significant protection against human leishmaniasis.

Vaccines and vaccination strategies against human cutaneous leishmaniasis.

PubMed

Okwor, Ifeoma; Uzonna, Jude

2009-05-01

One might think that the development of a vaccine against cutaneous leishmaniasis would be relatively straightforward because the type of immune response required for protection is known and natural immunity occurs following recovery from primary infection. However, there is as yet no effective vaccine against the disease in humans. Although vaccination in murine studies has yielded promising results, these vaccines have failed miserably when tested in primates or humans. The reasons behind these failures are unknown and remain a major hurdle for vaccine design and development against cutaneous leishmaniasis. In contrast, recovery from natural, deliberate or experimental infections results in development of long-lasting immunity to re-infection. This so called infection-induced resistance is the strongest anti-Leishmania immunity known. Here, we briefly review the different approaches to vaccination against cutaneous leishmaniasis and argue that vaccines composed of genetically modified (attenuated) parasites, which induce immunity akin to infection-induced resistance, may provide best protection against cutaneous leishmaniasis in humans.

Ability of immunodiagnostic tests to differentiate between dogs naturally infected with Leishmania infantum and Leishmune(®)-vaccinated dogs.

PubMed

Ribeiro, R A N; Teixeira-Neto, R G; Belo, V S; Ferreira, E C; Schallig, H D F H; Silva, E S

2015-06-01

Visceral leishmaniasis (VL) is a serious chronic disease with a lethality rate of up to 10% in humans. In urban areas of Brazil, dogs are the main reservoirs of the etiological agent (Leishmania infantum) of VL, and the Brazilian Ministry of Health recommends the euthanasia of animals that are seropositive in both the immunochromatographic dual path platform rapid test (DPP(®); Bio-Manguinhos) and the enzyme-linked immunosorbent assay (ELISA) with an L. major-like antigen (Bio-Manguinhos). Vaccination is an additional tool in the control of canine VL, but the use of Leishmune(®) (Zoetis Indústria de Produtos Veterinários, São Paulo, SP, Brazil), which contains the fucose mannose ligand (FML) isolated from L. donovani, is not currently recommended by the Brazilian Ministry of Health because vaccinated animals may exhibit positive serology and there are reservations regarding the efficacy of the vaccine. The aims of the present study were: (i) to verify the abilities of the fast agglutination screening test (FAST), the direct agglutination test (DAT), the indirect fluorescent-antibody test (IFAT), the DPP rapid test, and ELISA tests with L. major-like and FML antigens to differentiate between L. infantum-infected and Leishmune(®)-vaccinated dogs, and (ii) to analyze the sensitivities and specificities of the different methods. The reactivities to these tests of Leishmune(®)-vaccinated dogs (n = 71), asymptomatic (n = 20) and symptomatic (n = 20) naturally infected dogs, and unvaccinated healthy control dogs (n = 5) were compared. None of the Leishmune(®)-vaccinated dogs tested seropositive in FAST and DAT, although one dog was reactive to DPP and four dogs to ELISA/L. major-like and IFAT tests. While 69 (97%) of vaccinated dogs reacted to ELISA/FML, only one was seropositive in both ELISA/L. major-like and IFAT tests. Individually, all immunodiagnostic tests presented high specificities and positive likelihood ratios (LR+), and high specificity values were

Unraveling the genetic diversity and phylogeny of Leishmania RNA virus 1 strains of infected Leishmania isolates circulating in French Guiana

PubMed Central

Caballero, Ignacio S.; Bouchier, Christiane; Lavergne, Anne; Bourreau, Eliane; Mosnier, Emilie; Vantilcke, Vincent; Couppié, Pierre; Prevot, Ghislaine

2017-01-01

Introduction Leishmania RNA virus type 1 (LRV1) is an endosymbiont of some Leishmania (Vianna) species in South America. Presence of LRV1 in parasites exacerbates disease severity in animal models and humans, related to a disproportioned innate immune response, and is correlated with drug treatment failures in humans. Although the virus was identified decades ago, its genomic diversity has been overlooked until now. Methodology/Principles findings We subjected LRV1 strains from 19 L. (V.) guyanensis and one L. (V.) braziliensis isolates obtained from cutaneous leishmaniasis samples identified throughout French Guiana with next-generation sequencing and de novo sequence assembly. We generated and analyzed 24 unique LRV1 sequences over their full-length coding regions. Multiple alignment of these new sequences revealed variability (0.5%–23.5%) across the entire sequence except for highly conserved motifs within the 5’ untranslated region. Phylogenetic analyses showed that viral genomes of L. (V.) guyanensis grouped into five distinct clusters. They further showed a species-dependent clustering between viral genomes of L. (V.) guyanensis and L. (V.) braziliensis, confirming a long-term co-evolutionary history. Noteworthy, we identified cases of multiple LRV1 infections in three of the 20 Leishmania isolates. Conclusions/Significance Here, we present the first-ever estimate of LRV1 genomic diversity that exists in Leishmania (V.) guyanensis parasites. Genetic characterization and phylogenetic analyses of these viruses has shed light on their evolutionary relationships. To our knowledge, this study is also the first to report cases of multiple LRV1 infections in some parasites. Finally, this work has made it possible to develop molecular tools for adequate identification and genotyping of LRV1 strains for diagnostic purposes. Given the suspected worsening role of LRV1 infection in the pathogenesis of human leishmaniasis, these data have a major impact from a

Cationic liposomes containing soluble Leishmania antigens (SLA) plus CpG ODNs induce protection against murine model of leishmaniasis.

PubMed

Heravi Shargh, Vahid; Jaafari, Mahmoud Reza; Khamesipour, Ali; Jalali, Seyed Amir; Firouzmand, Hengameh; Abbasi, Azam; Badiee, Ali

2012-07-01

Development of an effective vaccine against leishmaniasis is possible due to the fact that individuals cured from cutaneous leishmaniasis (CL) are protected from further infection. First generation Leishmania vaccines consisting of whole killed parasites reached to phase 3 clinical trials but failed to show enough efficacies mainly due to the lack of an appropriate adjuvant. In this study, an efficient liposomal protein-based vaccine against Leishmania major infection was developed using soluble Leishmania antigens (SLA) as a first generation vaccine and cytidine phosphate guanosine oligodeoxynucleotides (CpG ODNs) as an immunostimulatory adjuvant. 1, 2-Dioleoyl-3-trimethylammonium-propane was used as a cationic lipid to prepare the liposomes due to its intrinsic adjuvanticity. BALB/c mice were immunized subcutaneously (SC), three times in 2-week intervals, with Lip-SLA-CpG, Lip-SLA, SLA + CpG, SLA, or HEPES buffer. As criteria for protection, footpad swelling at the site of challenge and spleen parasite loads were assessed, and the immune responses were evaluated by determination of IFN-γ and IL-4 levels of cultured splenocytes, and IgG subtypes. The group of mice that received Lip-SLA-CpG showed a significantly smaller footpad swelling, lower spleen parasite burden, higher IgG2a antibody, and lower IL-4 level compared to the control groups. It is concluded that cationic liposomes containing SLA and CpG ODNs are appropriate to induce Th1 type of immune response and protection against leishmaniasis.

In vitro interaction between SURFACEN® and surfactant protein A against Leishmania amazonensis.

PubMed

de Guevara, Yuliannis Lugones Ladrón; Hidalgo, Odalys Blanco; Faure, Roberto; Fidalgo, Lianet Monzote

2013-01-01

Leishmaniasis is caused by a parasite of the Leishmania genus, affecting more than 12 million people in 98 countries. The control of leishmaniasis remains a serious problem. There are currently no vaccines for leishmaniasis. The drugs available are toxic, expensive and frequently ineffective. The in vitro activity of SURFACEN® and SP-A against Leishmania amazonensis was evaluated. The combination of both products resulted in a synergic pharmacology effect, demonstrated by a fractional inhibitory concentration index <0.5. A more effective combination was a SURFACEN/SP-A ratio of 4:1, using a method of fixed ratio. The therapeutic effect of SURFACEN and SP-A as antileishmanial compounds was demonstrated, with a potentiation of activity when they were incubated in conjunction. Our results propose an exploration of these products in order to design new formulations against the Leishmania parasite. © 2014 S. Karger AG, Basel.

Vaccination with a Leishmania infantum HSP70-II null mutant confers long-term protective immunity against Leishmania major infection in two mice models

PubMed Central

Solana, José Carlos; Ramírez, Laura; Corvo, Laura; de Oliveira, Camila Indiani; Barral-Netto, Manoel; Requena, José María

2017-01-01

Background The immunization with genetically attenuated Leishmania cell lines has been associated to the induction of memory and effector T cell responses against Leishmania able to control subsequent challenges. A Leishmania infantum null mutant for the HSP70-II genes has been described, possessing a non-virulent phenotype. Methodology/Principal findings The L. infantum attenuated parasites (LiΔHSP70-II) were inoculated in BALB/c (intravenously and subcutaneously) and C57BL/6 (subcutaneously) mice. An asymptomatic infection was generated and parasites diminished progressively to become undetectable in most of the analyzed organs. However, inoculation resulted in the long-term induction of parasite specific IFN-γ responses able to control the disease caused by a challenge of L. major infective promastigotes. BALB/c susceptible mice showed very low lesion development and a drastic decrease in parasite burdens in the lymph nodes draining the site of infection and internal organs. C57BL/6 mice did not show clinical manifestation of disease, correlated to the rapid migration of Leishmania specific IFN-γ producing T cells to the site of infection. Conclusion/Significance Inoculation of the LiΔHSP70-II attenuated line activates mammalian immune system for inducing moderate pro-inflammatory responses. These responses are able to confer long-term protection in mice against the infection of L. major virulent parasites. PMID:28558043

Expression, purification, immunogenicity and protective efficacy of a recombinant nucleoside hydrolase from Leishmania donovani, a vaccine candidate for preventing cutaneous leishmaniasis.

PubMed

McAtee, C Patrick; Seid, Christopher A; Hammond, Molly; Hudspeth, Elissa; Keegan, Brian P; Liu, Zhuyun; Wei, Junfei; Zhan, Bin; Arjona-Sabido, Raul; Cruz-Chan, Vladimir; Dumonteil, Eric; Hotez, Peter J; Bottazzi, Maria Elena

2017-02-01

The nucleoside hydrolase gene from Leishmania donovani was cloned and expressed in Escherichia coli as a full length 36-kDa protein (LdNH36). Following lysis and extraction, the protein was purified by anion exchange and gel filtration chromatography. The purified protein had a molecular mass of approximately 36-kDa and was confirmed to be >99% pure. Using a nucleoside hydrolase assay, the protein was found to exhibit a Km of 741 ± 246 μM. Protein integrity was confirmed by lithium dodecyl sulfate polyacrylamide gel electrophoresis (LDS-PAGE), mass spectrometry (MS), and enzymatic assay. Analysis of antibody levels from immunized mice indicated that LdNH36 alone or in a stable emulsion with the Toll-like receptor-4 ligand glucopyranosyl lipid adjuvant (GLA-SE) as immunostimulant induced high levels of antigen-specific IgG antibodies. The cellular immune response indicated a T h 1 response in mice immunized with LdNH36, but only when formulated with GLA-SE. Mice immunized with the LdNH36 antigen in combination with the GLA-SE adjuvant and challenged with Leishmania mexicana showed significant reductions (>20 fold) in parasite burden, confirming the protective efficacy of this vaccine candidate. Copyright © 2016 Elsevier Inc. All rights reserved.

Deception and Manipulation: The Arms of Leishmania, a Successful Parasite

PubMed Central

Cecílio, Pedro; Pérez-Cabezas, Begoña; Santarém, Nuno; Maciel, Joana; Rodrigues, Vasco; Cordeiro da Silva, Anabela

2014-01-01

Leishmania spp. are intracellular parasitic protozoa responsible for a group of neglected tropical diseases, endemic in 98 countries around the world, called leishmaniasis. These parasites have a complex digenetic life cycle requiring a susceptible vertebrate host and a permissive insect vector, which allow their transmission. The clinical manifestations associated with leishmaniasis depend on complex interactions between the parasite and the host immune system. Consequently, leishmaniasis can be manifested as a self-healing cutaneous affliction or a visceral pathology, being the last one fatal in 85–90% of untreated cases. As a result of a long host–parasite co-evolutionary process, Leishmania spp. developed different immunomodulatory strategies that are essential for the establishment of infection. Only through deception and manipulation of the immune system, Leishmania spp. can complete its life cycle and survive. The understanding of the mechanisms associated with immune evasion and disease progression is essential for the development of novel therapies and vaccine approaches. Here, we revise how the parasite manipulates cell death and immune responses to survive and thrive in the shadow of the immune system. PMID:25368612

Evaluation of Live Recombinant Nonpathogenic Leishmania tarentolae Expressing Cysteine Proteinase and A2 Genes as a Candidate Vaccine against Experimental Canine Visceral Leishmaniasis.

PubMed

Shahbazi, Mehdi; Zahedifard, Farnaz; Taheri, Tahereh; Taslimi, Yasaman; Jamshidi, Shahram; Shirian, Sadegh; Mahdavi, Niousha; Hassankhani, Mehdi; Daneshbod, Yahya; Zarkesh-Esfahani, Sayyed Hamid; Papadopoulou, Barbara; Rafati, Sima

2015-01-01

Canine Visceral Leishmaniasis (CVL) is a major veterinary and public health problem caused by Leishmania infantum (L. infantum) in many endemic countries. It is a severe chronic disease with generalized parasite spread to the reticuloendothelial system, such as spleen, liver and bone marrow and is often fatal when left untreated. Control of VL in dogs would dramatically decrease infection pressure of L. infantum for humans, since dogs are the main domestic reservoir. In the past decade, various subunits and DNA antigens have been identified as potential vaccine candidates in experimental animal models, but none has been approved for human use so far. In this study, we vaccinated outbreed dogs with a prime-boost regimen based on recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinase genes (CPA and CPB without its unusual C-terminal extension (CPB-CTE) and evaluated its immunogenicity and protective immunity against L. infantum infectious challenge. We showed that vaccinated animals produced significantly higher levels of IgG2, but not IgG1, and also IFN-γ and TNF-α, but low IL-10 levels, before and after challenge as compared to control animals. Protection in dogs was also correlated with a strong DTH response and low parasite burden in the vaccinated group. Altogether, immunization with recombinant L. tarentolae A2-CPA-CPB-CTE was proven to be immunogenic and induced partial protection in dogs, hence representing a promising live vaccine candidate against CVL.

Evaluation of Live Recombinant Nonpathogenic Leishmania tarentolae Expressing Cysteine Proteinase and A2 Genes as a Candidate Vaccine against Experimental Canine Visceral Leishmaniasis

PubMed Central

Shahbazi, Mehdi; Zahedifard, Farnaz; Taheri, Tahereh; Taslimi, Yasaman; Jamshidi, Shahram; Shirian, Sadegh; Mahdavi, Niousha; Hassankhani, Mehdi; Daneshbod, Yahya; Zarkesh-Esfahani, Sayyed Hamid; Papadopoulou, Barbara; Rafati, Sima

2015-01-01

Canine Visceral Leishmaniasis (CVL) is a major veterinary and public health problem caused by Leishmania infantum (L. infantum) in many endemic countries. It is a severe chronic disease with generalized parasite spread to the reticuloendothelial system, such as spleen, liver and bone marrow and is often fatal when left untreated. Control of VL in dogs would dramatically decrease infection pressure of L. infantum for humans, since dogs are the main domestic reservoir. In the past decade, various subunits and DNA antigens have been identified as potential vaccine candidates in experimental animal models, but none has been approved for human use so far. In this study, we vaccinated outbreed dogs with a prime-boost regimen based on recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinase genes (CPA and CPB without its unusual C-terminal extension (CPB-CTE) and evaluated its immunogenicity and protective immunity against L. infantum infectious challenge. We showed that vaccinated animals produced significantly higher levels of IgG2, but not IgG1, and also IFN-γ and TNF-α, but low IL-10 levels, before and after challenge as compared to control animals. Protection in dogs was also correlated with a strong DTH response and low parasite burden in the vaccinated group. Altogether, immunization with recombinant L. tarentolae A2-CPA-CPB-CTE was proven to be immunogenic and induced partial protection in dogs, hence representing a promising live vaccine candidate against CVL. PMID:26197085

Progress toward development of photodynamic vaccination against infectious/malignant diseases and photodynamic mosquitocides

NASA Astrophysics Data System (ADS)

Chang, Kwang Poo; Kolli, Bala K.; Fan, Chia-Kwung; Ng, Dennis K. P.; Wong, Clarence T. T.; Manna, Laura; Corso, Raffaele; Shih, Neng-Yao; Elliott, Robert; Jiang, X. P.; Shiao, Shin-Hong; Fu, Guo-Liang

2018-02-01

Photodynamic therapy (PDT) uses photosensitizers (PS) that are excited with light to generate ROS in the presence of oxygen for treating various diseases. PS also has the potential use as photodynamic insecticides (PDI) and for light-inactivation of Leishmania for photodynamic vaccination (PDV). PDT-inactivated Leishmania are non-viable, but remain immunologically competent as whole-cell vaccines against leishmaniasis, and as a universal carrier for delivery of add-on vaccines against other infectious and malignant diseases. We have screened novel PS, including Zn- and Si-phthalocyanines (PC) for differential PDT activities against Leishmania, insect and mammalian cells in vitro to assess their PDI and PDV potential. Here, Zn-PC were conjugated with various functional groups. The conjugates were examined for uptake by cells as a prerequisite for their susceptibility to light-inactivation. PDT sensitivity was found to vary with cell types and PS used. PDI potential of several PS was demonstrated by their mosquito larvicidal PDT activities in vitro. PDT-inactivated Leishmania were stored frozen for PDV in several ongoing studies: [1] Open label trial with 20 sick dogs for immunotherapy of canine leishmaniasis after chemotherapy in Naples, Italy. Clinical follow-up for >3 years indicate that the PDV prolongs their survival; [2] PDV of murine models with a human lung cancer vaccine showed dramatic tumor suppression; [3] Open label trial of multiple PDV via compassionate access to 4 advanced cancer patients showed no clinically adverse effects. Two subjects remain alive. Genetic modifications of Leishmania are underway to further enhance their safety and efficacy for PDV by installation of activable mechanisms for self-destruction and spontaneous light-emission.

Anti-Leishmania Activity of Osthole

PubMed Central

Kermani, Elaheh Kordzadeh; Sajjadi, Seyed Ebrahim; Hejazi, Seyed Hossein; Arjmand, Reza; Saberi, Sedigheh; Eskandarian, Abbas Ali

2016-01-01

Background: Treatment of cutaneous leishmaniasis (CL) is occasionally highly resistant to pentavalent antimonials, the gold standard in pharmacotherapy of CL. Since there is no effective vaccine, the discovery of natural antileishmanial products as complementary therapeutic agents could be used to improve the current regimens. Objective: In this study in vitro and in vivo antileishmanial activities of osthole, a natural coumarin known to possess antibacterial and parasiticidal activities are evaluated. Materials and Methods: Leishmania major infected J774.A1 macrophages were treated with increasing concentrations of osthole. CL lesions of BALB/c mice were treated topically with 0.2% osthole. Results: Osthole exhibited dose-dependent leishmanicidal activity against intracellular amastigotes with IC50 value of 14.95 μg/ml. Treatment of CL lesions in BALB/c mice with osthole significantly declined lesion progression compared to untreated mice (P < 0.05), however did not result in recovery. Conclusion: Osthole demonstrated remarkable leishmanicidal activity in vitro. Higher concentrations of osthole may demonstrate the therapeutic property in vivo. SUMMARY In vitro and in vivo antileishmanial activities of osthole, a pernylated coumarin extracted from Prangos asperula Boiss., are studied against Leishmania major. PMID:27114685

Persistent parasites and immunologic memory in cutaneous leishmaniasis: implications for vaccine designs and vaccination strategies.

PubMed

Okwor, Ifeoma; Uzonna, Jude

2008-01-01

Despite a plethora of publications on the murine model of cutaneous leishmaniasis and their contribution to our understanding of the factors that regulate the development of CD4+ T cell immunity in vivo, there is still no effective vaccine against the human disease. While recovery from natural or experimental infection with Leishmania major, the causative agent of human cutaneous leishmaniasis, results in persistence of parasites at the primary infection site and the development of long-lasting immunity to reinfection, vaccination with killed parasites or recombinant proteins induces only short-term protection. The reasons for the difference in protective immunity following recovery from live infection and vaccination with heat-killed parasites are not known. This may in part be related to persistence of live parasites following healing of primary cutaneous lesions, because complete clearance of parasites leads to rapid loss of infection-induced immunity. Recent reports indicate that in addition to persistent parasites, IL-10-producing natural regulatory T cells may also play critical roles in the maintenance and loss of infection-induced immunity. This review focuses on current understanding of the factors that regulate the development, maintenance and loss of anti-Leishmania memory responses and highlights the role of persistent parasites and regulatory T cells in this process. Understanding these factors is crucial for designing effective vaccines and vaccination strategies against cutaneous leishmaniasis.

Is influenza vaccination in asthma helpful?

PubMed

Bueving, Herman J; Thomas, Siep; Wouden, Johannes C van der

2005-02-01

Influenza infections are frequently involved in asthma exacerbations. During influenza epidemics substantial excess morbidity due to respiratory tract complications is reported in all age categories as well as excess mortality among the elderly. Vaccines are available for protection against influenza. Worldwide, vaccination is advised and considered a quality point for asthma care. However, the protective effect of influenza vaccination in patients with asthma is still disputed. In order to establish the current state of affairs we reviewed the recent literature on the protective effect of influenza vaccination and its usefulness in patients with asthma. Several studies were found addressing influenza and the protective aspects of vaccination. They discussed the incidence, the adverse effects of vaccination, the coverage of influenza vaccination among patients with asthma and the effectiveness of the vaccine. Influenza vaccination can safely be used in patients with asthma. Allegations that vaccination could provoke asthma exacerbations are convincingly invalidated by previous and recent research. Although patients with asthma are one of the major target groups for immunization, vaccine coverage in all age categories remains low. So far, no unequivocal beneficial effect of influenza vaccination in patients with asthma was found in observational and experimental studies in the sense of reduction of asthma exacerbations and other complications. Recent studies confirm these negative findings. More long-term randomized, placebo-controlled studies, focusing on influenza- proven illness in patients with asthma, are needed to address the question of how helpful influenza vaccination is in these patients.

Strategic evaluation of vaccine candidate antigens for the prevention of Visceral Leishmaniasis.

PubMed

Duthie, Malcolm S; Favila, Michelle; Hofmeyer, Kimberley A; Tutterrow, Yeung L; Reed, Steven J; Laurance, John D; Picone, Alessandro; Guderian, Jeffrey; Bailor, H Remy; Vallur, Aarthy C; Liang, Hong; Mohamath, Raodoh; Vergara, Julie; Howard, Randall F; Coler, Rhea N; Reed, Steven G

2016-05-27

Infection with Leishmania parasites results in a range of clinical manifestations and outcomes, the most severe of which is visceral leishmaniasis (VL). Vaccination will likely provide the most effective long-term control strategy, as the large number of vectors and potential infectious reservoirs renders sustained interruption of Leishmania parasite transmission extremely difficult. Selection of the best vaccine is complicated because, although several vaccine antigen candidates have been proposed, they have emerged following production in different platforms. To consolidate the information that has been generated into a single vaccine platform, we expressed seven candidates as recombinant proteins in E. coli. After verifying that each recombinant protein could be recognized by VL patients, we evaluated their protective efficacy against experimental L. donovani infection of mice. Administration in formulation with the Th1-potentiating adjuvant GLA-SE indicated that each antigen could elicit antigen-specific Th1 responses that were protective. Considering the ability to reduce parasite burden along with additional factors such as sequence identity across Leishmania species, we then generated a chimeric fusion protein comprising a combination of the 8E, p21 and SMT proteins. This E. coli -expressed fusion protein was also demonstrated to protect against L. donovani infection. These data indicate a novel recombinant vaccine antigen with the potential for use in VL control programs. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Pneumococcal vaccination and chronic respiratory diseases.

PubMed

Froes, Filipe; Roche, Nicolas; Blasi, Francesco

2017-01-01

Patients with COPD and other chronic respiratory diseases are especially vulnerable to viral and bacterial pulmonary infections, which are major causes of exacerbations, hospitalization, disease progression, and mortality in COPD patients. Effective vaccines could reduce the burden of respiratory infections and acute exacerbations in COPD patients, but what is the evidence for this? This article reviews and discusses the existing evidence for pneumococcal vaccination efficacy and its changing role in patients with chronic respiratory diseases, especially COPD. Specifically, the recent Community-Acquired Pneumonia Immunization Trial in Adults (CAPITA) showed the efficacy of pneumococcal conjugate vaccine in older adults, many of whom had additional risk factors for pneumococcal disease, including chronic lung diseases. Taken together, the evidence suggests that pneumococcal and influenza vaccinations can prevent community-acquired pneumonia and acute exacerbations in COPD patients, while pneumococcal vaccination early in the course of COPD could help maintain stable health status. Despite the need to prevent pulmonary infections in patients with chronic respiratory diseases and evidence for the efficacy of pneumococcal conjugate vaccine, pneumococcal vaccine coverage and awareness are low and need to be improved. Respiratory physicians need to communicate the benefits of vaccination more effectively to their patients who suffer from chronic respiratory diseases.

Neutrophils Contribute to the Protection Conferred by ArtinM against Intracellular Pathogens: A Study on Leishmania major

PubMed Central

Ricci-Azevedo, Rafael; Oliveira, Aline Ferreira; Conrado, Marina C. A. V.; Carvalho, Fernanda Caroline; Roque-Barreira, Maria Cristina

2016-01-01

ArtinM, a D-mannose binding lectin from Artocarpus heterophyllus, has immunomodulatory activities through its interaction with N-glycans of immune cells, culminating with the establishment of T helper type 1 (Th1) immunity. This interaction protects mice against intracellular pathogens, including Leishmania major and Leishmania amazonensis. ArtinM induces neutrophils activation, which is known to account for both resistance to pathogens and host tissue injury. Although exacerbated inflammation was not observed in ArtinM-treated animals, assessment of neutrophil responses to ArtinM is required to envisage its possible application to design a novel immunomodulatory agent based on carbohydrate recognition. Herein, we focus on the mechanisms through which neutrophils contribute to ArtinM-induced protection against Leishmania, without exacerbating inflammation. For this purpose, human neutrophils treated with ArtinM and infected with Leishmania major were analyzed together with untreated and uninfected controls, based on their ability to eliminate the parasite, release cytokines, degranulate, produce reactive oxygen species (ROS), form neutrophil extracellular traps (NETs) and change life span. We demonstrate that ArtinM-stimulated neutrophils enhanced L. major clearance and at least duplicated tumor necrosis factor (TNF) and interleukin-1beta (IL-1β) release; otherwise, transforming growth factor-beta (TGF-β) production was reduced by half. Furthermore, ROS production and cell degranulation were augmented. The life span of ArtinM-stimulated neutrophils decreased and they did not form NETs when infected with L. major. We postulate that the enhanced leishmanicidal ability of ArtinM-stimulated neutrophils is due to augmented release of inflammatory cytokines, ROS production, and cell degranulation, whereas host tissue integrity is favored by their shortened life span and the absence of NET formation. Our results reinforce the idea that ArtinM may be considered an

Neutrophils Contribute to the Protection Conferred by ArtinM against Intracellular Pathogens: A Study on Leishmania major.

PubMed

Ricci-Azevedo, Rafael; Oliveira, Aline Ferreira; Conrado, Marina C A V; Carvalho, Fernanda Caroline; Roque-Barreira, Maria Cristina

2016-04-01

ArtinM, a D-mannose binding lectin from Artocarpus heterophyllus, has immunomodulatory activities through its interaction with N-glycans of immune cells, culminating with the establishment of T helper type 1 (Th1) immunity. This interaction protects mice against intracellular pathogens, including Leishmania major and Leishmania amazonensis. ArtinM induces neutrophils activation, which is known to account for both resistance to pathogens and host tissue injury. Although exacerbated inflammation was not observed in ArtinM-treated animals, assessment of neutrophil responses to ArtinM is required to envisage its possible application to design a novel immunomodulatory agent based on carbohydrate recognition. Herein, we focus on the mechanisms through which neutrophils contribute to ArtinM-induced protection against Leishmania, without exacerbating inflammation. For this purpose, human neutrophils treated with ArtinM and infected with Leishmania major were analyzed together with untreated and uninfected controls, based on their ability to eliminate the parasite, release cytokines, degranulate, produce reactive oxygen species (ROS), form neutrophil extracellular traps (NETs) and change life span. We demonstrate that ArtinM-stimulated neutrophils enhanced L. major clearance and at least duplicated tumor necrosis factor (TNF) and interleukin-1beta (IL-1β) release; otherwise, transforming growth factor-beta (TGF-β) production was reduced by half. Furthermore, ROS production and cell degranulation were augmented. The life span of ArtinM-stimulated neutrophils decreased and they did not form NETs when infected with L. major. We postulate that the enhanced leishmanicidal ability of ArtinM-stimulated neutrophils is due to augmented release of inflammatory cytokines, ROS production, and cell degranulation, whereas host tissue integrity is favored by their shortened life span and the absence of NET formation. Our results reinforce the idea that ArtinM may be considered an

Novel selective inhibitor of Leishmania (Leishmania) amazonensis arginase.

PubMed

da Silva, Edson R; Boechat, Nubia; Pinheiro, Luiz C S; Bastos, Monica M; Costa, Carolina C P; Bartholomeu, Juliana C; da Costa, Talita H

2015-11-01

Arginase is a glycosomal enzyme in Leishmania that is involved in polyamine and trypanothione biosynthesis. The central role of arginase in Leishmania (Leishmania) amazonensis was demonstrated by the generation of two mutants: one with an arginase lacking the glycosomal addressing signal and one in which the arginase-coding gene was knocked out. Both of these mutants exhibited decreased infectivity. Thus, arginase seems to be a potential drug target for Leishmania treatment. In an attempt to search for arginase inhibitors, 29 derivatives of the [1,2,4]triazolo[1,5-a]pyrimidine system were tested against Leishmania (Leishmania) amazonensis arginase in vitro. The [1,2,4]triazolo[1,5-a]pyrimidine scaffold containing R1  = CF3 exhibited greater activity against the arginase rather than when the substituent R1  = CH3 in the 2-position. The novel compound 2-(5-methyl-2-(trifluoromethyl)-[1,2,4]triazolo[1,5-a]pyrimidin-7-yl)hydrazinecarbothioamide (30) was the most potent, inhibiting arginase by a non-competitive mechanism, with the Ki and IC50 values for arginase inhibition estimated to be 17 ± 1 μm and 16.5 ± 0.5 μm, respectively. These results can guide the development of new drugs against leishmaniasis based on [1,2,4]triazolo[1,5-a]pyrimidine derivatives targeting the arginase enzyme. © 2015 John Wiley & Sons A/S.

Prevalence and Distribution of Leishmania RNA Virus 1 in Leishmania Parasites from French Guiana

PubMed Central

Ginouvès, Marine; Simon, Stéphane; Bourreau, Eliane; Lacoste, Vincent; Ronet, Catherine; Couppié, Pierre; Nacher, Mathieu; Demar, Magalie; Prévot, Ghislaine

2016-01-01

In South America, the presence of the Leishmania RNA virus type 1 (LRV1) was described in Leishmania guyanensis and Leishmania braziliensis strains. The aim of this study was to determine the prevalence distribution of LRV1 in Leishmania isolates in French Guiana given that, in this French overseas department, most Leishmania infections are due to these parasite species. The presence of the virus was observed in 74% of Leishmania sp. isolates, with a highest presence in the internal areas of the country. PMID:26598572

Parasite Carbohydrate Vaccines.

PubMed

Jaurigue, Jonnel A; Seeberger, Peter H

2017-01-01

Vaccination is an efficient means of combating infectious disease burden globally. However, routine vaccines for the world's major human parasitic diseases do not yet exist. Vaccines based on carbohydrate antigens are a viable option for parasite vaccine development, given the proven success of carbohydrate vaccines to combat bacterial infections. We will review the key components of carbohydrate vaccines that have remained largely consistent since their inception, and the success of bacterial carbohydrate vaccines. We will then explore the latest developments for both traditional and non-traditional carbohydrate vaccine approaches for three of the world's major protozoan parasitic diseases-malaria, toxoplasmosis, and leishmaniasis. The traditional prophylactic carbohydrate vaccine strategy is being explored for malaria. However, given that parasite disease biology is complex and often arises from host immune responses to parasite antigens, carbohydrate vaccines against deleterious immune responses in host-parasite interactions are also being explored. In particular, the highly abundant glycosylphosphatidylinositol molecules specific for Plasmodium, Toxoplasma , and Leishmania spp. are considered exploitable antigens for this non-traditional vaccine approach. Discussion will revolve around the application of these protozoan carbohydrate antigens for vaccines currently in preclinical development.

Parasite Carbohydrate Vaccines

PubMed Central

Jaurigue, Jonnel A.; Seeberger, Peter H.

2017-01-01

Vaccination is an efficient means of combating infectious disease burden globally. However, routine vaccines for the world's major human parasitic diseases do not yet exist. Vaccines based on carbohydrate antigens are a viable option for parasite vaccine development, given the proven success of carbohydrate vaccines to combat bacterial infections. We will review the key components of carbohydrate vaccines that have remained largely consistent since their inception, and the success of bacterial carbohydrate vaccines. We will then explore the latest developments for both traditional and non-traditional carbohydrate vaccine approaches for three of the world's major protozoan parasitic diseases—malaria, toxoplasmosis, and leishmaniasis. The traditional prophylactic carbohydrate vaccine strategy is being explored for malaria. However, given that parasite disease biology is complex and often arises from host immune responses to parasite antigens, carbohydrate vaccines against deleterious immune responses in host-parasite interactions are also being explored. In particular, the highly abundant glycosylphosphatidylinositol molecules specific for Plasmodium, Toxoplasma, and Leishmania spp. are considered exploitable antigens for this non-traditional vaccine approach. Discussion will revolve around the application of these protozoan carbohydrate antigens for vaccines currently in preclinical development. PMID:28660174

A Randomised, Double-Blind, Controlled Efficacy Trial of the LiESP/QA-21 Vaccine in Naïve Dogs Exposed to Two Leishmania infantum Transmission Seasons

PubMed Central

Oliva, Gaetano; Nieto, Javier; Foglia Manzillo, Valentina; Cappiello, Silvia; Fiorentino, Eleonora; Di Muccio, Trentina; Scalone, Aldo; Moreno, Javier; Chicharro, Carmen; Carrillo, Eugenia; Butaud, Therese; Guegand, Laurie; Martin, Virginie; Cuisinier, Anne-Marie; McGahie, David; Gueguen, Sylvie; Cañavate, Carmen; Gradoni, Luigi

2014-01-01

Canine leishmaniasis is an important zoonosis caused by uncontrolled infection with Leishmania infantum, where an inappropriate immune response is not only responsible for permitting this intracellular parasite to multiply, but is also responsible for several of the pathological processes seen in this disease. Effective canine vaccines are therefore a highly desirable prevention tool. In this randomised, double-blinded, controlled trial, the efficacy of the LiESP/QA-21 vaccine (CaniLeish, Virbac, France) was assessed by exposing 90 naïve dogs to natural L. infantum infection during 2 consecutive transmission seasons, in two highly endemic areas of the Mediterranean basin. Regular PCR, culture, serological and clinical examinations were performed, and the infection/disease status of the dogs was classified at each examination. The vaccine was well-tolerated, and provided a significant reduction in the risk of progressing to uncontrolled active infection (p = 0.025) or symptomatic disease (p = 0.046), with an efficacy of 68.4% and a protection rate of 92.7%. The probability of becoming PCR positive was similar between groups, but the probability of returning to a PCR negative condition was higher in the vaccinated group (p = 0.04). In conclusion, we confirmed the interest of using this vaccine as part of a comprehensive control program for canine leishmaniasis, and validated the use of a protocol based on regular in-depth assessments over time to assess the efficacy of a canine leishmaniasis vaccine. PMID:25299614

Prevalence and Distribution of Leishmania RNA Virus 1 in Leishmania Parasites from French Guiana.

PubMed

Ginouvès, Marine; Simon, Stéphane; Bourreau, Eliane; Lacoste, Vincent; Ronet, Catherine; Couppié, Pierre; Nacher, Mathieu; Demar, Magalie; Prévot, Ghislaine

2016-01-01

In South America, the presence of the Leishmania RNA virus type 1 (LRV1) was described in Leishmania guyanensis and Leishmania braziliensis strains. The aim of this study was to determine the prevalence distribution of LRV1 in Leishmania isolates in French Guiana given that, in this French overseas department, most Leishmania infections are due to these parasite species. The presence of the virus was observed in 74% of Leishmania spp. isolates, with a highest presence in the internal areas of the country. © The American Society of Tropical Medicine and Hygiene.

DNA vaccine against visceral leishmaniasis: a promising approach for prevention and control.

PubMed

Kumar, A; Samant, M

2016-05-01

The visceral leishmaniasis (VL) caused by Leishmania donovani parasite severely affects large populations in tropical and subtropical regions of the world. The arsenal of drugs available is limited, and resistance is common in clinical field isolates. Therefore, vaccines could be an important alternative for prevention against VL. Recently, some investigators advocated the protective efficacy of DNA vaccines, which induces the T cell-based immunity against VL. The vaccine antigens are selected as conserved in various Leishmania species and provide a viable strategy for DNA vaccine development. Our understanding for DNA vaccine development against VL is not enough and much technological advancement is required. Improved formulations and methods of delivery are required, which increase the uptake of DNA vaccine by cells; optimization of vaccine vectors/encoded antigens to augment and direct the host immune response in VL. Despite the many genes identified as vaccine candidates, the disappointing potency of the DNA vaccines in VL underscores the challenges encountered in the efforts to translate efficacy in preclinical models into clinical realities. This review will provide a brief background of DNA vaccines including the insights gained about the design, strategy, safety issues, varied candidates, progress and challenges that play a role in their ability against VL. © 2016 John Wiley & Sons Ltd.

Recombinant small glutamine-rich tetratricopeptide repeat-containing protein of Leishmania infantum: Potential vaccine and diagnostic application against visceral leishmaniasis.

PubMed

Dias, Daniel S; Ribeiro, Patrícia A F; Martins, Vívian T; Lage, Daniela P; Portela, Áquila S B; Costa, Lourena E; Salles, Beatriz C S; Lima, Mariana P; Ramos, Fernanda F; Santos, Thaís T O; Caligiorne, Rachel B; Chávez-Fumagalli, Miguel A; Silveira, Julia A G; Magalhães-Soares, Danielle F; Gonçalves, Denise U; Oliveira, Jamil S; Roatt, Bruno M; Duarte, Mariana C; Menezes-Souza, Daniel; Silva, Eduardo S; Galdino, Alexsandro S; Machado-de-Ávila, Ricardo A; Teixeira, Antonio L; Coelho, Eduardo A F

2017-11-01

Different Leishmania proteins have been evaluated in order to find a potential vaccine candidate or diagnostic marker capable of providing long lasting protection against infection or helping to identify infected mammalian hosts, respectively. However, just few molecules have fulfilled all the requirements to be evaluated. In the current study, we evaluated the prophylactic and diagnostic value against visceral leishmaniasis (VL) of a small glutamine-rich tetratricopeptide repeat-containing (SGT) protein from Leishmania infantum species. In a first step, the immune response elicited by the immunization using the recombinant protein (rSGT) plus saponin was evaluated in BALB/c mice. Immunized animals had a low parasitism in all evaluated organs. They developed a specific Th1 immune response, which was based on protein-specific production of IFN-γ, IL-12 and GM-CSF, and a humoral response dominated by antibodies of the IgG2a isotype. Both CD4 + and CD8 + T cells contributed to the IFN-γ production, showing that both T cell subtypes contribute to the resistance against infection. Regarding its value as a diagnostic marker, rSGT showed maximum sensitivity and specificity to serologically identify L. infantum-infected dog and human sera. No cross-reactivity with sera from humans or dogs that had other diseases was found. Although further studies are necessary to validate these findings, data showed here suggest immunogenicity of rSGT and its protective effect against murine VL, as well as its potential for the serodiagnosis of human and canine VL. Copyright © 2017 Elsevier Ltd. All rights reserved.

Vaccines for leishmaniasis in the fore coming 25 years.

PubMed

Palatnik-de-Sousa, Clarisa B

2008-03-25

Human vaccination against leishmaniasis using live Leishmania was used in Middle East and Russia (1941-1980). First-generation vaccines, composed by killed parasites induce low efficacies (54%) and were tested in humans and dogs Phase III trials in Asia and South America since 1940. Second-generation vaccines using live genetically modified parasites, or bacteria or viruses containing Leishmania genes, recombinant or native fractions are known since the 1990s. Due to the loss of PAMPs, the use of adjuvants increased vaccine efficacies of the purified antigens to 82%, in Phase III dog trials. Recombinant second-generation vaccines and third-generation DNA vaccines showed average values of parasite load reduction of 68% and 59% in laboratory animal models, respectively, but their success in field trials had not yet been reported. This review is focused on vaccine candidates that show any efficacy against leishmaniasis and that are already in different phase trials. A lot of interest though was generated in recent years, by the studies going on in experimental models. The promising candidates may find a place in the forth coming years. Among them most probably are the multiple-gene DNA vaccines that are stable and do not require cold-chain transportation. In the mean time, second-generation vaccines with native antigens and effective adjuvants are likely to be licensed and used in Public Health control programs in the fore coming 25 years. To date, only three vaccines have been licensed for use: one live vaccine for humans in Uzbekistan, one killed vaccine for human immunotherapy in Brazil and a second-generation vaccine for dog prophylaxis in Brazil.

Detection and molecular identification of leishmania RNA virus (LRV) in Iranian Leishmania species.

PubMed

Hajjaran, Homa; Mahdi, Maryam; Mohebali, Mehdi; Samimi-Rad, Katayoun; Ataei-Pirkooh, Angila; Kazemi-Rad, Elham; Naddaf, Saied Reza; Raoofian, Reza

2016-12-01

Leishmania RNA virus (LRV) was first detected in members of the subgenus Leishmania (Viannia), and later, the virulence and metastasis of the New World species were attributed to this virus. The data on the presence of LRV in Old World species are confined to Leishmania major and a few Leishmania aethiopica isolates. The aim of this study was to survey the presence of LRV in various Iranian Leishmania species originating from patients and animal reservoir hosts. Genomic nucleic acids were extracted from 50 cultured isolates belonging to the species Leishmania major, Leishmania tropica, and Leishmania infantum. A partial sequence of the viral RNA-dependent RNA polymerase (RdRp) gene was amplified, sequenced and compared with appropriate sequences from the GenBank database. We detected the virus in two parasite specimens: an isolate of L. infantum derived from a visceral leishmaniasis (VL) patient who was unresponsive to meglumine antimoniate treatment, and an L. major isolate originating from a great gerbil, Rhombomys opimus. The Iranian LRV sequences showed the highest similarities to an Old World L. major LRV2 and were genetically distant from LRV1 isolates detected in New World Leishmania parasites. We could not attribute treatment failure in VL patient to the presence of LRV due to the limited number of specimens analyzed. Further studies with inclusion of more clinical samples are required to elucidate the potential role of LRVs in pathogenesis or treatment failure of Old World leishmaniasis.

Leishmaniasis in humans: drug or vaccine therapy?

PubMed Central

Ghorbani, Masoud; Farhoudi, Ramin

2018-01-01

Leishmania is an obligate intracellular pathogen that invades phagocytic host cells. Approximately 30 different species of Phlebotomine sand flies can transmit this parasite either anthroponotically or zoonotically through their bites. Leishmaniasis affects poor people living around the Mediterranean Basin, East Africa, the Americas, and Southeast Asia. Affected regions are often remote and unstable, with limited resources for treating this disease. Leishmaniasis has been reported as one of the most dangerous neglected tropical diseases, second only to malaria in parasitic causes of death. People can carry some species of Leishmania for long periods without becoming ill, and symptoms depend on the form of the disease. There are many drugs and candidate vaccines available to treat leishmaniasis. For instance, antiparasitic drugs, such as amphotericin B (AmBisome), are a treatment of choice for leishmaniasis depending on the type of the disease. Despite the availability of different treatment approaches to treat leishmaniasis, therapeutic tools are not adequate to eradicate this infection. In the meantime, drug therapy has been limited because of adverse side effects and unsuccessful vaccine preparation. However, it can immediately make infections inactive. According to other studies, vaccination cannot eradicate leishmaniasis. There is no perfect vaccine or suitable drug to eradicate leishmaniasis completely. So far, no vaccine or drug has been provided to induce long-term protection and ensure effective immunity against leishmaniasis. Therefore, it is necessary that intensive research should be performed in drug and vaccine fields to achieve certain results. PMID:29317800

High density of Leishmania major and rarity of other mammals' Leishmania in zoonotic cutaneous leishmaniasis foci, Iran.

PubMed

Bordbar, Ali; Parvizi, Parviz

2014-03-01

Only Leishmania major is well known as a causative agent of zoonotic cutaneous leishmaniasis (ZCL) in Iran. Our objective was to find Leishmania parasites circulating in reservoir hosts, sand flies and human simultaneously. Sand flies, rodents and prepared smears of humans were sampled. DNA of Leishmania parasites was extracted, and two fragments of ITS-rDNA gene amplified by PCR. RFLP and sequencing were employed to identify Leishmania parasites. Leishmania major and L. turanica were identified unequivocally by targeting and sequencing ITS-rDNA from humans, rodents and sand flies. The new Leishmania species close to gerbilli (GenBank Accession Nos. EF413076; EF413087) was discovered only in sand flies. Based on parasite detection of ITS-rDNA in main and potential reservoir hosts and vectors and humans, we conclude that at least two Leishmania species are common in the Turkmen Sahra ZCL focus. Phylogenetic analysis proved that the new Leishmania is closely related to Leishmania mammal parasites (Leishmania major, Leishmania turanica, Leishmania gerbilli). Its role as a principal agent of ZCL is unknown because it was found only in sand flies. Our findings shed new light on the transmission cycles of several Leishmania parasites in sand flies, reservoir hosts and humans. © 2014 John Wiley & Sons Ltd.

Genetically Engineered Ascorbic acid-deficient Live Mutants of Leishmania donovani induce long lasting Protective Immunity against Visceral Leishmaniasis.

PubMed

Anand, Sneha; Madhubala, Rentala

2015-06-02

Visceral leishmaniasis caused by Leishmania donovani is the most severe systemic form of the disease. There are still no vaccines available for humans and there are limitations associated with the current therapeutic regimens for leishmaniasis. Recently, we reported functional importance of Arabino-1, 4-lactone oxidase (ALO) enzyme from L. donovani involved in ascorbate biosynthesis pathway. In this study, we have shown that ΔALO parasites do not affect the ability of null mutants to invade visceral organs but severely impair parasite persistence beyond 16 week in BALB/c mice and hence are safe as an immunogen. Both short term (5 week) and long term (20 week) immunization with ΔALO parasites conferred sustained protection against virulent challenge in BALB/c mice, activated splenocytes and resulted in induction of pro-inflammatory cytokine response. Protection in immunized mice after challenge correlated with the stimulation of IFN-γ producing CD4(+) and CD8(+) T cells. Antigen-mediated cell immunity correlated with robust nitrite and superoxide generation, macrophage-derived oxidants critical in controlling Leishmania infection. Our data shows that live attenuated ΔALO parasites are safe, induce protective immunity and can provide sustained protection against Leishmania donovani. We further conclude that the parasites attenuated in their anti-oxidative defence mechanism can be exploited as vaccine candidates.

Vaccine candidates for leishmaniasis: a review.

PubMed

Nagill, Rajeev; Kaur, Sukhbir

2011-10-01

Leishmaniasis is a diverse group of clinical syndromes caused by protozoan parasites of the genus Leishmania. The clinical manifestation of the disease varies from self-limiting cutaneous lesions to progressive visceral disease. It is estimated that 350 million people are at risk in 88 countries, with a global incidence of 1-1.5 million cases of cutaneous and 500,000 cases of visceral leishmaniasis. The key control measures mainly rely on early case detection and chemotherapy which has been hampered by the toxicity of drugs, side-effects and by the emergence of drug resistance in parasites. Control of reservoir host and vector is difficult due to operational difficulties and frequent relapses in the host. Therefore, the development of effective and affordable vaccine against leishmaniasis is highly desirable. Although considerable progress has been made over the last decade in understanding immune mechanisms underlying potential candidate antigens, including killed, live attenuated parasites, crude parasites, pure or recombinant Leishmania proteins or DNA encoding leishmanial proteins, as well as immunomodulators from sand fly saliva, very few candidate vaccines have progressed beyond the experimental stage. As such there is no vaccine against any form of human leishmaniasis. In recent years, however, much interest has been stimulated towards vaccination against leishmaniasis focused mainly on cutaneous leishmaniasis with fewer attempts against visceral leishmaniasis. Copyright © 2011 Elsevier B.V. All rights reserved.

Possibilities and challenges for developing a successful vaccine for leishmaniasis.

PubMed

Srivastava, Saumya; Shankar, Prem; Mishra, Jyotsna; Singh, Sarman

2016-05-12

Leishmaniasis is a vector-borne disease caused by different species of protozoan parasites of the genus Leishmania. It is a major health problem yet neglected tropical diseases, with approximately 350 million people worldwide at risk and more than 1.5 million infections occurring each year. Leishmaniasis has different clinical manifestations, including visceral (VL or kala-azar), cutaneous (CL), mucocutaneous (MCL), diffuse cutaneous (DCL) and post kala-azar dermal leishmaniasis (PKDL). Currently, the only mean to treat and control leishmaniasis is by rational medications and vector control. However, the number of available drugs is limited and even these are either exorbitantly priced, have toxic side effects or prove ineffective due to the emergence of resistant strains. On the other hand, the vector control methods are not so efficient. Therefore, there is an urgent need for developing a safe, effective, and affordable vaccine for the prevention of leishmaniasis. Although in recent years a large body of researchers has concentrated their efforts on this issue, yet only three vaccine candidates have gone for clinical trial, until date. These are: (i) killed vaccine in Brazil for human immunotherapy; (ii) live attenuated vaccine for humans in Uzbekistan; and (iii) second-generation vaccine for dog prophylaxis in Brazil. Nevertheless, there are at least half a dozen vaccine candidates in the pipeline. One can expect that, in the near future, the understanding of the whole genome of Leishmania spp. will expand the vaccine discovery and strategies that may provide novel vaccines. The present review focuses on the development and the status of various vaccines and potential vaccine candidates against leishmaniasis.

Establishment of Stable, Cell-Mediated Immunity that Makes "Susceptible" Mice Resistant to Leishmania major

NASA Astrophysics Data System (ADS)

Bretscher, Peter A.; Wei, Guojian; Menon, Juthika N.; Bielefeldt-Ohmann, Helle

1992-07-01

Cell-mediated, but not antibody-mediated, immune responses protect humans against certain pathogens that produce chronic diseases such as leishmaniasis. Effective vaccination against such pathogens must therefore produce an immunological "imprint" so that stable, cell-mediated immunity is induced in all individuals after natural infection. BALB/c mice "innately susceptible" to Leishmania major produce antibodies after substantial infection. In the present study, "susceptible" mice injected with a small number of parasites mounted a cell-mediated response and acquired resistance to a larger, normally pathogenic, challenge. This vaccination strategy may be applicable in diseases in which protection is dependent on cell-mediated immunity.

Dectin-1 Positive Dendritic Cells Expand after Infection with Leishmania major Parasites and Represent Promising Targets for Vaccine Development

PubMed Central

Zimara, Nicole; Chanyalew, Menberework; Aseffa, Abraham; van Zandbergen, Ger; Lepenies, Bernd; Schmid, Maximilian; Weiss, Richard; Rascle, Anne; Wege, Anja Kathrin; Jantsch, Jonathan; Schatz, Valentin; Brown, Gordon D.; Ritter, Uwe

2018-01-01

Resistant mouse strains mount a protective T cell-mediated immune response upon infection with Leishmania (L.) parasites. Healing correlates with a T helper (Th) cell-type 1 response characterized by a pronounced IFN-γ production, while susceptibility is associated with an IL-4-dependent Th2-type response. It has been shown that dermal dendritic cells are crucial for inducing protective Th1-mediated immunity. Additionally, there is growing evidence that C-type lectin receptor (CLR)-mediated signaling is involved in directing adaptive immunity against pathogens. However, little is known about the function of the CLR Dectin-1 in modulating Th1- or Th2-type immune responses by DC subsets in leishmaniasis. We characterized the expression of Dectin-1 on CD11c+ DCs in peripheral blood, at the site of infection, and skin-draining lymph nodes of L. major-infected C57BL/6 and BALB/c mice and in peripheral blood of patients suffering from cutaneous leishmaniasis (CL). Both mouse strains responded with an expansion of Dectin-1+ DCs within the analyzed tissues. In accordance with the experimental model, Dectin-1+ DCs expanded as well in the peripheral blood of CL patients. To study the role of Dectin-1+ DCs in adaptive immunity against L. major, we analyzed the T cell stimulating potential of bone marrow-derived dendritic cells (BMDCs) in the presence of the Dectin-1 agonist Curdlan. These experiments revealed that Curdlan induces the maturation of BMDCs and the expansion of Leishmania-specific CD4+ T cells. Based on these findings, we evaluated the impact of Curdlan/Dectin-1 interactions in experimental leishmaniasis and were able to demonstrate that the presence of Curdlan at the site of infection modulates the course of disease in BALB/c mice: wild-type BALB/c mice treated intradermally with Curdlan developed a protective immune response against L. major whereas Dectin-1−/− BALB/c mice still developed the fatal course of disease after Curdlan treatment. Furthermore

Leishmania proteophosphoglycans regurgitated from infected sand flies accelerate dermal wound repair and exacerbate leishmaniasis via insulin-like growth factor 1-dependent signalling

PubMed Central

Giraud, Emilie; Martin, Oihane; Dillon, Rod J.; Műller, Ingrid

2018-01-01

Leishmania parasites are transmitted to vertebrate hosts by female phlebotomine sand flies as they bloodfeed by lacerating the upper capillaries of the dermis with their barbed mouthparts. In the sand fly midgut secreted proteophosphoglycans from Leishmania form a biological plug known as the promastigote secretory gel (PSG), which blocks the gut and facilitates the regurgitation of infective parasites. The interaction between the wound created by the sand fly bite and PSG is not known. Here we nanoinjected a sand fly egested dose of PSG into BALB/c mouse skin that lead to the differential expression of 7,907 transcripts. These transcripts were transiently up-regulated during the first 6 hours post-wound and enriched for pathways involved in inflammation, cell proliferation, fibrosis, epithelial cell differentiation and wound remodelling. We found that PSG significantly accelerated wound healing in vitro and in mice; which was associated with an early up-regulation of transcripts involved in inflammation (IL-1β, IL-6, IL-10, TNFα) and inflammatory cell recruitment (CCL2, CCL3, CCL4, CXCL2), followed 6 days later by enhanced expression of transcripts associated with epithelial cell proliferation, fibroplasia and fibrosis (FGFR2, EGF, EGFR, IGF1). Dermal expression of IGF1 was enhanced following an infected sand fly bite and was acutely responsive to the deposition of PSG but not the inoculation of parasites or sand fly saliva. Antibody blockade of IGF1 ablated the gel’s ability to promote wound closure in mouse ears and significantly reduced the virulence of Leishmania mexicana infection delivered by an individual sand fly bite. Dermal macrophages recruited to air-pouches on the backs of mice revealed that IGF1 was pivotal to the PSG’s ability to promote macrophage alternative activation and Leishmania infection. Our data demonstrate that through the regurgitation of PSG Leishmania exploit the wound healing response of the host to the vector bite by promoting

Effects of nitro-heterocyclic derivatives against Leishmania (Leishmania) infantum promastigotes and intracellular amastigotes.

PubMed

Petri e Silva, Simone Carolina Soares; Palace-Berl, Fanny; Tavares, Leoberto Costa; Soares, Sandra Regina Castro; Lindoso, José Angelo Lauletta

2016-04-01

Leishmaniasis is an overlooked tropical disease affecting approximately 1 million people in several countries. Clinical manifestation depends on the interaction between Leishmania and the host's immune response. Currently available treatment options for leishmaniasis are limited and induce severe side effects. In this research, we tested nitro-heterocyclic compounds (BSF series) as a new alternative against Leishmania. Its activity was measured in Leishmania (Leishmania) infantum promastigotes and intracellular amastigotes using MTT colorimetric assay. Additionally, we assessed the phosphatidylserine exposure by promastigotes, measured by flow cytometry, as well as nitric oxide production, measured by Griess' method. The nitro-heterocyclic compounds (BSF series) showed activity against L. (L.) infantum promastigotes, inducting the phosphatidylserine exposition by promastigotes, decreasing intracellular amastigotes and increasing oxide nitric production. The selectivity index was more prominent to Leishmania than to macrophages. Compared to amphotericin b, our compounds presented higher IC50, however the selectivity index was more specific to parasite than to amphotericin b. In conclusion, these nitro-heterocyclic compounds showed to be promising as an anti-Leishmania drug, in in vitro studies. Copyright © 2016 Elsevier Inc. All rights reserved.

Recent developments in leishmaniasis vaccine delivery systems.

PubMed

Bhowmick, Sudipta; Ali, Nahid

2008-07-01

The observation that recovery from infection with Leishmania confers immunity to reinfection suggests that control of leishmaniasis by vaccination may be possible. New generation vaccines, particularly those based on recombinant proteins and DNA, are found to be less immunogenic. There is an urgent need for the development of new and improved vaccine adjuvants. Based on their principal mechanisms of action, adjuvants can be broadly separated into two classes: immunostimulatory adjuvants and vaccine delivery systems. Vaccine delivery systems can carry both antigen and adjuvant for effective delivery to the antigen-presenting cells (APCs). In this article, we review the adjuvants, the delivery systems and their combinations used in the search of an effective vaccine against leishmaniasis. Based on current knowledge, cationic liposomes appear to have better prospects as effective delivery systems for developing a vaccine for leishmaniasis.

Histological study of cell migration in the dermis of hamsters after immunisation with two different vaccines against visceral leishmaniasis.

PubMed

Moreira, Nádia das Dores; Giunchetti, Rodolfo Cordeiro; Carneiro, Cláudia Martins; Vitoriano-Souza, Juliana; Roatt, Bruno Mendes; Malaquias, Luiz Cosme Cotta; Corrêa-Oliveira, Rodrigo; Reis, Alexandre Barbosa

2009-04-15

Vaccine candidates, including live and/or killed parasites, Leishmania-purified fractions, defined recombinant antigens and antigen-encoding DNA-plasmids have been proposed to use as vaccine anti-Leishmania. More recently, the hamsters have been used to pre-selection of antigens candidate to apply in further experiments using canine model. In this report we evaluated the kinetics of cell migration in dermal inflammatory infiltrate, circulating leukocytes and the presence of nitric oxide (NO)/induced nitric oxide synthase during the early (1-24h) and late (48-168h) periods following inoculation of hamsters with antigenic components of anti-canine visceral leishmaniasis vaccines Leishmune and Leishmania braziliensis antigen (LB) with and without saponin (Sap) adjuvant. Our results show that LB caused an early reduction of lymphocytes in the dermis while Sap and LBSap triggered a late recruitment, suggesting the role of the adjuvant in the traffic of antigen-presenting cells and the induction of lymphocyte migration. In that manner our results suggest that the kinetics of cell migration on hamster model may be of value in the selection of vaccine antigens prior the tests in dogs particularly in respect of the toxicity of the preparations.

Infectivity of five different types of macrophages by Leishmania infantum.

PubMed

Maia, C; Rolão, N; Nunes, M; Gonçalves, L; Campino, L

2007-08-01

Leishmania are intracellular parasites that multiply as the amastigote form in the macrophages of their vertebrate hosts. Since vaccines against leishmaniases are still under development, the control of these diseases relies on prompt diagnosis and chemotherapy in infected humans as well as in dogs, which are the main reservoir of Leishmania infantum, in Mediterranean countries. To establish the macrophage type to be used as an in vitro model for antileishmanial chemotherapeutic studies, we analysed the susceptibility of human peripheral blood derived macrophages, macrophages derived from mouse bone marrow, mouse peritoneal macrophages and macrophages differentiated from cell lines U-937 and DH82 to infection by two L. infantum strains, one obtained from a human leishmanial infection and other from a canine infection. Both strains displayed comparable behaviour in their capacity of infecting the different macrophage types. Human peripheral blood macrophages and DH82 cells were less infectable by both strains. U-937, mouse peritoneal macrophages and mouse bone marrow derived macrophages are the most active cells to phagocytose the parasites. However, U-937 cell line appears to be the most useful as Leishmania infection model providing an unlimited source of homogeneous host cells with reproducibility of the results, is less time consuming, less expensive and tolerate high doses of first line drugs for human and canine visceral leishmaniasis treatment.

Detection and Characterization of Leishmania (Leishmania) and Leishmania (Viannia) by SYBR Green-Based Real-Time PCR and High Resolution Melt Analysis Targeting Kinetoplast Minicircle DNA

PubMed Central

Ceccarelli, Marcello; Galluzzi, Luca; Migliazzo, Antonella; Magnani, Mauro

2014-01-01

Leishmaniasis is a neglected disease with a broad clinical spectrum which includes asymptomatic infection. A thorough diagnosis, able to distinguish and quantify Leishmania parasites in a clinical sample, constitutes a key step in choosing an appropriate therapy, making an accurate prognosis and performing epidemiological studies. Several molecular techniques have been shown to be effective in the diagnosis of leishmaniasis. In particular, a number of PCR methods have been developed on various target DNA sequences including kinetoplast minicircle constant regions. The first aim of this study was to develop a SYBR green-based qPCR assay for Leishmania (Leishmania) infantum detection and quantification, using kinetoplast minicircle constant region as target. To this end, two assays were compared: the first used previously published primer pairs (qPCR1), whereas the second used a nested primer pairs generating a shorter PCR product (qPCR2). The second aim of this study was to evaluate the possibility to discriminate among subgenera Leishmania (Leishmania) and Leishmania (Viannia) using the qPCR2 assay followed by melting or High Resolution Melt (HRM) analysis. Both assays used in this study showed good sensitivity and specificity, and a good correlation with standard IFAT methods in 62 canine clinical samples. However, the qPCR2 assay allowed to discriminate between Leishmania (Leishmania) and Leishmania (Viannia) subgenera through melting or HRM analysis. In addition to developing assays, we investigated the number and genetic variability of kinetoplast minicircles in the Leishmania (L.) infantum WHO international reference strain (MHOM/TN/80/IPT1), highlighting the presence of minicircle subclasses and sequence heterogeneity. Specifically, the kinetoplast minicircle number per cell was estimated to be 26,566±1,192, while the subclass of minicircles amplifiable by qPCR2 was estimated to be 1,263±115. This heterogeneity, also observed in canine clinical samples

The Leishmania infantum PUF proteins are targets of the humoral response during visceral leishmaniasis

PubMed Central

2010-01-01

Background RNA-binding proteins of the PUF family share a conserved domain consisting of tandemly repeated 36-40 amino acid motifs (typically eight) known as Puf repeats. Proteins containing tandem repeats are often dominant targets of humoral responses during infectious diseases. Thus, we considered of interest to analyze whether Leishmania PUF proteins result antigenic during visceral leishmaniasis (VL). Findings Here, employing whole-genome databases, we report the composition, and structural features, of the PUF family in Leishmania infantum. Additionally, the 10 genes of the L. infantum PUF family were cloned and used to express the Leishmania PUFs in bacteria as recombinant proteins. Finally, the antigenicity of these PUF proteins was evaluated by determining levels of specific antibodies in sera from experimentally infected hamsters. The Leishmania PUFs were all recognized by the sera, even though with different degree of reactivity and/or frequency of recognition. The reactivity of hamster sera against recombinant LiPUF1 and LiPUF2 was particularly prominent, and these proteins were subsequently assayed against sera from human patients. High antibody responses against rLiPUF1 and rLiPUF2 were found in sera from VL patients, but these proteins resulted also recognized by sera from Chagas' disease patients. Conclusion Our results suggest that Leishmania PUFs are targets of the humoral response during L. infantum infection and may represent candidates for serodiagnosis and/or vaccine reagents; however, it should be kept in mind the cross-reactivity of LiPUFs with antibodies induced against other trypanosomatids such as Trypanosoma cruzi. PMID:20180988

Identification of Proteins in Promastigote and Amastigote-like Leishmania Using an Immunoproteomic Approach

PubMed Central

Coelho, Vinicio T. S.; Oliveira, Jamil S.; Valadares, Diogo G.; Chávez-Fumagalli, Miguel A.; Duarte, Mariana C.; Lage, Paula S.; Soto, Manuel; Santoro, Marcelo M.; Tavares, Carlos A. P.; Fernandes, Ana Paula; Coelho, Eduardo A. F.

2012-01-01

Background The present study aims to identify antigens in protein extracts of promastigote and amastigote-like Leishmania (Leishmania) chagasi syn. L. (L.) infantum recognized by antibodies present in the sera of dogs with asymptomatic and symptomatic visceral leishmaniasis (VL). Methodology/Principal Findings Proteins recognized by sera samples were separated by two-dimensional electrophoresis (2DE) and identified by mass spectrometry. A total of 550 spots were observed in the 2DE gels, and approximately 104 proteins were identified. Several stage-specific proteins could be identified by either or both classes of sera, including, as expected, previously known proteins identified as diagnosis, virulence factors, drug targets, or vaccine candidates. Three, seven, and five hypothetical proteins could be identified in promastigote antigenic extracts; while two, eleven, and three hypothetical proteins could be identified in amastigote-like antigenic extracts by asymptomatic and symptomatic sera, as well as a combination of both, respectively. Conclusions/Significance The present study represents a significant contribution not only in identifying stage-specific L. infantum molecules, but also in revealing the expression of a large number of hypothetical proteins. Moreover, when combined, the identified proteins constitute a significant source of information for the improvement of diagnostic tools and/or vaccine development to VL. PMID:22272364

Multicomponent LBSap vaccine displays immunological and parasitological profiles similar to those of Leish-Tec® and Leishmune® vaccines against visceral leishmaniasis.

PubMed

de Mendonça, Ludmila Zanandreis; Resende, Lucilene Aparecida; Lanna, Mariana Ferreira; Aguiar-Soares, Rodrigo Dian de Oliveira; Roatt, Bruno Mendes; Castro, Renata Alves de Oliveira E; Batista, Maurício Azevedo; Silveira-Lemos, Denise; Gomes, Juliana de Assis Silva; Fujiwara, Ricardo Toshio; Rezende, Simone Aparecida; Martins-Filho, Olindo Assis; Corrêa-Oliveira, Rodrigo; Dutra, Walderez Ornelas; Reis, Alexandre Barbosa; Giunchetti, Rodolfo Cordeiro

2016-08-30

In past years, many researchers have sought canine visceral leishmaniasis (CVL) prevention through the characterization of Leishmania antigens as vaccine candidates. Despite these efforts, there is still no efficient vaccine for CVL control. In the present study, we performed a pre-clinical vaccine trial using BALB/c mice to compare the effects of the multicomponent LBSap vaccine with those of Leish-Tec® and Leishmune®. Blood was collected to determine the frequency of peripheral blood cells and to evaluate hematologic and immunophenotypic parameters. Liver and spleen samples were collected for parasitological quantification, and spleen samples were used to access the cytokine profile. When measuring total IgG and IgG1 anti-Leishmania levels after the third vaccination and L. infantum challenge, it was evident that all vaccines were able to induce humoral immune response. Regarding the innate immune response, increased levels of NK CD3(-)CD49(+) cells were the hallmark of all vaccinated groups, whereas only the Leish-Tec® group displayed a high frequency of CD14(+) monocytes after L. infantum challenge. Moreover, CD3(+)CD4(+) T cells were the main circulating lymphocytes induced after L. infantum challenge with all evaluated vaccines. Importantly, after L. infantum challenge, splenocytes from the Leishmune® vaccine produced high levels of IL-2, whereas a prominent type 1 immune response was the hallmark of the LBSap vaccine, which presented high levels of IL-2, IL-6, TNF-α, and IFN-γ. The efficacy analysis using real-time polymerase chain reaction demonstrated a reduction in the parasitism in the spleen (Leishmune®: 64 %; LBSap: 42 %; and Leish-Tec®: 36 %) and liver (Leishmune®: 71 %; LBSap: 62 %; and Leish-Tec®: 48 %). The dataset led to the conclusion that the LBSap vaccination was able to induce immune and efficacy profiles comparable with those of commercial vaccines, thus demonstrating its potential as a promising vaccine candidate for visceral

A Chimera Containing CD4+ and CD8+ T-Cell Epitopes of the Leishmania donovani Nucleoside Hydrolase (NH36) Optimizes Cross-Protection against Leishmania amazonesis Infection.

PubMed

Alves-Silva, Marcus Vinícius; Nico, Dirlei; Morrot, Alexandre; Palatnik, Marcos; Palatnik-de-Sousa, Clarisa B

2017-01-01

The Leishmania donovani nucleoside hydrolase (NH36) and NH A34480 of Leishmania amazonensis share 93% of sequence identity. In mice, the NH36 induced protection against visceral leishmaniasis is mediated by a CD4+ T cell response against its C-terminal domain (F3). Besides this CD4+ Th1 response, prevention and cure of L. amazonensis infection require also additional CD8+ and regulatory T-cell responses to the NH36 N-terminal (F1 domain). We investigated if mice vaccination with F1 and F3 domains cloned in tandem, in a recombinant chimera, with saponin, optimizes the vaccine efficacy against L. amazonensis infection above the levels promoted by the two admixed domains or by each domain independently. The chimera induced the highest IgA, IgG, and IgG2a anti-NH36 antibody, IDR, IFN-γ, and IL-10 responses, while TNF-α was more secreted by mice vaccinated with F3 or all F3-contaning vaccines. Additionally, the chimera and the F1 vaccine also induced the highest proportions of CD4+ and CD8+ T cells secreting IL-2, TNF-α, or IFN-γ alone, TNF-α in combination with IL-2 or IFN-γ, and of CD4+ multifunctional cells secreting IL-2, TNF-α, and IFN-γ. Correlating with the immunological results, the strongest reductions of skin lesions sizes were determined by the admixed domains (80%) and by the chimera (84%), which also promoted the most pronounced and significant reduction of the parasite load (99.8%). Thus, the epitope presentation in a recombinant chimera optimizes immunogenicity and efficacy above the levels induced by the independent or admixed F1 and F3 domains. The multiparameter analysis disclosed that the Th1-CD4+ T helper response induced by the chimera is mainly directed against its FRYPRPKHCHTQVA epitope. Additionally, the YPPEFKTKL epitope of F1 induced the second most important CD4+ T cell response, and, followed by the DVAGIVGVPVAAGCT, FMLQILDFYTKVYE, and ELLAITTVVGNQ sequences, also the most potent CD8+ T cell responses and IL-10 secretion. Remarkably

A Chimera Containing CD4+ and CD8+ T-Cell Epitopes of the Leishmania donovani Nucleoside Hydrolase (NH36) Optimizes Cross-Protection against Leishmania amazonesis Infection

PubMed Central

Alves-Silva, Marcus Vinícius; Nico, Dirlei; Morrot, Alexandre; Palatnik, Marcos; Palatnik-de-Sousa, Clarisa B.

2017-01-01

The Leishmania donovani nucleoside hydrolase (NH36) and NH A34480 of Leishmania amazonensis share 93% of sequence identity. In mice, the NH36 induced protection against visceral leishmaniasis is mediated by a CD4+ T cell response against its C-terminal domain (F3). Besides this CD4+ Th1 response, prevention and cure of L. amazonensis infection require also additional CD8+ and regulatory T-cell responses to the NH36 N-terminal (F1 domain). We investigated if mice vaccination with F1 and F3 domains cloned in tandem, in a recombinant chimera, with saponin, optimizes the vaccine efficacy against L. amazonensis infection above the levels promoted by the two admixed domains or by each domain independently. The chimera induced the highest IgA, IgG, and IgG2a anti-NH36 antibody, IDR, IFN-γ, and IL-10 responses, while TNF-α was more secreted by mice vaccinated with F3 or all F3-contaning vaccines. Additionally, the chimera and the F1 vaccine also induced the highest proportions of CD4+ and CD8+ T cells secreting IL-2, TNF-α, or IFN-γ alone, TNF-α in combination with IL-2 or IFN-γ, and of CD4+ multifunctional cells secreting IL-2, TNF-α, and IFN-γ. Correlating with the immunological results, the strongest reductions of skin lesions sizes were determined by the admixed domains (80%) and by the chimera (84%), which also promoted the most pronounced and significant reduction of the parasite load (99.8%). Thus, the epitope presentation in a recombinant chimera optimizes immunogenicity and efficacy above the levels induced by the independent or admixed F1 and F3 domains. The multiparameter analysis disclosed that the Th1-CD4+ T helper response induced by the chimera is mainly directed against its FRYPRPKHCHTQVA epitope. Additionally, the YPPEFKTKL epitope of F1 induced the second most important CD4+ T cell response, and, followed by the DVAGIVGVPVAAGCT, FMLQILDFYTKVYE, and ELLAITTVVGNQ sequences, also the most potent CD8+ T cell responses and IL-10 secretion. Remarkably

Alum-precipitated autoclaved Leishmania major plus bacille Calmette-Guérrin, a candidate vaccine for visceral leishmaniasis: safety, skin-delayed type hypersensitivity response and dose finding in healthy volunteers.

PubMed

Kamil, A A; Khalil, E A G; Musa, A M; Modabber, F; Mukhtar, M M; Ibrahim, M E; Zijlstra, E E; Sacks, D; Smith, P G; Zicker, F; El-Hassan, A M

2003-01-01

In a previous efficacy study, autoclaved Leishmania major (ALM) + bacille Calmette-Guérrin (BCG) vaccine was shown to be safe, but not superior to BCG alone, in protecting against visceral leishmaniasis. From June 1999 to June 2000, we studied the safety and immunogenicity of different doses of alum-precipitated ALM + BCG vaccine mixture administered intradermally to evaluate whether the addition of alum improved the immunogenicity of ALM. Twenty-four healthy adult volunteers were recruited and sequentially allocated to receive either 10 microg, 100 microg, 200 microg, or 400 microg of leishmanial protein in the alum-precipitated ALM + BCG vaccine mixture. Side effects were minimal for all doses and confined to the site of injection. All volunteers in the 10 microg, 100 microg, and 400 microg groups had a leishmanin skin test (LST) reaction of > or = 5 mm by day 42 and this response was maintained when tested after 90 d. Only 1 volunteer out of 5 in the 200 microg group had a LST reaction of > or = 5 mm by day 42 and the reasons for the different LST responses in this group are unclear. This is the first time that an alum adjuvant with ALM has been in used in humans and the vaccine mixture was safe and induced a strong delayed type hypersensitivity (DTH) reaction in the study volunteers. On the basis of this study we suggest that 100 1 microg of leishmanial protein in the vaccine mixture is a suitable dose for future efficacy studies, as it induced the strongest DTH reaction following vaccination.

Leishmune vaccine: the newest tool for prevention and control of canine visceral leishmaniosis and its potential as a transmission-blocking vaccine.

PubMed

Dantas-Torres, Filipe

2006-10-10

Canine visceral leishmaniosis is a life-threatening disease caused by Leishmania infantum. For quite some time, specialists in leishmaniosis have tried to develop more affordable and effective control measures against this disease. In this search, the first vaccine against canine visceral leishmaniosis was recently licensed in Brazil. In the light of recent research, the Leishmune vaccine might be seen as the newest tool for prevention and control of canine visceral leishmaniosis. Moreover, the potential of the Leishmune as a transmission-blocking vaccine has recently been demonstrated, indicating its usefulness in the control of zoonotic visceral leishmaniosis.

Selective effects of Euterpe oleracea (açai) on Leishmania (Leishmania) amazonensis and Leishmania infantum.

PubMed

Da Silva, Bruno José Martins; Souza-Monteiro, José Rogério; Rogez, Herve; Crespo-López, Maria Elena; Do Nascimento, Jose Luiz M; Silva, Edilene O

2018-01-01

Leishmania (Leishmania) amazonensis and Leishmania infantum (=Leishmania chagasi) are protozoa that cause American cutaneous and visceral leishmaniasis, respectively. These diseases show a high incidence in developing countries such as Brazil. The treatments used for leishmaniasis are still limited due to their high cost and toxicity. Currently, some natural products are considered an important alternative source of new leishmanicidal agents. Euterpe oleracea Martius, a palm producing black fruits, is frequently consumed in the Amazon region, as a juice, known as açai, with potent antioxidant, anti-inflammatory and anticonvulsant properties. Interestingly, the biological activity of clarified açai juice (EO) on L. (L.) amazonensis and L. infantum (=L. chagasi) is unknown. Therefore, the mechanism of anti-leishmanial action of EO has been evaluated on L. (L.) amazonensis and L. infantum (=L. chagasi). EO reduced the number of promastigotes and caused morphological alterations, increased the production of reactive oxygen species (ROS) and induced cell death phenotypes probably seems by apoptosis in the promastigotes of L. (L.) amazonensis (IC 50  = 1:40) and L. infantum (=L. chagasi) (IC 50  = 1:38). EO also presented activity against Leishmania amastigotes. Treatment with EO for 72 h strongly reduced IL-17 cytokine levels at all tested concentrations and decreased the number of intracellular amastigotes in macrophages infected with L. (L.) amazonensis (IC 50  = 1:30) and L. infantum (=L. chagasi) (IC 50  = 1:38). Additionally, no cytotoxic effect was observed in murine macrophages treated with EO (72 h - CC 50  > 1:1). Our results demonstrated that EO has leishmanicidal activity against two different species that cause American visceral and cutaneous leishmaniasis without cytotoxic effects for the host cell. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

[Development of new vaccines].

PubMed

González-Romo, Fernando; Picazo, Juan J

2015-10-01

Recent and important advances in the fields of immunology, genomics, functional genomics, immunogenetics, immunogenomics, bioinformatics, microbiology, genetic engineering, systems biology, synthetic biochemistry, proteomics, metabolomics and nanotechnology, among others, have led to new approaches in the development of vaccines. The better identification of ideal epitopes, the strengthening of the immune response due to new adjuvants, and the search of new routes of vaccine administration, are good examples of advances that are already a reality and that will favour the development of more vaccines, their use in indicated population groups, or its production at a lower cost. There are currently more than 130 vaccines are under development against the more wished (malaria or HIV), difficult to get (CMV or RSV), severe re-emerging (Dengue or Ebola), increasing importance (Chagas disease or Leishmania), and nosocomial emerging (Clostridium difficile or Staphylococcus aureus) infectious diseases. Copyright © 2015. Published by Elsevier España, S.L.U.

Leishmanization revisited: immunization with a naturally attenuated cutaneous Leishmania donovani isolate from Sri Lanka protects against visceral leishmaniasis.

PubMed

McCall, Laura-Isobel; Zhang, Wen-Wei; Ranasinghe, Shanlindra; Matlashewski, Greg

2013-02-27

Leishmaniasis is a neglected tropical disease caused by Leishmania protozoa and associated with three main clinical presentations: cutaneous, mucocutaneous and visceral leishmaniasis. Visceral leishmaniasis is the second most lethal parasitic disease after malaria and there is so far no human vaccine. Leishmania donovani is a causative agent of visceral leishmaniasis in South East Asia and Eastern Africa. However, in Sri Lanka, L. donovani causes mainly cutaneous leishmaniasis, while visceral leishmaniasis is rare. We investigate here the possibility that the cutaneous form of L. donovani can provide immunological protection against the visceral form of the disease, as a potential explanation for why visceral leishmaniasis is rare in Sri Lanka. Subcutaneous immunization with a cutaneous clinical isolate from Sri Lanka was significantly protective against visceral leishmaniasis in BALB/c mice. Protection was associated with a mixed Th1/Th2 response. These results provide a possible rationale for the scarcity of visceral leishmaniasis in Sri Lanka and could guide leishmaniasis vaccine development efforts. Copyright © 2012 Elsevier Ltd. All rights reserved.

Detection of Leishmania in Unaffected Mucosal Tissues of Patients with Cutaneous Leishmaniasis Caused by Leishmania (Viannia) Species

PubMed Central

Figueroa, Roger Adrian; Lozano, Leyder Elena; Romero, Ibeth Cristina; Cardona, Maria Teresa; Prager, Martin; Pacheco, Robinson; Diaz, Yira Rosalba; Tellez, Jair Alexander; Saravia, Nancy Gore

2016-01-01

Background Leishmania (Viannia) species are the principal cause of mucosal leishmaniasis. The natural history and pathogenesis of mucosal disease are enigmatic. Parasitological evaluation of mucosal tissues has been constrained by the invasiveness of conventional sampling methods. Methods We evaluated the presence ofLeishmania in the mucosa of 26 patients with cutaneous leishmaniasis and 2 patients with mucocutaneous leishmaniasis. Swab samples of the nasal mucosa, tonsils, and conjunctiva were analyzed using polymerase chain reaction with LV-B1 primers and Southern blot hybridization. Results Two patients with mucocutaneous leishmaniasis and 21 (81%) of 26 patients with cutaneous leishmaniasis had Leishmania kinetoplast minicircle DNA (kDNA) in mucosal tissues. kDNA was amplified from swab samples of nasal mucosa from 14 (58%) of 24 patients, tonsils from 13 (46%) of 28 patients, and conjunctiva from 6 (25%) of 24 patients. kDNA was detected in the mucosa of patients with cutaneous disease caused by Leishmania panamensis, Leishmania guyanensis, and Leishmania braziliensis. Conclusion The asymptomatic presence of parasites in mucosal tissues may be common in patients with Leishmania (Viannia) infection. PMID:19569974

First evidence of Leishmania infection in European brown hare (Lepus europaeus) in Greece: GIS analysis and phylogenetic position within the Leishmania spp.

PubMed

Tsokana, C N; Sokos, C; Giannakopoulos, A; Mamuris, Z; Birtsas, P; Papaspyropoulos, K; Valiakos, G; Spyrou, V; Lefkaditis, M; Chatzopoulos, D C; Kantere, M; Manolakou, K; Touloudi, A; Burriel, A Rodi; Ferroglio, E; Hadjichristodoulou, C; Billinis, C

2016-01-01

Although the existence of a sylvatic transmission cycle of Leishmania spp., independent from the domestic cycle, has been proposed, data are scarce on Leishmania infection in wild mammals in Greece. In this study, we aimed to investigate the presence of Leishmania infection in the European brown hare in Greece, to infer the phylogenetic position of the Leishmania parasites detected in hares in Greece, and to identify any possible correlation between Leishmania infection in hares with environmental parameters, using the geographical information system (GIS). Spleen samples from 166 hares were tested by internal transcribed spacer-1 (ITS-1)-nested PCR for the detection of Leishmania DNA. Phylogenetic analysis was performed on Leishmania sequences from hares in Greece in conjunction with Leishmania sequences from dogs in Greece and 46 Leishmania sequences retrieved from GenBank. The Leishmania DNA prevalence in hares was found to be 23.49 % (95 % confidence interval (CI) 17.27-30.69). The phylogenetic analysis confirmed that the Leishmania sequences from hares in Greece belong in the Leishmania donovani complex. The widespread Leishmania infection in hares should be taken into consideration because under specific circumstances, this species can act as a reservoir host. This study suggests that the role of wild animals, including hares, in the epidemiology of Leishmania spp. in Greece deserves further elucidation.

Performance of LBSap Vaccine after Intradermal Challenge with L. infantum and Saliva of Lu. longipalpis: Immunogenicity and Parasitological Evaluation

PubMed Central

Roatt, Bruno Mendes; Aguiar-Soares, Rodrigo Dian de Oliveira; Vitoriano-Souza, Juliana; Coura-Vital, Wendel; Braga, Samuel Leôncio; Corrêa-Oliveira, Rodrigo; Martins-Filho, Olindo Assis; Teixeira-Carvalho, Andréa; de Lana, Marta; Gontijo, Nelder Figueiredo; Marques, Marcos José; Giunchetti, Rodolfo Cordeiro; Reis, Alexandre Barbosa

2012-01-01

In the last decade, the search for new vaccines against canine visceral leishmaniasis has intensified. However, the pattern related to immune protection during long periods after experimental infection in vaccine trials is still not fully understood. Herein, we investigated the immunogenicity and parasitological levels after intradermal challenge with Leishmania infantum plus salivary gland extract in dogs immunized with a vaccine composed of L. braziliensis antigens plus saponin as an adjuvant (LBSap vaccine). The LBSap vaccine elicited higher levels of total anti-Leishmania IgG as well as both IgG1 and IgG2. Furthermore, dogs vaccinated had increased levels of lymphocytes, particularly circulating B cells (CD21+) and both CD4+ and CD8+ T lymphocytes. LBSap also elicited an intense in vitro cell proliferation associated with higher levels of CD4+ T lymphocytes specific for vaccine soluble antigen and soluble lysate of L. infantum antigen even 885 days after experimental challenge. Furthermore, LBSap vaccinated dogs presented high IFN-γ and low IL-10 and TGF-β1 expression in spleen with significant reduction of parasite load in this tissue. Overall, our results validate the potential of LBSap vaccine to protect against L. infantum experimental infection and strongly support further evaluation of efficiency of LBSap against CVL in natural infection conditions. PMID:23189161

Direct detection of Leishmania from clinical samples.

PubMed

Waitumbi, John N; Bast, Joshua; Nyakoe, Nancy; Magiri, Charles; Quintana, Miguel; Takhampunya, Ratree; Schuster, Anthony L; Van de Wyngaerde, Marshall T; McAvin, James C; Coleman, Russell E

2017-01-01

The ability to rapidly and accurately diagnose leishmaniasis is a military priority. Testing was conducted to evaluate diagnostic sensitivity and specificity of field-expedient Leishmania genus and visceral Leishmania specific dual-fluorogenic, hydrolysis probe (TaqMan), polymerase chain reaction assays previously established for use in vector surveillance. Blood samples of patients with confirmed visceral leishmaniasis and controls without the disease from Baringo District, Kenya, were tested. Leishmania genus assay sensitivity was 100% (14/14) and specificity was 84% (16/19). Visceral Leishmania assay sensitivity was 93% (13/14) and specificity 80% (4/5). Cutaneous leishmaniasis (CL) skin scrapes of patients from Honduras were also evaluated. Leishmania genus assay sensitivity was 100% (10/10). Visceral Leishmania assay specificity was 100% (10/10) from cutaneous leishmaniasis samples; no fluorescence above background was reported. These results show promise in a rapid, sensitive, and specific method for Leishmania direct detection from clinical samples.

Development of Leishmania vaccines in the era of visceral leishmaniasis elimination.

PubMed

Engwerda, Christian R; Matlashewski, Greg

2015-07-01

A visceral leishmaniasis (VL) elimination target set for the Indian subcontinent in 2005 is being met in many endemic areas without a vaccine. This begs a question: is a VL vaccine needed if elimination targets can be met with current control programs? Here, we argue that a vaccine will be critical if the success of recent VL control efforts are to be sustained. However, not only do we require a safe and effective vaccine, but we also need to know how this should be used for maximum impact. In particular, identifying appropriate target populations to vaccinate will be crucial. © The Author 2015. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Canine leishmaniosis caused by Leishmania major and Leishmania tropica: comparative findings and serology.

PubMed

Baneth, Gad; Yasur-Landau, Daniel; Gilad, Matan; Nachum-Biala, Yaarit

2017-03-13

Infection and clinical disease associated with Leishmania major and Leishmania tropica, two common agents of human cutaneous leishmaniosis, have rarely been reported in dogs. This study describes dogs infected with these Leishmania spp. prevalent in the Middle East and North Africa, and compares the serological response of dogs infected with Leishmania infantum, L. major or L. tropica to whole promastigote antigen enzyme-linked immunosorbent assay (ELISA) of each species and to rK39 dipstick. Leishmania major infection in a 5-month-old male dog was associated with alopecic and ulcerative periocular and limb skin lesions which responded to allopurinol treatment. Infection was detected by skin and blood polymerase chain reaction (PCR) and confirmed by DNA sequencing but the dog was seronegative. Leishmania tropica infection was detected in a 3-month-old female dog co-infected with Babesia vogeli and Anaplasma platys and with no skin lesions. PCR and DNA sequencing of the blood and parasite culture were positive for L. tropica. Sera from 11 dogs infected with L. infantum, L. major or L. tropica were reactive with all three Leishmania spp. antigens except for sera from a dog with L. major infection. No significant differences were found between reactivity of dog sera to the antigen of the infecting species, or to the other Leishmania spp. antigens. Sera from dogs infected with L. infantum and L. tropica were positive with the rK39 antigen kit, while dogs with L. major infection were seronegative. Skin lesions in L. major infected dogs from this study and previous reports (n = 2) were ulcerative and located on the muzzle, feet and foot pads and not associated with generalized lymphadenomegaly and splenomegaly. In previous L. tropica infections, skin lesions were proliferative mucocutaneous in young dogs (n = 2), or associated with widespread dermatitis, lymphadenomegaly and splenomegaly in older dogs with similarity to L. infantum infection (n = 2). This

Effects of written action plan adherence on COPD exacerbation recovery.

PubMed

Bischoff, Erik W M A; Hamd, Dina H; Sedeno, Maria; Benedetti, Andrea; Schermer, Tjard R J; Bernard, Sarah; Maltais, François; Bourbeau, Jean

2011-01-01

The effects of written action plans on recovery from exacerbations of chronic obstructive pulmonary disease (COPD) have not been well studied. The aims of this study were to assess the effects of adherence to a written action plan on exacerbation recovery time and unscheduled healthcare utilisation and to explore factors associated with action plan adherence. This was a 1-year prospective cohort study embedded in a randomised controlled trial. Exacerbation data were recorded for 252 patients with COPD who received a written action plan for prompt treatment of exacerbations with the instructions to initiate standing prescriptions for both antibiotics and prednisone within 3 days of exacerbation onset. Following the instructions was defined as adherence to the action plan. From the 288 exacerbations reported by 143 patients, start dates of antibiotics or prednisone were provided in 217 exacerbations reported by 119 patients (53.8% male, mean age 65.4 years, post-bronchodilator forced expiratory volume in 1 s (FEV(1)) 43.9% predicted). In 40.1% of exacerbations, patients adhered to their written action plan. Adherence reduced exacerbation recovery time with statistical (p=0.0001) and clinical (-5.8 days) significance, but did not affect unscheduled healthcare utilisation (OR 0.94, 95% CI 0.49 to 1.83). Factors associated with an increased likelihood of adherence were influenza vaccination, cardiac comorbidity, younger age and lower FEV(1) as percentage predicted. This study shows that adherence to a written action plan is associated with a reduction in exacerbation recovery time by prompt treatment. Knowing the factors that are associated with proper and prompt utilisation of an action plan permits healthcare professionals to better focus their self-management support on appropriate patients.

First evidence of autochthonous cases of Leishmania (Leishmania) infantum in horse (Equus caballus) in the Americas and mixed infection of Leishmania infantum and Leishmania (Viannia) braziliensis.

PubMed

Soares, Isabel R; Silva, Soraia O; Moreira, Filipe Moraghi; Prado, Luan Gavião; Fantini, Priscila; Maranhão, Renata de Pino Albuquerque; da Silva Filho, José Monteiro; Melo, Maria Norma; Palhares, Maristela S

2013-11-08

This study reports the first evidence of infection by Leishmania infantum in Equus caballus in Americas and the first mixed infection of L. infantum/Leishmania braziliensis on this mammalian species in the world. The diagnoses was based on presence of parasites in lesions and bone marrow aspirates, their identification by using specific primers for L. infantum and L. braziliensis complexes and also serological methods IFAT and ELISA. The analysis of the PCR products suggested mixed infection in three animals. Further studies involving equine leishmaniasis are carrying out in order to clarify the dynamic of Leishmania sp. in this mammalian specie and their role in the transmission of those parasites in urban endemic area of Belo Horizonte, Minas Gerais State, Brazil. Copyright © 2013 Elsevier B.V. All rights reserved.

Peptide Vaccines for Leishmaniasis.

PubMed

De Brito, Rory C F; Cardoso, Jamille M De O; Reis, Levi E S; Vieira, Joao F; Mathias, Fernando A S; Roatt, Bruno M; Aguiar-Soares, Rodrigo Dian D O; Ruiz, Jeronimo C; Resende, Daniela de M; Reis, Alexandre B

2018-01-01

Due to an increase in the incidence of leishmaniases worldwide, the development of new strategies such as prophylactic vaccines to prevent infection and decrease the disease have become a high priority. Classic vaccines against leishmaniases were based on live or attenuated parasites or their subunits. Nevertheless, the use of whole parasite or their subunits for vaccine production has numerous disadvantages. Therefore, the use of Leishmania peptides to design more specific vaccines against leishmaniases seems promising. Moreover, peptides have several benefits in comparison with other kinds of antigens, for instance, good stability, absence of potentially damaging materials, antigen low complexity, and low-cost to scale up. By contrast, peptides are poor immunogenic alone, and they need to be delivered correctly. In this context, several approaches described in this review are useful to solve these drawbacks. Approaches, such as, peptides in combination with potent adjuvants, cellular vaccinations, adenovirus, polyepitopes, or DNA vaccines have been used to develop peptide-based vaccines. Recent advancements in peptide vaccine design, chimeric, or polypeptide vaccines and nanovaccines based on particles attached or formulated with antigenic components or peptides have been increasingly employed to drive a specific immune response. In this review, we briefly summarize the old, current, and future stands on peptide-based vaccines, describing the disadvantages and benefits associated with them. We also propose possible approaches to overcome the related weaknesses of synthetic vaccines and suggest future guidelines for their development.

Vaccination with poly(D,L-lactide-co-glycolide) nanoparticles loaded with soluble Leishmania antigens and modified with a TNFα-mimicking peptide or monophosphoryl lipid A confers protection against experimental visceral leishmaniasis.

PubMed

Margaroni, Maritsa; Agallou, Maria; Athanasiou, Evita; Kammona, Olga; Kiparissides, Costas; Gaitanaki, Catherine; Karagouni, Evdokia

2017-01-01

Visceral leishmaniasis (VL) persists as a major public health problem, and since the existing chemotherapy is far from satisfactory, development of an effective vaccine emerges as the most appropriate strategy for confronting VL. The development of an effective vaccine relies on the selection of the appropriate antigen and also the right adjuvant and/or delivery vehicle. In the present study, the protective efficacy of poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles (NPs), which were surface-modified with a TNFα-mimicking eight-amino-acid peptide (p8) and further functionalized by encapsulating soluble Leishmania infantum antigens (sLiAg) and monophosphoryl lipid A (MPLA), a TLR4 ligand, was evaluated against challenge with L. infantum parasites in BALB/c mice. Vaccination with these multifunctionalized PLGA nanoformulations conferred significant protection against parasite infection in vaccinated mice. In particular, vaccination with PLGA-sLiAg-MPLA or p8-PLGA-sLiAg NPs resulted in almost complete elimination of the parasite in the spleen for up to 4 months post-challenge. Parasite burden reduction was accompanied by antigen-specific humoral and cellular immune responses. Specifically, injection with PLGA-sLiAg-MPLA raised exclusively anti-sLiAg IgG1 antibodies post-vaccination, while in p8-PLGA-sLiAg-vaccinated mice, no antibody production was detected. However, 4 months post-challenge, in mice vaccinated with all the multifunctionalized NPs, antibody class switching towards IgG2a subtype was observed. The study of cellular immune responses revealed the increased proliferation capacity of spleen cells against sLiAg, consisting of IFNγ-producing CD4 + and CD8 + T cells. Importantly, the activation of CD8 + T cells was exclusively attributed to vaccination with PLGA NPs surface-modified with the p8 peptide. Moreover, characterization of cytokine production in vaccinated-infected mice revealed that protection was accompanied by significant increase of IFN�

Impact and prevention of severe exacerbations of COPD: a review of the evidence

PubMed Central

Halpin, David MG; Miravitlles, Marc; Metzdorf, Norbert; Celli, Bartolomé

2017-01-01

Severe exacerbations of COPD, ie, those leading to hospitalization, have profound clinical implications for patients and significant economic consequences for society. The prevalence and burden of severe COPD exacerbations remain high, despite recognition of the importance of exacerbation prevention and the availability of new treatment options. Severe COPD exacerbations are associated with high mortality, have negative impact on quality of life, are linked to cardiovascular complications, and are a significant burden on the health-care system. This review identified risk factors that contribute to the development of severe exacerbations, treatment options (bronchodilators, antibiotics, corticosteroids [CSs], oxygen therapy, and ventilator support) to manage severe exacerbations, and strategies to prevent readmission to hospital. Risk factors that are amenable to change have been highlighted. A number of bronchodilators have demonstrated successful reduction in risk of severe exacerbations, including long-acting muscarinic antagonist or long-acting β2-agonist mono- or combination therapies, in addition to vaccination, mucolytic and antibiotic therapy, and nonpharmacological interventions, such as pulmonary rehabilitation. Recognition of the importance of severe exacerbations is an essential step in improving outcomes for patients with COPD. Evidence-based approaches to prevent and manage severe exacerbations should be implemented as part of targeted strategies for disease management. PMID:29062228

LaRbp38: A Leishmania amazonensis protein that binds nuclear and kinetoplast DNAs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lira, C.B.B.; Instituto de Biologia, UNICAMP, Campinas, SP; Siqueira Neto, J.L.

Leishmania amazonensis causes a wide spectrum of leishmaniasis. There are no vaccines or adequate treatment for leishmaniasis, therefore there is considerable interest in the identification of new targets for anti-leishmania drugs. The central role of telomere-binding proteins in cell maintenance makes these proteins potential targets for new drugs. In this work, we used a combination of purification chromatographies to screen L. amazonensis proteins for molecules capable of binding double-stranded telomeric DNA. This approach resulted in the purification of a 38 kDa polypeptide that was identified by mass spectrometry as Rbp38, a trypanosomatid protein previously shown to stabilize mitochondrial RNA andmore » to associate with nuclear and kinetoplast DNAs. Western blotting and supershift assays confirmed the identity of the protein as LaRbp38. Competition and chromatin immunoprecipitation assays confirmed that LaRbp38 interacted with kinetoplast and nuclear DNAs in vivo and suggested that LaRbp38 may have dual cellular localization and more than one function.« less

Immunogenicity Evaluation of a Rationally Designed Polytope Construct Encoding HLA-A*0201 Restricted Epitopes Derived from Leishmania major Related Proteins in HLA-A2/DR1 Transgenic Mice: Steps toward Polytope Vaccine

PubMed Central

Seyed, Negar; Taheri, Tahereh; Vauchy, Charline; Dosset, Magalie; Godet, Yann; Eslamifar, Ali; Sharifi, Iraj; Adotevi, Olivier; Borg, Christophe; Rohrlich, Pierre Simon; Rafati, Sima

2014-01-01

Background There are several reports demonstrating the role of CD8 T cells against Leishmania species. Therefore peptide vaccine might represent an effective approach to control the infection. We developed a rational polytope-DNA construct encoding immunogenic HLA-A2 restricted peptides and validated the processing and presentation of encoded epitopes in a preclinical mouse model humanized for the MHC-class-I and II. Methods and Findings HLA-A*0201 restricted epitopes from LPG-3, LmSTI-1, CPB and CPC along with H-2Kd restricted peptides, were lined-up together as a polytope string in a DNA construct. Polytope string was rationally designed by harnessing advantages of ubiquitin, spacers and HLA-DR restricted Th1 epitope. Endotoxin free pcDNA plasmid expressing the polytope was inoculated into humanized HLA-DRB1*0101/HLA-A*0201 transgenic mice intramuscularly 4 days after Cardiotoxin priming followed by 2 boosters at one week interval. Mice were sacrificed 10 days after the last booster, and splenocytes were subjected to ex-vivo and in-vitro evaluation of specific IFN-γ production and in-vitro cytotoxicity against individual peptides by ELISpot and standard chromium-51(51Cr) release assay respectively. 4 H-2Kd and 5 HLA-A*0201 restricted peptides were able to induce specific CD8 T cell responses in BALB/C and HLA-A2/DR1 mice respectively. IFN-γ and cytolytic activity together discriminated LPG-3-P1 as dominant, LmSTI-1-P3 and LmSTI-1-P6 as subdominant with both cytolytic activity and IFN-γ production, LmSTI-1-P4 and LPG-3-P5 as subdominant with only IFN-γ production potential. Conclusions Here we described a new DNA-polytope construct for Leishmania vaccination encompassing immunogenic HLA-A2 restricted peptides. Immunogenicity evaluation in HLA-transgenic model confirmed CD8 T cell induction with expected affinities and avidities showing almost efficient processing and presentation of the peptides in relevant preclinical model. Further evaluation will determine

Immuno-informatics based approaches to identify CD8+ T cell epitopes within the Leishmania donovani 3-ectonucleotidase in cured visceral leishmaniasis subjects.

PubMed

Vijayamahantesh; Amit, Ajay; Dikhit, Manas R; Singh, Ashish K; Venkateshwaran, T; Das, V N R; Das, Pradeep; Bimal, Sanjiva

2017-06-01

Leishmaniases are vector-borne diseases for which no vaccine exists. These diseases are caused by the Leishmania species complex. Activation of the CD8 + T cell is crucial for protection against intracellular pathogens, and peptide antigens are attractive strategies for the precise activation of CD8 + T in vaccine development against intracellular infections. The traditional approach to mine the epitopes is an arduous task. However, with the advent of immunoinformatics, in silico epitope prediction tools are available to expedite epitope identification. In this study, we employ different immunoinformatics tools to predict CD8 + T cell specific 9 mer epitopes presented by HLA-A*02 and HLA-B40 within the highly conserved 3'-ectonucleotidase of Leishmania donovani. We identify five promiscuous epitopes, which have no homologs in humans, theoretically cover 85% of the world's population and are highly conserved (100%) among Leishmania species. Presentation of selected peptides was confirmed by T2 cell line based HLA-stabilization assay, and three of them were found to be strong binders. The in vitro peptide stimulation of peripheral blood mononuclear cells (PBMC) from cured HLA-A02 + visceral leishmaniasis (VL) subjects produced significantly higher IFN-γ, IL-2 and IL-12 compared to no peptide control healthy subjects. Further, CD8 + cells from treated VL subjects produced significantly higher intracellular IFN-γ, lymphocyte proliferation and cytotoxic activity against selected peptides from the PBMCs of treated HLA-A02 + VL subjects. Thus, the CD8 + T cell specific epitopes shown in this study will speed up the development of polytope vaccines for leishmaniasis. Copyright © 2017. Published by Elsevier Masson SAS.

Leishmaniasis: vaccine candidates and perspectives.

PubMed

Singh, Bhawana; Sundar, Shyam

2012-06-06

Leishmania is a protozoan parasite and a causative agent of the various clinical forms of leishmaniasis. High cost, resistance and toxic side effects of traditional drugs entail identification and development of therapeutic alternatives. The sound understanding of parasite biology is key for identifying novel drug targets, that can induce the cell mediated immunity (mainly CD4+ and CD8+ IFN-gamma mediated responses) polarized towards a Th1 response. These aspects are important in designing a new vaccine along with the consideration of the candidates with respect to their ability to raise memory response in order to improve the vaccine performance. This review is an effort to identify molecules according to their homology with the host and their ability to be used as potent vaccine candidates. Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

Gluconeogenesis in Leishmania mexicana

PubMed Central

Rodriguez-Contreras, Dayana; Hamilton, Nicklas

2014-01-01

Gluconeogenesis is an active pathway in Leishmania amastigotes and is essential for their survival within the mammalian cells. However, our knowledge about this pathway in trypanosomatids is very limited. We investigated the role of glycerol kinase (GK), phosphoenolpyruvate carboxykinase (PEPCK), and pyruvate phosphate dikinase (PPDK) in gluconeogenesis by generating the respective Leishmania mexicana Δgk, Δpepck, and Δppdk null mutants. Our results demonstrated that indeed GK, PEPCK, and PPDK are key players in the gluconeogenesis pathway in Leishmania, although stage-specific differences in their contribution to this pathway were found. GK participates in the entry of glycerol in promastigotes and amastigotes; PEPCK participates in the entry of aspartate in promastigotes, and PPDK is involved in the entry of alanine in amastigotes. Furthermore, the majority of alanine enters into the pathway via decarboxylation of pyruvate in promastigotes, whereas pathway redundancy is suggested for the entry of aspartate in amastigotes. Interestingly, we also found that l-lactate, an abundant glucogenic precursor in mammals, was used by Leishmania amastigotes to synthesize mannogen, entering the pathway through PPDK. On the basis of these new results, we propose a revision in the current model of gluconeogenesis in Leishmania, emphasizing the differences between amastigotes and promastigotes. This work underlines the importance of studying the trypanosomatid intracellular life cycle stages to gain a better understanding of the pathologies caused in humans. PMID:25288791

Serological diagnosis of canine leishmaniosis: comparison of three commercial ELISA tests (Leiscan®, ID Screen® and Leishmania 96®), a rapid test (Speed Leish K®) and an in-house IFAT

PubMed Central

2014-01-01

Background Speed Leish K® is used as a serological screening test for Leishmania infection prior to vaccination. Limited comparative serological studies with Speed Leish K® have been performed. The aim of this study was to evaluate the diagnostic performance of four commercially available serologic tests including ELISAs (Leiscan®, ID Screen® and Leishmania 96®), a rapid test (Speed Leish K®) and an in-house IFAT for the detection of specific antibodies against Leishmania infantum antigen in dogs in different states of infection. Methods Sick infected dogs (n = 36), healthy infected dogs (n = 18), L. infantum seropositive dogs with low to high levels of antibodies (n = 53), dogs seropositive to other pathogens (to evaluate cross reaction) (n = 14) and uninfected dogs from a non-endemic area (n = 50) and from an endemic area (n = 32) were analysed by the serological methods mentioned above. Results The sensitivity was as follows: ID Screen® (0.953), Leiscan® and Leishmania 96® (0.925), IFAT (0.869) and Speed Leish K® (0.636). The maximum specificity (1.000) was attained for all diagnostic tests except the Leishmania 96® (0.896) and IFAT (0.917). The accuracy was as follows: ID Screen® (0.975), Leiscan® (0.961), Leishmania 96® (0.911), IFAT (0.892) and Speed Leish K® (0.808). In relation to the area under the ROC curve (AUC-ROC), the maximum value was attained with the ID Screen® (0.993) closely followed by Leiscan® (0.990), then, Leishmania 96® (0.962), IFAT (0.926) and Speed Leish K® (0.818). For the Kappa index, the best result was obtained by the ID Screen® (0.951) followed by Leiscan® (0.921), Leishmania 96® (0.822), IFAT (0.783) and Speed Leish K® (0.622). Statistically significant differences were found between the AUC-ROC of quantitative serological tests and the only qualitative rapid test evaluated. There were also statistically significant differences between AUC-ROC of the ELISAs (ID Screen® and Leiscan

Extreme inbreeding in Leishmania braziliensis

PubMed Central

Rougeron, Virginie; De Meeûs, Thierry; Hide, Mallorie; Waleckx, Etienne; Bermudez, Herman; Arevalo, Jorge; Llanos-Cuentas, Alejandro; Dujardin, Jean-Claude; De Doncker, Simone; Le Ray, Dominique; Ayala, Francisco J.; Bañuls, Anne-Laure

2009-01-01

Leishmania species of the subgenus Viannia and especially Leishmania braziliensis are responsible for a large proportion of New World leishmaniasis cases. The reproductive mode of Leishmania species has often been assumed to be predominantly clonal, but remains unsettled. We have investigated the genetic polymorphism at 12 microsatellite loci on 124 human strains of Leishmania braziliensis from 2 countries, Peru and Bolivia. There is substantial genetic diversity, with an average of 12.4 ± 4.4 alleles per locus. There is linkage disequilibrium at a genome-wide scale, as well as a substantial heterozygote deficit (more than 50% the expected value from Hardy−Weinberg equilibrium), which indicates high levels of inbreeding. These observations are inconsistent with a strictly clonal model of reproduction, which implies excess heterozygosity. Moreover, there is large genetic heterogeneity between populations within countries (Wahlund effect), which evinces a strong population structure at a microgeographic scale. Our findings are compatible with the existence of population foci at a microgeographic scale, where clonality alternates with sexuality of an endogamic nature, with possible occasional recombination events between individuals of different genotypes. These findings provide key clues on the ecology and transmission patterns of Leishmania parasites. PMID:19497885

Seroprevalence of Leishmania infection and molecular detection of Leishmania tropica and Leishmania infantum in stray cats of İzmir, Turkey.

PubMed

Can, Hüseyin; Döşkaya, Mert; Özdemir, H Gökhan; Şahar, Esra Atalay; Karakavuk, Muhammet; Pektaş, Bayram; Karakuş, Mehmet; Töz, Seray; Caner, Ayşe; Döşkaya, Aysu Değirmenci; İz, Sultan Gülce; Özbel, Yusuf; Gürüz, Yüksel

2016-08-01

Leishmaniasis caused by more than 20 species of genus Leishmania is transmitted by the bite of infected phlebotomine sand flies. The studies on Leishmania infection in cats is very few in Turkey and therefore we aimed to screen stray cats living in city of İzmir located in western Turkey using nested PCR targeting kinetoplast DNA and serological techniques (ELISA and IFA). Leishmania DNA positive samples were also studied by ITS1 real time PCR. Whole blood and serum samples were obtained from stray cats (n: 1101) living in different counties of İzmir. In serological assays, a serum sample was considered positive in 1:40 dilution in IFA and for ELISA a serum sample was accepted positive when the absorbance value (AV) exceeded the mean AV + Standard Deviation (SD) of the negative control serum samples. According to the results, the seropositivity rates were 10.8% (119/1101) and 15.2% (167/1101) by in house ELISA and IFA, respectively. Among serology coherent samples, the seropositivity rate was 11.1% (116/1047) as detected by both assays after discordant samples (n: 54) were discarded. Of the 1101 stray cats, six (0.54%) were positive by nested PCR while only one of these six samples was positive by ITS1 real time PCR. During PCR, three controls designated as Leishmania infantum, Leishmania tropica, and Leishmania major were used for species identification. According to nested PCR results, L. tropica was identified in two cats (no.76 and 95). In another cat (no. 269), there were two bands in which one of them was well-matched with L. infantum and the other band had ∼850 bp size which does not match with any controls. Remaining three cats (no. 86, 514, and 622) also had the ∼850 bp atypical band size. ITS1 real time PCR detected L. tropica in only one cat (no. 622) which showed an atypical band size in nested PCR. These results indicated that three cats with only one atypical band (no. 86, 514, and 622) and the cat with mixed infection (no. 269) were

Assessment of nuclear and mitochondrial genes in precise identification and analysis of genetic polymorphisms for the evaluation of Leishmania parasites.

PubMed

Fotouhi-Ardakani, Reza; Dabiri, Shahriar; Ajdari, Soheila; Alimohammadian, Mohammad Hossein; AlaeeNovin, Elnaz; Taleshi, Neda; Parvizi, Parviz

2016-12-01

.85 & 2.91, P<0.01). More different clinical lesions with extensive phylogenetic and evolutionary analyses should be employed to avoid confusion in the diagnosis of leishmaniasis and development of vaccines for eradicating Leishmania parasites. Copyright © 2016 Elsevier B.V. All rights reserved.

Phylogenetic analysis of HSP70 and cyt b gene sequences for Chinese Leishmania isolates and ultrastructural characteristics of Chinese Leishmania sp.

PubMed

Yuan, Dongmei; Qin, Hanxiao; Zhang, Jianguo; Liao, Lin; Chen, Qiwei; Chen, Dali; Chen, Jianping

2017-02-01

Leishmaniasis is a worldwide epidemic disease caused by the genus Leishmania, which is still endemic in the west and northwest areas of China. Some viewpoints of the traditional taxonomy of Chinese Leishmania have been challenged by recent phylogenetic researches based on different molecular markers. However, the taxonomic positions and phylogenetic relationships of Chinese Leishmania isolates remain controversial, which need for more data and further analysis. In this study, the heat shock protein 70 (HSP70) gene and cytochrome b (cyt b) gene were used for phylogenetic analysis of Chinese Leishmania isolates from patients, dogs, gerbils, and sand flies in different geographic origins. Besides, for the interesting Leishmania sp. in China, the ultrastructure of three Chinese Leishmania sp. strains (MHOM/CN/90/SC10H2, SD, GL) were observed by transmission electron microscopy. Bayesian trees from HSP70 and cyt b congruently indicated that the 14 Chinese Leishmania isolates belong to three Leishmania species including L. donovani complex, L. gerbilli, and L. (Sauroleishmania) sp. Their identity further confirmed that the undescribed Leishmania species causing visceral Leishmaniasis (VL) in China is closely related to L. tarentolae. The phylogenetic results from HSP70 also suggested the classification of subspecies within L. donovani complex: KXG-918, KXG-927, KXG-Liu, KXG-Xu, 9044, SC6, and KXG-65 belong to L. donovani; Cy, WenChuan, and 801 were proposed to be L. infantum. Through transmission electron microscopy, unexpectedly, the Golgi apparatus were not observed in SC10H2, SD, and GL, which was similar to previous reports of reptilian Leishmania. The statistical analysis of microtubule counts separated SC10H2, SD, and GL as one group from any other reference strain (L. donovani MHOM/IN/80/DD8; L. tropica MHOM/SU/74/K27; L. gerbilli MRHO/CN/60/GERBILLI). The ultrastructural characteristics of Leishmania sp. partly lend support to the phylogenetic inference that

[Approaches and problems in vaccine development against leishmaniasis].

PubMed

Allahverdiyev, Adil; Bağirova, Melahat; Cakir Koç, Rabia; Oztel, Olga Nehir; Elçıçek, Serhat; Ateş, Sezen Canım; Karaca, Tuğçe Deniz

2010-01-01

Leishmaniasis is a major public health problem of the world and Turkey. Recently there has been increasing interest in vaccine studies among strategies for control of leishmaniasis. Recently the increase of interest in vaccine studies among leishmaniasis control strategies makes the subject more up to date. So the aim of this review is to present information about recent vaccine studies, problems and new strategies for vaccine development studies. There are 3 generations of vaccine against leishmaniasis. First-generation vaccines are killed or live attenuated parasites; second-generation vaccines are recombinant or native antigens and live genetically modified parasites (knock out and suicidal cassettes), third generation vaccines are DNA vaccines. Also vector salivary proteins, dendritic cells and non-pathogenic L. tarentolae have been used as vaccine candidates. However there is still no effective vaccine against leishmaniasis. Since polymer conjugates considerably increase immunogenicity, polymer based vaccine studies have gained importance in recent years. However, there has not been such a study for an antileishmanial vaccine yet. LPG, surface antigen of Leishmania promastigotes, and polymer conjugates may be promising in antileishmanial vaccine studies so we are carrying out a TUBITAK Project on this subject which has been given the number, 1085170SBAG-4007.

Vaccination with plasmid DNA encoding TSA/LmSTI1 leishmanial fusion proteins confers protection against Leishmania major infection in susceptible BALB/c mice.

PubMed

Campos-Neto, A; Webb, J R; Greeson, K; Coler, R N; Skeiky, Y A W; Reed, S G

2002-06-01

We have recently shown that a cocktail containing two leishmanial recombinant antigens (LmSTI1 and TSA) and interleukin-12 (IL-12) as an adjuvant induces solid protection in both a murine and a nonhuman primate model of cutaneous leishmaniasis. However, because IL-12 is difficult to prepare, is expensive, and does not have the stability required for a vaccine product, we have investigated the possibility of using DNA as an alternative means of inducing protective immunity. Here, we present evidence that the antigens TSA and LmSTI1 delivered in a plasmid DNA format either as single genes or in a tandem digene construct induce equally solid protection against Leishmania major infection in susceptible BALB/c mice. Immunization of mice with either TSA DNA or LmSTI1 DNA induced specific CD4(+)-T-cell responses of the Th1 phenotype without a requirement for specific adjuvant. CD8 responses, as measured by cytotoxic-T-lymphocyte activity, were generated after immunization with TSA DNA but not LmSTI1 DNA. Interestingly, vaccination of mice with TSA DNA consistently induced protection to a much greater extent than LmSTI1 DNA, thus supporting the notion that CD8 responses might be an important accessory arm of the immune response for acquired resistance against leishmaniasis. Moreover, the protection induced by DNA immunization was specific for infection with Leishmania, i.e., the immunization had no effect on the course of infection of the mice challenged with an unrelated intracellular pathogen such as Mycobacterium tuberculosis. Conversely, immunization of BALB/c mice with a plasmid DNA that is protective against challenge with M. tuberculosis had no effect on the course of infection of these mice with L. major. Together, these results indicate that the protection observed with the leishmanial DNA is mediated by acquired specific immune response rather than by the activation of nonspecific innate immune mechanisms. In addition, a plasmid DNA containing a fusion construct

Natural infection of bats with Leishmania in Ethiopia.

PubMed

Kassahun, Aysheshm; Sadlova, Jovana; Benda, Petr; Kostalova, Tatiana; Warburg, Alon; Hailu, Asrat; Baneth, Gad; Volf, Petr; Votypka, Jan

2015-10-01

The leishmaniases, a group of diseases with a worldwide-distribution, are caused by different species of Leishmania parasites. Both cutaneous and visceral leishmaniasis remain important public health problems in Ethiopia. Epidemiological cycles of these protozoans involve various sand fly (Diptera: Psychodidae) vectors and mammalian hosts, including humans. In recent years, Leishmania infections in bats have been reported in the New World countries endemic to leishmaniasis. The aim of this study was to survey natural Leishmania infection in bats collected from various regions of Ethiopia. Total DNA was isolated from spleens of 163 bats belonging to 23 species and 18 genera. Leishmania infection was detected by real-time (RT) PCR targeting a kinetoplast (k) DNA and internal transcribed spacer one (ITS1) gene of the parasite. Detection was confirmed by sequencing of the PCR products. Leishmania kDNA was detected in eight (4.9%) bats; four of them had been captured in the Aba-Roba and Awash-Methara regions that are endemic for leishmaniasis, while the other four specimens originated from non-endemic localities of Metu, Bedele and Masha. Leishmania isolates from two bats were confirmed by ITS1 PCR to be Leishmania tropica and Leishmania major, isolated from two individual bats, Cardioderma cor and Nycteris hispida, respectively. These results represent the first confirmed observation of natural infection of bats with the Old World Leishmania. Hence, bats should be considered putative hosts of Leishmania spp. affecting humans with a significant role in the transmission. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

In vitro evaluation of photodynamic therapy using curcumin on Leishmania major and Leishmania braziliensis.

PubMed

Pinto, Juliana Guerra; Fontana, Letícia Correa; de Oliveira, Marco Antonio; Kurachi, Cristina; Raniero, Leandro José; Ferreira-Strixino, Juliana

2016-07-01

Cutaneous leishmaniasis is an infectious disease caused by the Leishmania protozoan. The conventional treatment is long-lasting and aggressive, in addition to causing harmful effect. Photodynamic therapy has emerged as a promising alternative treatment, which allows local administration with fewer side effects. This study investigated the photodynamic activity of curcumin on Leishmania major and Leishmania braziliensis promastigote. Both species were submitted to incubation with curcumin in serial dilutions from 500 μg/ml up to 7.8 μg/ml. Control groups were kept in the dark while PDT groups received a fluency of 10 J/cm(2) at 450 nm. Mitochondrial activity was assessed by MTT assay 18 h after light treatment, and viability was measured by Trypan blue dye exclusion test. Morphological alterations were observed by Giemsa staining. Confocal microscopy showed the uptake of curcumin by both tested Leishmania species. Mitochondrial activity was inconclusive to determine viability; however, Trypan blue test was able to show that curcumin photodynamic treatment had a significant effect on viability of parasites. The morphology of promastigotes was highly affected by the photodynamic therapy. These results indicated that curcumin may be a promising alternative photosensitizer, because it presents no toxicity in the dark; however, further tests in co-culture with macrophages and other species of Leishmania should be conducted to determine better conditions before in vivo tests are performed.

Leishmania hijacking of the macrophage intracellular compartments.

PubMed

Liévin-Le Moal, Vanessa; Loiseau, Philippe M

2016-02-01

Leishmania spp., transmitted to humans by the bite of the sandfly vector, are responsible for the three major forms of leishmaniasis, cutaneous, diffuse mucocutaneous and visceral. Leishmania spp. interact with membrane receptors of neutrophils and macrophages. In macrophages, the parasite is internalized within a parasitophorous vacuole and engages in a particular intracellular lifestyle in which the flagellated, motile Leishmania promastigote metacyclic form differentiates into non-motile, metacyclic amastigote form. This phenomenon is induced by Leishmania-triggered events leading to the fusion of the parasitophorous vacuole with vesicular members of the host cell endocytic pathway including recycling endosomes, late endosomes and the endoplasmic reticulum. Maturation of the parasitophorous vacuole leads to the intracellular proliferation of the Leishmania amastigote forms by acquisition of host cell nutrients while escaping host defense responses. © 2015 FEBS.

Evaluation of the adjuvant effect of agonists of toll-like receptor 4 and 7/8 in a vaccine against leishmaniasis in BALB/c mice.

PubMed

Rostamian, Mosayeb; Niknam, Hamid M

2017-11-01

There is no effective vaccine against human leishmaniasis. Achieving successful vaccines seems to need powerful adjuvants. Separate or combined use of toll like receptor (TLR) agonists as adjuvant is a promising approach in Leishmania vaccine research. In present study, we evaluated adjuvant effect of separate or combined use of a TLR7/8 agonist, R848 and a TLR4 agonist, monophosphoryl lipid A (MPL) beside soluble Leishmania antigen (SLA) in BALB/c mice. Mice were vaccinated three times by SLA with separate or combined TLR7/8 and TLR4 agonists and were then challenged by Leishmania major. Delay type hypersensitivity, lesion development, parasite load, and cytokines (interferon gamma, and interleukin-10) response were assessed. Results showed: 1) MPL can slightly assist SLA in parasite load reduction, but it is not able to increase SLA ability in evoking DTH and cytokine responses or decreasing lesion diameter. 2) R848 does not affect the DTH response and parasite load of mice vaccinated with SLA, but it decreases/inhibits cytokine responses induced by SLA, leading to increase lesion diameter. 3) MPL neutralized inhibitory effect of R848. In overall, these data emphasize that MPL slightly assists SLA to make a more potent vaccine, but R848 is not a good adjuvant to induce T cell-dependent immune response in BALB/c mice, and therefore combination of these TLR agonists in the current formulation, is not recommended for making a more powerful adjuvant. Copyright © 2017 Elsevier Ltd. All rights reserved.

Experimental mixed infection of Leishmania (Leishmania) amazonensis and Leishmania (L.) infantum in hamsters (Mesocricetus auratus).

PubMed

DE Lima Celeste, Jordanna Luíza; Venuto Moura, Ana Paula; França-Silva, João Carlos; Matos DE Sousa, Gabriela; Oliveira Silva, Soraia; Norma Melo, Maria; Luiz Tafuri, Wagner; Carvalho Souza, Carolina; Monteiro DE Andrade, Hélida

2017-08-01

In South America, visceral leishmaniasis is frequently caused by Leishmania infantum and, at an unknown frequency, by Leishmania amazonensis. Therefore, mixed infections with these organisms are possible. Mixed infections might affect the clinical course, immune response, diagnosis, treatment and epidemiology of the disease. Here we describe the clinical course of mixed infections with L. amazonensis and L. infantum in a hamster model. We show that mixed infections are associated with more severe clinical disease than infection with L. amazonensis or L. infantum alone. In spleens with mixed infections, L. infantum outcompeted L. amazonensis in the tissue, but not in culture from tissue. We found increased levels of IgG in animals infected with L. infantum. Although more than 30 bands were revealed in a Western blot, the highest immunogenicity was observed with proteins having molecular masses of 95 and 90 kDa, whereas proteins with molecular masses of lower than 50 kDa were reactive frequently with serum from hamsters infected with L. amazonensis, and proteins with molecular masses of 80 and 70 kDa were reactive only with serum from hamsters infected with L. infantum. This finding has important implications regarding the biology of Leishmania and humoral immune responses to infections with these organisms.

Liposomal adjuvant development for leishmaniasis vaccines.

PubMed

Askarizadeh, Anis; Jaafari, Mahmoud Reza; Khamesipour, Ali; Badiee, Ali

2017-08-01

Leishmaniasis is a parasitic disease that ranges in severity from skin lesions to fatality. Since long-lasting protection is induced upon recovery from cutaneous leishmaniasis, development of an effective vaccine is promising. However, there is no vaccine for use in humans yet. It seems limited efficacy in leishmaniasis vaccines is due to lack of an appropriate adjuvant or delivery system. Hence, the use of particulate adjuvants such as liposomes for effective delivery to the antigen presenting cells (APCs) is a valuable strategy to enhance leishmaniasis vaccine efficacy. The extraordinary versatility of liposomes because of their unique amphiphilic and biphasic nature allows for using antigens or immunostimulators within the core, on the surface or within the bilayer, and modulates both the magnitude and the T-helper bias of the immune response. In this review article, we attempt to summarize the role of liposomal adjuvants in the development of Leishmania vaccines and describe the main physicochemical properties of liposomes like phospholipid composition, surface charge, and particle size during formulation design. We also suggest potentially useful formulation strategies in order for future experiments to have a chance to succeed as liposomal vaccines against leishmaniasis.

Liposomal adjuvant development for leishmaniasis vaccines

PubMed Central

Askarizadeh, Anis; Jaafari, Mahmoud Reza; Khamesipour, Ali; Badiee, Ali

2017-01-01

Leishmaniasis is a parasitic disease that ranges in severity from skin lesions to fatality. Since long-lasting protection is induced upon recovery from cutaneous leishmaniasis, development of an effective vaccine is promising. However, there is no vaccine for use in humans yet. It seems limited efficacy in leishmaniasis vaccines is due to lack of an appropriate adjuvant or delivery system. Hence, the use of particulate adjuvants such as liposomes for effective delivery to the antigen presenting cells (APCs) is a valuable strategy to enhance leishmaniasis vaccine efficacy. The extraordinary versatility of liposomes because of their unique amphiphilic and biphasic nature allows for using antigens or immunostimulators within the core, on the surface or within the bilayer, and modulates both the magnitude and the T-helper bias of the immune response. In this review article, we attempt to summarize the role of liposomal adjuvants in the development of Leishmania vaccines and describe the main physicochemical properties of liposomes like phospholipid composition, surface charge, and particle size during formulation design. We also suggest potentially useful formulation strategies in order for future experiments to have a chance to succeed as liposomal vaccines against leishmaniasis. PMID:29201374

No recent adaptive selection on the apyrase of Mediterranean Phlebotomus: implications for using salivary peptides to vaccinate against canine leishmaniasis.

PubMed

Mahamdallie, Shazia S; Ready, Paul D

2012-04-01

Vaccine development is informed by a knowledge of genetic variation among antigen alleles, especially the distribution of positive and balancing selection in populations and species. A combined approach using population genetic and phylogenetic methods to detect selective signatures can therefore be informative for identifying vaccine candidates. Parasitic Leishmania species cause the disease leishmaniasis in humans and mammalian reservoir hosts after inoculation by female phlebotomine sandflies. Like other arthropod vectors of disease agents, sandflies use salivary peptides to counteract host haemostatic and immunomodulatory responses during bloodfeeding, and these peptides are vaccine candidates because they can protect against Leishmania infection. We detected no contemporary adaptive selection on one salivary peptide, apyrase, in 20 populations of Phlebotomus ariasi, a European vector of Leishmania infantum. Maximum likelihood branch models on a gene phylogeny showed apyrase to be a single copy in P. ariasi but an ancient duplication event associated with temporary positive selection was observed in its sister group, which contains most Mediterranean vectors of L. infantum. The absence of contemporary adaptive selection on the apyrase of P. ariasi may result from this sandfly's opportunistic feeding behaviour. Our study illustrates how the molecular population genetics of arthropods can help investigate the potential of salivary peptides for disease control and for understanding geographical variation in vector competence.

The Comparative Genomics and Phylogenomics of Leishmania amazonensis Parasite.

PubMed

Tschoeke, Diogo A; Nunes, Gisele L; Jardim, Rodrigo; Lima, Joana; Dumaresq, Aline Sr; Gomes, Monete R; de Mattos Pereira, Leandro; Loureiro, Daniel R; Stoco, Patricia H; de Matos Guedes, Herbert Leonel; de Miranda, Antonio Basilio; Ruiz, Jeronimo; Pitaluga, André; Silva, Floriano P; Probst, Christian M; Dickens, Nicholas J; Mottram, Jeremy C; Grisard, Edmundo C; Dávila, Alberto Mr

2014-01-01

Leishmaniasis is an infectious disease caused by Leishmania species. Leishmania amazonensis is a New World Leishmania species belonging to the Mexicana complex, which is able to cause all types of leishmaniasis infections. The L. amazonensis reference strain MHOM/BR/1973/M2269 was sequenced identifying 8,802 codifying sequences (CDS), most of them of hypothetical function. Comparative analysis using six Leishmania species showed a core set of 7,016 orthologs. L. amazonensis and Leishmania mexicana share the largest number of distinct orthologs, while Leishmania braziliensis presented the largest number of inparalogs. Additionally, phylogenomic analysis confirmed the taxonomic position for L. amazonensis within the "Mexicana complex", reinforcing understanding of the split of New and Old World Leishmania. Potential non-homologous isofunctional enzymes (NISE) were identified between L. amazonensis and Homo sapiens that could provide new drug targets for development.

The Comparative Genomics and Phylogenomics of Leishmania amazonensis Parasite

PubMed Central

Tschoeke, Diogo A; Nunes, Gisele L; Jardim, Rodrigo; Lima, Joana; Dumaresq, Aline SR; Gomes, Monete R; de Mattos Pereira, Leandro; Loureiro, Daniel R; Stoco, Patricia H; de Matos Guedes, Herbert Leonel; de Miranda, Antonio Basilio; Ruiz, Jeronimo; Pitaluga, André; Silva, Floriano P; Probst, Christian M; Dickens, Nicholas J; Mottram, Jeremy C; Grisard, Edmundo C; Dávila, Alberto MR

2014-01-01

Leishmaniasis is an infectious disease caused by Leishmania species. Leishmania amazonensis is a New World Leishmania species belonging to the Mexicana complex, which is able to cause all types of leishmaniasis infections. The L. amazonensis reference strain MHOM/BR/1973/M2269 was sequenced identifying 8,802 codifying sequences (CDS), most of them of hypothetical function. Comparative analysis using six Leishmania species showed a core set of 7,016 orthologs. L. amazonensis and Leishmania mexicana share the largest number of distinct orthologs, while Leishmania braziliensis presented the largest number of inparalogs. Additionally, phylogenomic analysis confirmed the taxonomic position for L. amazonensis within the “Mexicana complex”, reinforcing understanding of the split of New and Old World Leishmania. Potential non-homologous isofunctional enzymes (NISE) were identified between L. amazonensis and Homo sapiens that could provide new drug targets for development. PMID:25336895

The emergence of Leishmania major and Leishmania donovani in southern Turkey.

PubMed

Koltas, Ismail S; Eroglu, Fadime; Alabaz, Derya; Uzun, Soner

2014-03-01

In southern Turkey, Leishmania tropica and L. infantum are both the causative agents of cutaneous leishmaniasis (CL) and visceral leishmaniasis (VL), respectively. However, L. major and L. donovani were known to exist after the influx of Syrian refugees. Between the years of July 2003 and July 2013, a total of 167 smears and 113 bone marrow samples were taken from CL and VL-suspected cases, respectively. Samples were analysed through real-time PCR and ITS1 DNA sequencing. One hundred and seven 64% (107/167) smears and 56% (63/113) bone marrow samples were positive for leishmaniasis according to the real-time PCR. Three different Leishmania species were found in the 107 CL cases by real-time PCR: 42% (45/107) L. tropica, 36.5% (39/107) L. infantum and 21.5% (23/107) L. major. In addition, three different Leishmania species were identified in the 63 VL cases: 60.3% (38/63) L. infantum, 30.2% (19/63) L. donovani and 9.5% (6/63) L. tropica using real-time PCR. The results of real-time PCR were confirmed with Leishmania ITS1 DNA sequencing. This study revealed that in southern Turkey, L. major and L. donovani were the aetiological agents of CL and VL, respectively. It was assumed that emergence of L. major and L. donovani was due to influx of Syrian refugees, as well as the effects of global warming.

Vaccination with poly(D,L-lactide-co-glycolide) nanoparticles loaded with soluble Leishmania antigens and modified with a TNFα-mimicking peptide or monophosphoryl lipid A confers protection against experimental visceral leishmaniasis

PubMed Central

Margaroni, Maritsa; Agallou, Maria; Athanasiou, Evita; Kammona, Olga; Kiparissides, Costas; Gaitanaki, Catherine; Karagouni, Evdokia

2017-01-01

Visceral leishmaniasis (VL) persists as a major public health problem, and since the existing chemotherapy is far from satisfactory, development of an effective vaccine emerges as the most appropriate strategy for confronting VL. The development of an effective vaccine relies on the selection of the appropriate antigen and also the right adjuvant and/or delivery vehicle. In the present study, the protective efficacy of poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles (NPs), which were surface-modified with a TNFα-mimicking eight-amino-acid peptide (p8) and further functionalized by encapsulating soluble Leishmania infantum antigens (sLiAg) and monophosphoryl lipid A (MPLA), a TLR4 ligand, was evaluated against challenge with L. infantum parasites in BALB/c mice. Vaccination with these multifunctionalized PLGA nanoformulations conferred significant protection against parasite infection in vaccinated mice. In particular, vaccination with PLGA-sLiAg-MPLA or p8-PLGA-sLiAg NPs resulted in almost complete elimination of the parasite in the spleen for up to 4 months post-challenge. Parasite burden reduction was accompanied by antigen-specific humoral and cellular immune responses. Specifically, injection with PLGA-sLiAg-MPLA raised exclusively anti-sLiAg IgG1 antibodies post-vaccination, while in p8-PLGA-sLiAg-vaccinated mice, no antibody production was detected. However, 4 months post-challenge, in mice vaccinated with all the multifunctionalized NPs, antibody class switching towards IgG2a subtype was observed. The study of cellular immune responses revealed the increased proliferation capacity of spleen cells against sLiAg, consisting of IFNγ-producing CD4+ and CD8+ T cells. Importantly, the activation of CD8+ T cells was exclusively attributed to vaccination with PLGA NPs surface-modified with the p8 peptide. Moreover, characterization of cytokine production in vaccinated–infected mice revealed that protection was accompanied by significant increase of IFNγ and

The immunology of Leishmania/HIV co-infection.

PubMed

Okwor, Ifeoma; Uzonna, Jude Eze

2013-05-01

Leishmaniases are emerging as an important disease in human immunodeficiency virus (HIV)-infected persons living in several sub-tropical and tropical regions around the world, including the Mediterranean. The HIV/AIDS pandemic is spreading at an alarming rate in Africa and the Indian subcontinent, areas with very high prevalence of leishmaniases. The spread of HIV into rural areas and the concomitant spread of leishmaniases to suburban/urban areas have helped maintain the occurrence of Leishmania/HIV co-infection in many parts of the world. The number of cases of Leishmania/HIV co-infection is expected to rise owing to the overlapping geographical distribution of the two infections. In Southwestern Europe, there is also an increasing incidence of Leishmania/HIV co-infection (particularly visceral leishmaniasis) in such countries as France, Italy, Spain and Portugal. Studies suggest that in humans, very complex mechanisms involving dysregulation of host immune responses contribute to Leishmania-mediated immune activation and pathogenesis of HIV. In addition, both HIV-1 and Leishmania infect and multiply within cells of myeloid or lymphoid origin, thereby presenting a perfect recipe for reciprocal modulation of Leishmania and HIV-1-related disease pathogenesis. Importantly, because recovery from leishmaniases is associated with long-term persistence of parasites at the primary infection sites and their draining lymph nodes, there is very real possibility that HIV-mediated immunosuppression (due to CD4(+) T cell depletion) could lead to reactivation of latent infections (reactivation leishmaniasis) in immunocompromised patients. Here, we present an overview of the immunopathogenesis of Leishmania/HIV co-infection and the implications of this interaction on Leishmania and HIV disease outcome.

Leishmaniasis: focus on the design of nanoparticulate vaccine delivery systems.

PubMed

Doroud, Delaram; Rafati, Sima

2012-01-01

Although mass vaccination of the entire population of an endemic area would be the most cost-effective tool to diminish Leishmania burden, an effective vaccine is not yet commercially available. Practically, vaccines have failed to achieve the required level of protection, possibly owing to the lack of an appropriate adjuvant and/or delivery system. Therefore, there is still an imperative demand for an improved, safe and efficient delivery system to enhance the immunogenicity of available vaccine candidates. Nanoparticles are proficient in boosting the quality and magnitude of immune responses in a predictable fashion. Herein, we discuss how nanoparticulate vaccine delivery systems can be used to induce appropriate immune responses against leishmaniasis by controlling physicochemical properties of the vaccine. Stability, production reproducibility, low cost per dose and low risk-benefit ratios are desirable characteristics of an ideal vaccine formulation and solid lipid nanoparticles may serve as one of the most promising practical strategies to help to achieve such a leishmanial vaccine, at least in canine species in the developing world.

Analysis of the Antigenic and Prophylactic Properties of the Leishmania Translation Initiation Factors eIF2 and eIF2B in Natural and Experimental Leishmaniasis

PubMed Central

Garde, Esther; Ramírez, Laura; Corvo, Laura; Solana, José C.; Martín, M. Elena; González, Víctor M.; Gómez-Nieto, Carlos; Barral, Aldina; Barral-Netto, Manoel; Requena, José M.; Iborra, Salvador; Soto, Manuel

2018-01-01

Different members of intracellular protein families are recognized by the immune system of the vertebrate host infected by parasites of the genus Leishmania. Here, we have analyzed the antigenic and immunogenic properties of the Leishmania eIF2 and eIF2B translation initiation factors. An in silico search in Leishmania infantum sequence databases allowed the identification of the genes encoding the α, β, and γ subunits and the α, β, and δ subunits of the putative Leishmania orthologs of the eukaryotic initiation factors F2 (LieIF2) or F2B (LieIF2B), respectively. The antigenicity of these factors was analyzed by ELISA using recombinant versions of the different subunits. Antibodies against the different LieIF2 and LieIF2B subunits were found in the sera from human and canine visceral leishmaniasis patients, and also in the sera from hamsters experimentally infected with L. infantum. In L. infantum (BALB/c) and Leishmania major (BALB/c or C57BL/6) challenged mice, a moderate humoral response against these protein factors was detected. Remarkably, these proteins elicited an IL-10 production by splenocytes derived from infected mice independently of the Leishmania species employed for experimental challenge. When DNA vaccines based on the expression of the LieIF2 or LieIF2B subunit encoding genes were administered in mice, an antigen-specific secretion of IFN-γ and IL-10 cytokines was observed. Furthermore, a partial protection against murine CL development due to L. major infection was generated in the vaccinated mice. Also, in this work we show that the LieIF2α subunit and the LieIF2Bβ and δ subunits have the capacity to stimulate IL-10 secretion by spleen cells from naïve mice. B-lymphocytes were identified as the major producers of this anti-inflammatory cytokine. Taking into account the data found in this study, it may be hypothesized that these proteins act as virulence factors implicated in the induction of humoral responses as well as in the

Arginase expression modulates nitric oxide production in Leishmania (Leishmania) amazonensis.

PubMed

Acuña, Stephanie Maia; Aoki, Juliana Ide; Laranjeira-Silva, Maria Fernanda; Zampieri, Ricardo Andrade; Fernandes, Juliane Cristina Ribeiro; Muxel, Sandra Marcia; Floeter-Winter, Lucile Maria

2017-01-01

Arginase is an enzyme that converts L-arginine to urea and L-ornithine, an essential substrate for the polyamine pathway supporting Leishmania (Leishmania) amazonensis replication and its survival in the mammalian host. L-arginine is also the substrate of macrophage nitric oxide synthase 2 (NOS2) to produce nitric oxide (NO) that kills the parasite. This competition can define the fate of Leishmania infection. The transcriptomic profiling identified a family of oxidoreductases in L. (L.) amazonensis wild-type (La-WT) and L. (L.) amazonensis arginase knockout (La-arg-) promastigotes and axenic amastigotes. We highlighted the identification of an oxidoreductase that could act as nitric oxide synthase-like (NOS-like), due to the following evidences: conserved domain composition, the participation of NO production during the time course of promastigotes growth and during the axenic amastigotes differentiation, regulation dependence on arginase activity, as well as reduction of NO amount through the NOS activity inhibition. NO quantification was measured by DAF-FM labeling analysis in a flow cytometry. We described an arginase-dependent NOS-like activity in L. (L.) amazonensis and its role in the parasite growth. The increased detection of NO production in the mid-stationary and late-stationary growth phases of La-WT promastigotes could suggest that this production is an important factor to metacyclogenesis triggering. On the other hand, La-arg- showed an earlier increase in NO production compared to La-WT, suggesting that NO production can be arginase-dependent. Interestingly, La-WT and La-arg- axenic amastigotes produced higher levels of NO than those observed in promastigotes. As a conclusion, our work suggested that NOS-like is expressed in Leishmania in the stationary growth phase promastigotes and amastigotes, and could be correlated to metacyclogenesis and amastigotes growth in a dependent way to the internal pool of L-arginine and arginase activity.

Screening for Inhibitors of Essential Leishmania Glucose Transporters

DTIC Science & Technology

2010-07-01

TITLE: Screening for Inhibitors of Essential Leishmania Glucose Transporters PRINCIPAL INVESTIGATOR: Scott M. Landfear, Ph.D...TITLE AND SUBTITLE Screening for Inhibitors of Essential Leishmania Glucose Transporters 5a. CONTRACT NUMBER 5b. GRANT NUMBER...selective inhibitors of the essential glucose transporters of the parasite Leishmania mexicana. Toward this end, a cell growth assay has been

Screening for Inhibitors of Essential Leishmania Glucose Transporters

DTIC Science & Technology

2012-07-01

Leishmania Glucose Transporters PRINCIPAL INVESTIGATOR: Scott M. Landfear, Ph.D. CONTRACTING ORGANIZATION: Oregon Health & Science...3. DATES COVERED 1 JUL 2011 - 30 JUN 2012 4. TITLE AND SUBTITLE Screening for Inhibitors of Essential Leishmania Glucose Transporters...function as selective inhibitors of the essential glucose transporters of the parasite Leishmania mexicana. To identify such compounds, a cell growth

Prophylactic efficacy of high-molecular-weight antigenic fractions of a recent clinical isolate of Leishmania donovani against visceral leishmaniasis.

PubMed

Tripathi, P; Gupta, S K; Sinha, S; Sundar, S; Dube, A; Naik, S

2008-11-01

T-cell mediated immune responses are key determinants to the natural course of infection caused by intracellular parasites such as Leishmania. Thus, T-cell activating proteins of these microbes continue to generate active interest particularly in view of their possible role in the design and development of newer and more effective vaccines. We have recently reported the presence of T-cell immunostimulatory antigens with the high-molecular-weight (MW) fractions (134-64.2 kDa) of whole Leishmania donovani antigen (strain 2001), which stimulated variable amounts of IFN-gamma, IL-12 and IL-10 in exposed immune individuals. The present study was undertaken to further evaluate these high-MW antigenic fractions (MW range >100-60 kDa) for potential protective efficacy. The high-MW region of the parasite was resolved into five antigenic fractions (Prep A-E) using continuous elution gel electrophoresis. Prior to in vivo protection studies in hamsters, these fractions were used to evaluate in vitro cellular responses in eight Leishmania-exposed individuals and treated cured hamsters. The protective efficacy of prep (A + B), C, D and E in combination with BCG was evaluated in inbred hamsters using standard immunization protocol. Proliferative responses were seen in all eight of eight exposed individuals to prep D [median stimulation index (SI): 5.2 (range 3.9-7.1)] and E [median SI: 5.6 (range 4.4-8.2)], five of eight individuals to prep B and prep C and three of eight to prep A [median SI: 0.2 (range 0.1-7.2)]. The median proliferative responses to prep D and prep E were significantly higher than to fraction prep A; (P < 0.05) but not to prep B and prep C. However, prep A-E induced equivalent levels of IFN-gamma, IL-10 and IL-12 cytokines. Fractions D and E also exhibited marked parasite inhibition in spleen (52.5% and 73.7%) and liver (65% and 80.2%) as compared with prep (A + B) (23% in spleen and 24% in liver) and prep C (38% in spleen and 24% in liver). Prep D and prep E

The Leishmania-macrophage interaction: a metabolic perspective.

PubMed

Naderer, Thomas; McConville, Malcolm J

2008-02-01

Protozoan parasites belonging to the genus Leishmania exhibit a pronounced tropism for macrophages although they have the capacity to infect a variety of other phagocytic and non-phagocytic mammalian cells. Unlike most other intramacrophage pathogens, the major proliferative stage of Leishmania resides in the mature phagolysosomes of these host cells. In this review we highlight some of the strategies utilized by the intracellular amastigote stage of Leishmania to survive in this compartment. Remarkably, and in contrast to many other intracellular pathogens, Leishmania amastigotes have a minimalist surface glycocalyx which may facilitate uptake of essential lipids and promote exposure of phospholipids required for phagocytosis via macrophage apoptotic cell receptors. Leishmania amastigotes also differ from many other intracellular pathogens in having complex nutritional requirements which must be scavenged from the host cell. Amino acids and polyamines appear to be important carbon sources and growth-limiting nutrients, respectively, and their availability to intracellular amastigotes may be regulated by the activation state of host macrophages. Metabolic processes in both the parasite and host cell may thus be crucial determinants of disease outcome.

Vaccines for Canine Leishmaniasis

PubMed Central

Palatnik-de-Sousa, Clarisa B.

2012-01-01

Leishmaniasis is the third most important vector-borne disease worldwide. Visceral leishmaniasis (VL) is a severe and frequently lethal protozoan disease of increasing incidence and severity due to infected human and dog migration, new geographical distribution of the insect due to global warming, coinfection with immunosuppressive diseases, and poverty. The disease is an anthroponosis in India and Central Africa and a canid zoonosis (ZVL) in the Americas, the Middle East, Central Asia, China, and the Mediterranean. The ZVL epidemic has been controlled by one or more measures including the culling of infected dogs, treatment of human cases, and insecticidal treatment of homes and dogs. However, the use of vaccines is considered the most cost–effective control tool for human and canine disease. Since the severity of the disease is related to the generation of T-cell immunosuppression, effective vaccines should be capable of sustaining or enhancing the T-cell immunity. In this review we summarize the clinical and parasitological characteristics of ZVL with special focus on the cellular and humoral canine immune response and review state-of-the-art vaccine development against human and canine VL. Experimental vaccination against leishmaniasis has evolved from the practice of leishmanization with living parasites to vaccination with crude lysates, native parasite extracts to recombinant and DNA vaccination. Although more than 30 defined vaccines have been studied in laboratory models no human formulation has been licensed so far; however three second-generation canine vaccines have already been registered. As expected for a zoonotic disease, the recent preventive vaccination of dogs in Brazil has led to a reduction in the incidence of canine and human disease. The recent identification of several Leishmania proteins with T-cell epitopes anticipates development of a multiprotein vaccine that will be capable of protecting both humans and dogs against VL. PMID:22566950

In vitro additive interaction between ketoconazole and antimony against intramacrophage Leishmania (Leishmania) amazonensis amastigotes.

PubMed

Nunes, Débora Cristina de Oliveira; Bispo-da-Silva, Luiz Borges; Napolitano, Danielle Reis; Costa, Mônica Soares; Figueira, Márcia Moura Nunes Rocha; Rodrigues, Renata Santos; Rodrigues, Veridiana de Melo; Yoneyama, Kelly Aparecida Geraldo

2017-01-01

Leishmaniasis is a group of diseases caused by protozoa of Leishmania genus. The currently available treatments for this disease are expensive, present high toxicity and are associated to difficulties of healing and parasite resistance. Therefore, the development of strategies for leishmaniasis treatment is indispensable and includes reposition of existing drugs, as well as drug combination therapy. The aim of this study was to assess the nature of ketoconazole and antimony association on the cytotoxic effect against Leishmania (Leishmania) amazonensis amastigotes. The calculated mean sum of fractional 50% inhibitory concentration ([Formula: see text]ΣFIC50) was 2.54 and 1.43 for free and intracellular amastigotes, respectively, values that suggest an additive interaction between ketoconazole and antimony concerning to Leishmania toxicity only in the intramacrophage parasite form. Despite the clinical efficacy of ketoconazole-antimony combination has been shown in the literature, our study is the first to describe the nature of ketoconazole-antimony interaction against L. (L.) amazonensis amastigotes. Moreover, our results point out the need for future in vivo studies to confirm the nature of ketoconazole-antimony interaction and also to determine possible effective dosage regimens related to ketoconazole administration in association with the optimal lower dose of antimony.

Phylogenomic reconstruction supports supercontinent origins for Leishmania.

PubMed

Harkins, Kelly M; Schwartz, Rachel S; Cartwright, Reed A; Stone, Anne C

2016-03-01

Leishmania, a genus of parasites transmitted to human hosts and mammalian/reptilian reservoirs by an insect vector, is the causative agent of the human disease complex leishmaniasis. The evolutionary relationships within the genus Leishmania and its origins are the source of ongoing debate, reflected in conflicting phylogenetic and biogeographic reconstructions. This study employs a recently described bioinformatics method, SISRS, to identify over 200,000 informative sites across the genome from newly sequenced and publicly available Leishmania data. This dataset is used to reconstruct the evolutionary relationships of this genus. Additionally, we constructed a large multi-gene dataset, using it to reconstruct the phylogeny and estimate divergence dates for species. We conclude that the genus Leishmania evolved at least 90-100 million years ago, supporting a modified version of the Multiple Origins hypothesis that we call the Supercontinent hypothesis. According to this scenario, separate Leishmania clades emerged prior to, and during, the breakup of Gondwana. Additionally, we confirm that reptile-infecting Leishmania are derived from mammalian forms and that the species that infect porcupines and sloths form a clade long separated from other species. Finally, we firmly place the guinea-pig infecting species, Leishmaniaenriettii, the globally dispersed Leishmaniasiamensis, and the newly identified Australian species from a kangaroo, as sibling species whose distribution arises from the ancient connection between Australia, Antarctica, and South America. Copyright © 2015 Elsevier B.V. All rights reserved.

Vaccinating high-risk children with the intranasal live-attenuated influenza vaccine: the Quebec experience.

PubMed

Quach, Caroline

2014-12-01

Given the burden of illness associated with influenza, vaccination is recommended for individuals at high risk of complications. The live-attenuated influenza vaccine (LAIV) is administered by intranasal spray, thus directly stimulating mucosal immunity. In this review, we aimed to provide evidence for its efficacy and safety in different paediatric populations. We also share the Quebec experience of LAIV use through a publicly funded vaccination program for children with chronic, high-risk conditions. from randomized controlled trials in healthy children and in asthmatics have demonstrated superior efficacy of LAIV over the injectable vaccine (IIV). LAIV is well tolerated: its administration is associated with runny nose and nasal congestion, but not with asthma exacerbations and is well tolerated in children with cystic fibrosis, when compared to IIV. The vaccine is well accepted by children and parents and can easily be part of vaccination clinics in paediatric tertiary care centres targeting children with chronic, high-risk conditions, not leading to immunosuppression. Copyright © 2014 Elsevier Ltd. All rights reserved.

Identification of BALB/c Immune Markers Correlated with a Partial Protection to Leishmania infantum after Vaccination with a Rationally Designed Multi-epitope Cysteine Protease A Peptide-Based Nanovaccine

PubMed Central

Agallou, Maria; Margaroni, Maritsa; Athanasiou, Evita; Toubanaki, Dimitra K.; Kontonikola, Katerina; Karidi, Konstantina; Kammona, Olga; Kiparissides, Costas

2017-01-01

Background Through their increased potential to be engaged and processed by dendritic cells (DCs), nanovaccines consisting of Poly(D,L-lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) loaded with both antigenic moieties and adjuvants are attractive candidates for triggering specific defense mechanisms against intracellular pathogens. The aim of the present study was to evaluate the immunogenicity and prophylactic potential of a rationally designed multi-epitope peptide of Leishmania Cysteine Protease A (CPA160-189) co-encapsulated with Monophosphoryl lipid A (MPLA) in PLGA NPs against L. infantum in BALB/c mice and identify immune markers correlated with protective responses. Methodology/Principal Findings The DCs phenotypic and functional features exposed to soluble (CPA160-189, CPA160-189+MPLA) or encapsulated in PLGA NPs forms of peptide and adjuvant (PLGA-MPLA, PLGA-CPA160-189, PLGA-CPA160-189+MPLA) was firstly determined using BALB/c bone marrow-derived DCs. The most potent signatures of DCs maturation were obtained with the PLGA-CPA160-189+MPLA NPs. Subcutaneous administration of PLGA-CPA160-189+MPLA NPs in BALB/c mice induced specific anti-CPA160-189 cellular and humoral immune responses characterized by T cells producing high amounts of IL-2, IFN-γ and TNFα and IgG1/IgG2a antibodies. When these mice were challenged with 2x107 stationary phase L. infantum promastigotes, they displayed significant reduced hepatic (48%) and splenic (90%) parasite load at 1 month post-challenge. This protective phenotype was accompanied by a strong spleen lymphoproliferative response and high levels of IL-2, IFN-γ and TNFα versus low IL-4 and IL-10 secretion. Although, at 4 months post-challenge, the reduced parasite load was preserved in the liver (61%), an increase was detected in the spleen (30%), indicating a partial vaccine-induced protection. Conclusions/Significance This study provide a basis for the development of peptide-based nanovaccines against leishmaniasis

[Molecular typing of Leishmania (Leishmania) amazonensis and species of the subgenus Viannia associated with cutaneous and mucosal leishmaniasis in Colombia: A concordance study].

PubMed

Ovalle-Bracho, Clemencia; Camargo, Carolina; Díaz-Toro, Yira; Parra-Muñoz, Marcela

2018-03-15

Multilocus enzyme electrophoresis (MLEE) is the reference standard for the characterization of Leishmania species. The test is restricted to specialized laboratories due to its technical complexity, cost, and time required to obtain results. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) is used to identify Leishmania species. To establish the concordance between the two tests as identifying methods for circulating species in Colombia. A total of 96 isolates from patients with cutaneous or mucosal leishmaniasis were selected and identified by MLEE and PCR-RFLP with miniexon and hsp70 as the molecular targets, which were used sequentially. Restriction enzymes HaeIII and BccI were similarly applied. Cohen's kappa coefficient and the 95% confidence interval (CI) were calculated. The kappa coefficient and the 95% CI between MLEE and PCR-RFLP displayed "very good" concordance with a coefficient of 0.98 (CI95%: 0.98 to 1.00). The identified species were Leishmania Viannia braziliensis, Leishmania Viannia panamensis, Leishmania Viannia guyanensis and Leishmania Leishmania amazonensis. A total of 80 of the 96 isolates were sequenced and the results obtained by PCR-RFLP were confirmed. Due to the concordance obtained between tests results with the amplification of the genes miniexon and hsp70, PCR-RFLP is proposed as an alternative for identifying circulating Leishmania species in Colombia.

Characterisation of exacerbation risk and exacerbator phenotypes in the POET-COPD trial.

PubMed

Beeh, Kai M; Glaab, Thomas; Stowasser, Susanne; Schmidt, Hendrik; Fabbri, Leonardo M; Rabe, Klaus F; Vogelmeier, Claus F

2013-10-29

Data examining the characteristics of patients with frequent exacerbations of chronic obstructive pulmonary disease (COPD) and associated hospitalisations and mortality are scarce. Post-hoc analysis of the Prevention Of Exacerbations with Tiotropium in COPD (POET-COPD) trial, targeting exacerbations as the primary endpoint. Patients were classified as non-, infrequent, and frequent exacerbators (0, 1, or ≥ 2 exacerbations during study treatment), irrespective of study treatment. A multivariate Cox regression model assessed the effect of covariates on time to first exacerbation. In total, 7376 patients were included in the analysis: 63.5% non-exacerbators, 22.9% infrequent, 13.6% frequent exacerbators. Factors significantly associated with exacerbation risk were age, sex, body mass index, COPD duration and severity, smoking history, baseline inhaled corticosteroid use, and preceding antibiotic or systemic corticosteroid courses. Frequent exacerbators had greater severity and duration of COPD, received more pulmonary medication, and ≥ 2 systemic corticosteroid or antibiotic courses in the preceding year, and were more likely to be female and ex-smokers. The small proportion of frequent exacerbators (13.6%) accounted for 56.6% of exacerbation-related hospitalisations, which, overall, were associated with a three-fold increase in mortality. The frequent exacerbator phenotype was closely associated with exacerbation-related hospitalisations, and exacerbation-related hospitalisations were associated with poorer survival. NCT00563381; Study identifier: BI 205.389.

Comparison between three adjuvants for a vaccine against canine leishmaniasis: In vitro evaluation of macrophage killing ability.

PubMed

Trotta, T; Fasanella, A; Scaltrito, D; Gradoni, L; Mitolo, V; Brandonisio, O; Acquafredda, A; Panaro, M A

2010-03-01

The aim of this study was to evaluate, in terms of dog macrophage killing ability in vitro, a vaccine based on Leishmania infantum promastigote soluble antigen (LSA) formulated with three different adjuvants (BCG, AdjuPrime, MPL/TDM/CWS). A significant increase of the macrophage killing ability was observed in dogs vaccinated with LSA+MPL/TDM/CWS after 1 month from vaccination. A similar increase of macrophage parasitocidal ability was present only after 5 months in dogs vaccinated with LSA+BCG or LSA+AdjuPrime. In all dogs the augmented killing percentage was still present after 12 months from vaccination. Therefore, in particular LSA+MPL/TDM/CWS vaccine seems promising for further studies in dogs. 2009 Elsevier Ltd. All rights reserved.

Calmodulin Polymerase Chain Reaction–Restriction Fragment Length Polymorphism for Leishmania Identification and Typing

PubMed Central

Miranda, Aracelis; Samudio, Franklyn; González, Kadir; Saldaña, Azael; Brandão, Adeilton; Calzada, Jose E.

2016-01-01

A precise identification of Leishmania species involved in human infections has epidemiological and clinical importance. Herein, we describe a preliminary validation of a restriction fragment length polymorphism assay, based on the calmodulin intergenic spacer region, as a tool for detecting and typing Leishmania species. After calmodulin amplification, the enzyme HaeIII yielded a clear distinction between reference strains of Leishmania mexicana, Leishmania amazonensis, Leishmania infantum, Leishmania lainsoni, and the rest of the Viannia reference species analyzed. The closely related Viannia species: Leishmania braziliensis, Leishmania panamensis, and Leishmania guyanensis, are separated in a subsequent digestion step with different restriction enzymes. We have developed a more accessible molecular protocol for Leishmania identification/typing based on the exploitation of part of the calmodulin gene. This methodology has the potential to become an additional tool for Leishmania species characterization and taxonomy. PMID:27352873

Leishmania infections: Molecular targets and diagnosis.

PubMed

Akhoundi, Mohammad; Downing, Tim; Votýpka, Jan; Kuhls, Katrin; Lukeš, Julius; Cannet, Arnaud; Ravel, Christophe; Marty, Pierre; Delaunay, Pascal; Kasbari, Mohamed; Granouillac, Bruno; Gradoni, Luigi; Sereno, Denis

2017-10-01

Progress in the diagnosis of leishmaniases depends on the development of effective methods and the discovery of suitable biomarkers. We propose firstly an update classification of Leishmania species and their synonymies. We demonstrate a global map highlighting the geography of known endemic Leishmania species pathogenic to humans. We summarize a complete list of techniques currently in use and discuss their advantages and limitations. The available data highlights the benefits of molecular markers in terms of their sensitivity and specificity to quantify variation from the subgeneric level to species complexes, (sub) species within complexes, and individual populations and infection foci. Each DNA-based detection method is supplied with a comprehensive description of markers and primers and proposal for a classification based on the role of each target and primer in the detection, identification and quantification of leishmaniasis infection. We outline a genome-wide map of genes informative for diagnosis that have been used for Leishmania genotyping. Furthermore, we propose a classification method based on the suitability of well-studied molecular markers for typing the 21 known Leishmania species pathogenic to humans. This can be applied to newly discovered species and to hybrid strains originating from inter-species crosses. Developing more effective and sensitive diagnostic methods and biomarkers is vital for enhancing Leishmania infection control programs. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Identification of L. infantum chagasi proteins in VL patients' urine: a promising antigen discovery approach of vaccine candidates

PubMed Central

Kashino, Suely S.; Abeijon, Claudia; Qin, Lizeng; Kanunfre, Kelly A.; Kubrusly, Flávia S.; Silva, Fernando O.; Costa, Dorcas L.; Campos, Dioclécio; Costa, Carlos H.N.; Raw, Isaias; Campos-Neto, Antonio

2012-01-01

Visceral leishmaniasis (VL) is a serious lethal parasitic disease caused by Leishmania donovani in Asia and by Leishmania infantum chagasi in Southern Europe and South America. VL is endemic in 47 countries with an annual incidence estimated to be 500,000 cases. This high incidence is due in part to the lack of an efficacious vaccine. Here, we introduce an innovative approach to directly identify parasite vaccine candidate antigens that are abundantly produced in vivo in humans with VL. We combined RP-HPLC and mass spectrometry and categorized three L. infantum chagasi proteins, presumably produced in spleen, liver, and bone marrow lesions and excreted in the patients’ urine. Specifically, these proteins were the following: Li-isd1 (XP_001467866.1), Li-txn1 (XP_001466642.1), and Li-ntf2 (XP_001463738.1). Initial vaccine validation studies were performed with the rLi-ntf2 protein produced in E. coli mixed with the adjuvant BpMPLA-SE. This formulation stimulated potent Th1 response in BALB/c mice. Compared to control animals, mice immunized with Li-ntf2 + BpMPLASE had a marked parasite burden reduction in spleens at 40 days post-challenge with virulent L. infantum chagasi. These results strongly support the proposed antigen discovery strategy of vaccine candidates to kala-azar and opens novel possibilities for vaccine development to other serious infectious diseases. PMID:22443237

The TryPIKinome of five human pathogenic trypanosomatids: Trypanosoma brucei, Trypanosoma cruzi, Leishmania major, Leishmania braziliensis and Leishmania infantum--new tools for designing specific inhibitors.

PubMed

Bahia, Diana; Oliveira, Luciana Márcia; Lima, Fabio Mitsuo; Oliveira, Priscila; Silveira, José Franco da; Mortara, Renato Arruda; Ruiz, Jerônimo Conceição

2009-12-18

Phosphatidylinositol (PI) kinases are at the heart of one of the major pathways of intracellular signal transduction. Herein, we present the first report on a survey made by similarity searches against the five human pathogenic trypanosomatids Trypanosoma brucei, Trypanosoma cruzi, Leishmania major, Leishmania braziliensis and Leishmania infantum genomes available to date for phosphatidylinositol- and related-kinases (TryPIKs). In addition to generating a panel called "The TryPIKinome", we propose a model of signaling pathways for these TryPIKs. The involvement of TryPIKs in fundamental pathways, such as intracellular signal transduction and host invasion processes, makes the study of TryPIKs an important area for further inquiry. New subtype-specific inhibitors are expected to work on individual members of the PIK family and, therefore, can presumably neutralize trypanosomatid invasion processes.

Evaluation of methylene blue as photosensitizer in promastigotes of Leishmania major and Leishmania braziliensis.

PubMed

Pinto, Juliana Guerra; Martins, Jaciara Fagundes de Souza; Pereira, André Henrique Correia; Mittmann, Josane; Raniero, Leandro José; Ferreira-Strixino, Juliana

2017-06-01

The cutaneous leishmaniasis is caused by the protozoan of the genus Leishmania. It is considered by WHO as a public health issue and a neglected disease, which affects rural workers and it is also a risk to travelers in endemic areas. The conventional treatment is toxic and leads to severe side effects. The photodynamic therapy has been studied as an alternative treatment to cutaneous leishmaniasis. This study aimed to evaluate the methylene blue internalization and the impact of the PDT in the viability and morphology of Leishmania major and Leishmania braziliensis promastigote in culture medium. The fluorescence microscopy was used to determine the MB localization. To evaluate the mitochondrial activity (MTT), viability (Trypan blue test) and the morphological alterations both species were incubated with the MB in concentrations starting in 500μg/ml, in serial dilution, until 7,8μg/ml. The fluorescence microscopy demonstrated that the MB is internalized by both species after one hour of incubation. The MB presented low toxicity at the dark and the PDT was capable of decreasing the viability in more than 70% in the higher concentrations tested. The PDT also triggered significant morphological alterations in the Leishmania promastigotes. The results presented in this study are an indicative that the MB is a photosensitizer with promising potential to clinical application, besides its low cost. Copyright © 2017 Elsevier B.V. All rights reserved.

Physalis angulata induces death of promastigotes and amastigotes of Leishmania (Leishmania) amazonensis via the generation of reactive oxygen species.

PubMed

Da Silva, B J M; Da Silva, R R P; Rodrigues, A P D; Farias, L H S; Do Nascimento, J L M; Silva, E O

2016-03-01

Leishmaniasis are a neglected group of emerging diseases that have been found in 98 countries and are caused by protozoa of the genus Leishmania. The therapy for leishmaniasis causes several side effects and leads to drug-resistant strains. Natural products from plants have exhibited activities against Leishmania in various experimental models. Physalis angulata is a widely used plant in popular medicine, and in the literature it has well-documented leishmanicidal activity. However, its mechanism of action is still unknown. Thus, this study aims to evaluate the mechanism driving the leishmanicidal activity of an aqueous extract of P. angulata root (AEPa). AEPa was effective against both promastigotes and intracellular amastigote forms of Leishmania amazonensis. This effect was mediated by an increase of reactive oxygen species (ROS), but not of nitric oxide (NO). The increased production of ROS induces cell death by phenotypes seems by apoptosis cell death in Leishmania, but not autophagy or necrosis. In addition, morphological analysis of macrophages showed that AEPa induced a high number of cytoplasmic projections, increased the volume of cytoplasm and number of vacuoles, caused cytoskeleton alterations and resulted in high spreading ability. AEPa also promoted superoxide anion (O2(-)) production in both uninfected macrophages and those infected with Leishmania. Therefore, these results revealed that AEPa causes cell death by phenotypes seems by apoptosis cell death in L. amazonensis and modulates macrophage activation through morphofunctional alterations and O2(-) generation to induce Leishmania death. Copyright © 2015 Elsevier Ltd. All rights reserved.

Quantification of anti-Leishmania antibodies in saliva of dogs.

PubMed

Cantos-Barreda, Ana; Escribano, Damián; Bernal, Luis J; Cerón, José J; Martínez-Subiela, Silvia

2017-08-15

Detection of serum anti-Leishmania antibodies by quantitative or qualitative techniques has been the most used method to diagnose Canine Leishmaniosis (CanL). Nevertheless, saliva may represent an alternative to blood because it is easy to collect, painless and non-invasive in comparison with serum. In this study, two time-resolved immunofluorometric assays (TR-IFMAs) for quantification of anti-Leishmania IgG2 and IgA antibodies in saliva were developed and validated and their ability to distinguish Leishmania-seronegative from seropositive dogs was evaluated. The analytical study was performed by evaluation of assay precision, sensitivity and accuracy. In addition, serum from 48 dogs (21 Leishmania-seropositive and 27 Leishmania-seronegative) were analyzed by TR-IFMAs. The assays were precise, with an intra- and inter-assay coefficients of variation lower than 11%, and showed high level of accuracy, as determined by linearity under dilution (R 2 =0.99) and recovery tests (>88.60%). Anti-Leishmania IgG2 antibodies in saliva were significantly higher in the seropositive group compared with the seronegative (p<0.0001), whereas no significant differences for anti-Leishmania IgA antibodies between both groups were observed. Furthermore, TR-IFMA for quantification of anti-Leishmania IgG2 antibodies in saliva showed higher differences between seropositive and seronegative dogs than the commercial assay used in serum. In conclusion, TR-IFMAs developed may be used to quantify anti-Leishmania IgG2 and IgA antibodies in canine saliva with an adequate precision, analytical sensitivity and accuracy. Quantification of anti-Leishmania IgG2 antibodies in saliva could be potentially used to evaluate the humoral response in CanL. However, IgA in saliva seemed not to have diagnostic value for this disease. For future studies, it would be desirable to evaluate the ability of the IgG2 assay to detect dogs with subclinical disease or with low antibody titers in serum and also to study

Conjugation, characterization and toxicity of lipophosphoglycan-polyacrylic acid conjugate for vaccination against leishmaniasis.

PubMed

Topuzogullari, Murat; Cakir Koc, Rabia; Dincer Isoglu, Sevil; Bagirova, Melahat; Akdeste, Zeynep; Elcicek, Serhat; Oztel, Olga N; Yesilkir Baydar, Serap; Canim Ates, Sezen; Allahverdiyev, Adil M

2013-06-03

Research on the conjugates of synthetic polyelectrolytes with antigenic molecules, such as proteins, peptides, or carbohydrates, is an attractive area due to their highly immunogenic character in comparison to classical adjuvants. For example, polyacrylic acid (PAA) is a weak polyelectrolyte and has been used in several biomedical applications such as immunological studies, drug delivery, and enzyme immobilization. However, to our knowledge, there are no studies that document immune-stimulant properties of PAA in Leishmania infection. Therefore, we aimed to develop a potential vaccine candidate against leishmaniasis by covalently conjugating PAA with an immunologically vital molecule of lipophosphoglycan (LPG) found in Leishmania parasites. In the study, LPG and PAA were conjugated by a multi-step procedure, and final products were analyzed with GPC and MALDI-TOF MS techniques. In cytotoxicity experiments, LPG-PAA conjugates did not indicate toxic effects on L929 and J774 murine macrophage cells. We assume that LPG-PAA conjugate can be a potential vaccine candidate, and will be immunologically characterized in further studies to prove its potential.

Use of Phage Display technology in development of canine visceral leishmaniasis vaccine using synthetic peptide trapped in sphingomyelin/cholesterol liposomes.

PubMed

Toledo-Machado, Christina Monerat; Bueno, Lilian Lacerda; Menezes-Souza, Daniel; Machado-de-Avila, Ricardo Andrez; Nguyen, Christophe; Granier, Claude; Bartholomeu, Daniella Castanheira; Chávez-Olórtegui, Carlos; Fujiwara, Ricardo Toshio

2015-02-28

Leishmania parasites can cause visceral or cutaneous disease and are found in subtropical and tropical regions of the Old and New World. The pathology of the infection is determined by both host immune factors and species/strain differences of the parasite. Dogs represent the major reservoir of Leishmania infantum (syn. L. chagasi) and vaccines are considered the most cost-effective control tools for canine disease. Selection of immunodominant peptides was performed by Phage Display to identify sequences recognized by L. infantum naturally infected animals. Sera from Leishmania infected animals were used in the biopanning to selection of specific peptides. Serum samples from T. cruzi infected and healthy animals were used as control. After selection, synthetic peptides were produced in membrane (spot-synthesis) in soluble form and blotting and ELISA were performed for validation of serum reactivity. Selected peptide was formulated with aluminum hydroxide and liposomes and immunization was performed in BALB/c mice. Protection was determined by qPCR after challenge infection with virulent L. infantum. We reported the selection of Peptide 5 through Phage Display technique and demonstrate its ability to promote a state of immunity against L. infantum infection in murine model after immunization using liposomes as vaccine carrier. Our results demonstrate that immunization with Peptide 5 when formulated with aluminum hydroxide and liposomes is immunogenic and elicited significant protection associated with the induction of mixed Th1/Th2 immune response against L. infantum infection. Peptide 5 is a promising vaccine candidate and the findings obtained in the present study encourage canine trials to confirm the effectiveness of a vaccine against CVL.

The genome sequence of Leishmania (Leishmania) amazonensis: functional annotation and extended analysis of gene models.

PubMed

Real, Fernando; Vidal, Ramon Oliveira; Carazzolle, Marcelo Falsarella; Mondego, Jorge Maurício Costa; Costa, Gustavo Gilson Lacerda; Herai, Roberto Hirochi; Würtele, Martin; de Carvalho, Lucas Miguel; Carmona e Ferreira, Renata; Mortara, Renato Arruda; Barbiéri, Clara Lucia; Mieczkowski, Piotr; da Silveira, José Franco; Briones, Marcelo Ribeiro da Silva; Pereira, Gonçalo Amarante Guimarães; Bahia, Diana

2013-12-01

We present the sequencing and annotation of the Leishmania (Leishmania) amazonensis genome, an etiological agent of human cutaneous leishmaniasis in the Amazon region of Brazil. L. (L.) amazonensis shares features with Leishmania (L.) mexicana but also exhibits unique characteristics regarding geographical distribution and clinical manifestations of cutaneous lesions (e.g. borderline disseminated cutaneous leishmaniasis). Predicted genes were scored for orthologous gene families and conserved domains in comparison with other human pathogenic Leishmania spp. Carboxypeptidase, aminotransferase, and 3'-nucleotidase genes and ATPase, thioredoxin, and chaperone-related domains were represented more abundantly in L. (L.) amazonensis and L. (L.) mexicana species. Phylogenetic analysis revealed that these two species share groups of amastin surface proteins unique to the genus that could be related to specific features of disease outcomes and host cell interactions. Additionally, we describe a hypothetical hybrid interactome of potentially secreted L. (L.) amazonensis proteins and host proteins under the assumption that parasite factors mimic their mammalian counterparts. The model predicts an interaction between an L. (L.) amazonensis heat-shock protein and mammalian Toll-like receptor 9, which is implicated in important immune responses such as cytokine and nitric oxide production. The analysis presented here represents valuable information for future studies of leishmaniasis pathogenicity and treatment.

The Genome Sequence of Leishmania (Leishmania) amazonensis: Functional Annotation and Extended Analysis of Gene Models

PubMed Central

Real, Fernando; Vidal, Ramon Oliveira; Carazzolle, Marcelo Falsarella; Mondego, Jorge Maurício Costa; Costa, Gustavo Gilson Lacerda; Herai, Roberto Hirochi; Würtele, Martin; de Carvalho, Lucas Miguel; e Ferreira, Renata Carmona; Mortara, Renato Arruda; Barbiéri, Clara Lucia; Mieczkowski, Piotr; da Silveira, José Franco; Briones, Marcelo Ribeiro da Silva; Pereira, Gonçalo Amarante Guimarães; Bahia, Diana

2013-01-01

We present the sequencing and annotation of the Leishmania (Leishmania) amazonensis genome, an etiological agent of human cutaneous leishmaniasis in the Amazon region of Brazil. L. (L.) amazonensis shares features with Leishmania (L.) mexicana but also exhibits unique characteristics regarding geographical distribution and clinical manifestations of cutaneous lesions (e.g. borderline disseminated cutaneous leishmaniasis). Predicted genes were scored for orthologous gene families and conserved domains in comparison with other human pathogenic Leishmania spp. Carboxypeptidase, aminotransferase, and 3′-nucleotidase genes and ATPase, thioredoxin, and chaperone-related domains were represented more abundantly in L. (L.) amazonensis and L. (L.) mexicana species. Phylogenetic analysis revealed that these two species share groups of amastin surface proteins unique to the genus that could be related to specific features of disease outcomes and host cell interactions. Additionally, we describe a hypothetical hybrid interactome of potentially secreted L. (L.) amazonensis proteins and host proteins under the assumption that parasite factors mimic their mammalian counterparts. The model predicts an interaction between an L. (L.) amazonensis heat-shock protein and mammalian Toll-like receptor 9, which is implicated in important immune responses such as cytokine and nitric oxide production. The analysis presented here represents valuable information for future studies of leishmaniasis pathogenicity and treatment. PMID:23857904

Humoral and In Vivo Cellular Immunity against the Raw Insect-Derived Recombinant Leishmania infantum Antigens KMPII, TRYP, LACK, and papLe22 in Dogs from an Endemic Area

PubMed Central

Todolí, Felicitat; Solano-Gallego, Laia; de Juan, Rafael; Morell, Pere; del Carmen Núñez, Maria; Lasa, Rodrigo; Gómez-Sebastián, Silvia; Escribano, José M.; Alberola, Jordi; Rodríguez-Cortés, Alhelí

2010-01-01

Leishmania infantum causes visceral leishmaniasis, a severe zoonotic and systemic disease that is fatal if left untreated. Identification of the antigens involved in Leishmania-specific protective immune response is a research priority for the development of effective control measures. For this purpose, we evaluated, in 27 dogs from an enzootic zone, specific humoral and cellular immune response by delayed-type hypersensitivity (DTH) skin test both against total L. infantum antigen and the raw Trichoplusia ni insect-derived kinetoplastid membrane protein-11 (rKMPII), tryparedoxin peroxidase (rTRYP), Leishmania homologue of receptors for activated C kinase (rLACK), and 22-kDa potentially aggravating protein of Leishmania (rpapLe22) antigens from this parasite. rTRYP induced the highest number of positive DTH responses (55% of leishmanin skin test [LST]-positive dogs), showing that TRYP antigen is an important T cell immunogen, and it could be a promising vaccine candidate against this disease. When TRYP-DTH and KMPII-DTH tests were evaluated in parallel, 82% of LST-positive dogs were detected, suggesting that both antigens could be considered as components of a standardized DTH immunodiagnostic tool for dogs. PMID:21118936

Cryopreservation of Leishmania Species in Manisa Province.

PubMed

Çavuş, İbrahim; Ocak, Fulya; Kaya, Tuğba; Özbilgin, Ahmet

2017-09-01

It was aimed to assess the success of the cryopreservation process which is carried out in order to preserve the genetic material and the virulence of the Leishmania species that are an important health problem in our region. Leishmania tropica, L. infantum, L. major, and L. donovani strains in Novy-MacNeal-Nicolle (NNN) medium in MCBU were used. Promastigotes cultured in the NNN medium were transferred to RPMI 1640 medium; promastigotes in the logarithmic phase were washed three times with PBS, and 15% dimethylsulfoxide (DMSO) was added. Leishmania species were transferred to 12 separate tubes. The tubes were stored at -86°C for one night by placing them in Coolcell boxes. The tubes were transferred into a liquid nitrogen tank. One cryotube per Leishmania strain is thawed monthly and cultured in NNN medium. For the duration of study it was observed that each Leishmania isolate preserved 60-65% of their viability and entered the logarithmic phase on the 7th day following the inoculation in the NNN medium. Abnormalities in the structures and movements of the promastigotes were not observed in microscopic examinations. The following conclusions were made: cryopreservation is important for studies planned related to leishmaniasis and cryopreservation with DMSO is successful.

Understanding serine proteases implications on Leishmania spp lifecycle.

PubMed

Alves, Carlos Roberto; Souza, Raquel Santos de; Charret, Karen Dos Santos; Côrtes, Luzia Monteiro de Castro; Sá-Silva, Matheus Pereira de; Barral-Veloso, Laura; Oliveira, Luiz Filipe Gonçalves; da Silva, Franklin Souza

2018-01-01

Serine proteases have significant functions over a broad range of relevant biological processes to the Leishmania spp lifecycle. Data gathered here present an update on the Leishmania spp serine proteases and the status of these enzymes as part of the parasite degradome. The serine protease genes (n = 26 to 28) in Leishmania spp, which encode proteins with a wide range of molecular masses (35 kDa-115 kDa), are described along with their degrees of chromosomal and allelic synteny. Amid 17 putative Leishmania spp serine proteases, only ∼18% were experimentally demonstrated, as: signal peptidases that remove the signal peptide from secretory pre-proteins, maturases of other proteins and with metacaspase-like activity. These enzymes include those of clans SB, SC and SF. Classical inhibitors of serine proteases are used as tools for the characterization and investigation of Leishmania spp. Endogenous serine protease inhibitors, which are ecotin-like, can act modulating host actions. However, crude or synthetic based-natural serine protease inhibitors, such as potato tuber extract, Stichodactyla helianthus protease inhibitor I, fukugetin and epoxy-α-lapachone act on parasitic serine proteases and are promising leishmanicidal agents. The functional interrelationship between serine proteases and other Leishmania spp proteins demonstrate essential functions of these enzymes in parasite physiology and therefore their value as targets for leishmaniasis treatment. Copyright © 2017 Elsevier Inc. All rights reserved.

Advances in our understanding of immunization and vaccines for patients with systemic lupus erythematosus.

PubMed

Bragazzi, Nicola Luigi; Watad, Abdulla; Sharif, Kassem; Adawi, Mohammad; Aljadeff, Gali; Amital, Howard; Shoenfeld, Yehuda

2017-10-01

Systemic lupus erythematosus (SLE) is a chronic systemic autoimmune disease. In SLE, immune system dysfunction is postulated to result by virtue of the disease itself as well as by the impact of treatment modalities employed. A myriad of immune dysregulations occur including complement system dysfunction among others. Infectious agents are known to complicate the disease course in close to 25-45% of SLE patients. Areas covered: In this review a discussion of the immunogenicity and safety of viral and bacterial vaccinations in SLE was performed. The search included ISI Web of Science (WoS), Scopus, MEDLINE/PubMed, Google-Scholar, DOAJ, EbscoHOST, Scirus, Science Direct, Cochrane Library and ProQuest. Proper string made up of a key-words including 'SLE', 'vaccination', 'safety' and 'efficacy' was used. Expert commentary: Vaccination of SLE patients is proven to be immunogenic. Concerns regarding vaccine safety are postulated, yet no direct relationship between vaccination and disease exacerbation were established. While live virus vaccines are generally contraindicated in immunosuppressive states, generally live attenuated vaccinations are recommended in SLE patients on a case-to-case basis. In SLE patients, clinical parameters such as vaccination during disease exacerbations have not been intensively studied and therefore while apparently safe, vaccination is generally recommended while disease is quiescent.

Pathways of iron acquisition and utilization in Leishmania

PubMed Central

Flannery, Andrew R.; Renberg, Rebecca L.; Andrews, Norma W.

2013-01-01

Iron is essential for many metabolic pathways, but is toxic in excess. Recent identification of the ferric iron reductase LFR1, the ferrous iron transporter LIT1, and the heme transporter LHR1 greatly advanced our understanding of how Leishmania parasites acquire iron and regulate its uptake. LFR1 and LIT1 have close orthologs in plants, and are required for Leishmania virulence. Consistent with the lack of heme biosynthesis in trypanosomatids, LHR1 and LABCG5, a protein involved in heme salvage from hemoglobin, seem essential for Leishmania survival. LFR1, LIT1 and LHR1 are upregulated under low iron availability, in agreement with the need to prevent excessive iron uptake. Future studies should clarify how Leishmania interacts with the iron homeostasis machinery of its host cell, the macrophage. PMID:23962817

Prevalence and risk factors associated with Leishmania infection in Trang Province, southern Thailand.

PubMed

Manomat, Jipada; Leelayoova, Saovanee; Bualert, Lertwut; Tan-Ariya, Peerapan; Siripattanapipong, Suradej; Mungthin, Mathirut; Naaglor, Tawee; Piyaraj, Phunlerd

2017-11-01

Autochthonous cutaneous and visceral leishmaniasis (VL) caused by Leishmania martiniquensis and Leishmania siamensis have been considered emerging infectious diseases in Thailand. The disease burden is significantly underestimated, especially the prevalence of Leishmania infection among HIV-positive patients. A cross-sectional study was conducted to determine the prevalence and risk factors associated with Leishmania infection among patients with HIV/AIDS living in Trang province, southern Thailand, between 2015 and 2016. Antibodies against Leishmania infection were assayed using the direct agglutination test (DAT). DNA of Leishmania was detected by ITS1-PCR using the buffy coat. Species of Leishmania were also identified. Of 724 participants, the prevalence of Leishmania infection was 25.1% (182/724) using either DAT or PCR assays. Seroprevalence of Leishmania infection was 18.5% (134/724), while Leishmania DNA detected by the PCR method was 8.4% (61/724). Of these, 24.9% (180/724) were asymptomatic, whereas 0.3% (2/724) were symptomatic VL and VL/CL (cutaneous leishmaniasis). At least five species were identified: L. siamensis, L. martiniquensis, L. donovani complex, L. lainsoni, and L. major. Multivariate analysis showed that CD4+ levels <500 cells/μL and living in stilt houses were independently associated with Leishmania infection. Those who were PCR positive for Leishmania DNA were significantly associated with a detectable viral load, whereas non-injection drug use (NIDU) and CD4+ levels <500 cells/μL were potential risk factors of Leishmania seropositivity. A magnitude of the prevalence of underreporting Leishmania infection among Thai patients with HIV was revealed in this study. Effective public health policy to prevent and control disease transmission is urgently needed.

Immunity to LuloHya and Lundep, the salivary spreading factors from Lutzomyia longipalpis, protects against Leishmania major infection

PubMed Central

Amo, Laura; Moore, Ian N.; DeSouza-Vieira, Thiago S.; Sanchez, Elda E.; Suntravat, Montamas; Ribeiro, Jose M. C.

2018-01-01

Salivary components from disease vectors help arthropods to acquire blood and have been shown to enhance pathogen transmission in different model systems. Here we show that two salivary enzymes from Lutzomyia longipalpis have a synergist effect that facilitates a more efficient blood meal intake and diffusion of other sialome components. We have previously shown that Lundep, a highly active endonuclease, enhances parasite infection and prevent blood clotting by inhibiting the intrinsic pathway of coagulation. To investigate the physiological role of a salivary hyaluronidase in blood feeding we cloned and expressed a recombinant hyaluronidase from Lu. longipalpis. Recombinant hyaluronidase (LuloHya) was expressed in mammalian cells and biochemically characterized in vitro. Our study showed that expression of neutrophil CXC chemokines and colony stimulating factors were upregulated in HMVEC cells after incubation with LuloHya and Lundep. These results were confirmed by the acute hemorrhage, edema and inflammation in a dermal necrosis (dermonecrotic) assay involving a massive infiltration of leukocytes, especially neutrophils, in mice co-injected with hemorrhagic factor and these two salivary proteins. Moreover, flow cytometry results showed that LuloHya and Lundep promote neutrophil recruitment to the bite site that may serve as a vehicle for establishment of Leishmania infection. A vaccination experiment demonstrated that LuloHya and Lundep confer protective immunity against cutaneous leishmaniasis using the Lu. longipalpis—Leishmania major combination as a model. Animals (C57BL/6) immunized with LuloHya or Lundep showed minimal skin damage while lesions in control animals remained ulcerated. This protective immunity was abrogated when B-cell-deficient mice were used indicating that antibodies against both proteins play a significant role for disease protection. Rabbit-raised anti-LuloHya antibodies completely abrogated hyaluronidase activity in vitro. Moreover

Arginase activity of Leishmania isolated from patients with cutaneous leishmaniasis.

PubMed

Badirzadeh, A; Taheri, T; Abedi-Astaneh, F; Taslimi, Y; Abdossamadi, Z; Montakhab-Yeganeh, H; Aghashahi, M; Niyyati, M; Rafati, S

2017-09-01

Cutaneous leishmaniasis (CL) is one of the most important vector-borne parasitic diseases, highly endemic in Iran, and its prevalence is increasing all over the country. Arginase (ARG) activity in isolated Leishmania parasites from CL patients is yet to be explored. This study aimed to compare the ARG activity of isolated Leishmania promastigotes from CL patients with a standard strain of Leishmania major and its influences on the disease pathogenesis. We recruited 16 confirmed CL patients from Qom Province, in central Iran; after detection of Leishmania species using PCR-RFLP, we assessed the levels of ARG in the isolated promastigotes and determined the parasites' growth rate. Only L. major was identified from CL patients. The level of ARG activity in the isolated Leishmania promastigotes from CL patients was significantly higher than that obtained from the standard strain of L. major. No significant correlations between ARG activity and lesion size, number or duration were observed; in contrast, a significant negative correlation was seen between ARG level and Leishmania' growth rate. The obtained results suggest that increased ARG expression and activity in the isolated Leishmania promastigotes might contribute to the higher parasite infectivity and play a major role in the pathogenicity of the CL. © 2017 John Wiley & Sons Ltd.

Re-designing the Mozambique vaccine supply chain to improve access to vaccines.

PubMed

Lee, Bruce Y; Haidari, Leila A; Prosser, Wendy; Connor, Diana L; Bechtel, Ruth; Dipuve, Amelia; Kassim, Hidayat; Khanlawia, Balbina; Brown, Shawn T

2016-09-22

Populations and routine childhood vaccine regimens have changed substantially since supply chains were designed in the 1980s, and introducing new vaccines during the "Decade of Vaccine" may exacerbate existing bottlenecks, further inhibiting the flow of all vaccines. Working with the Mozambique Ministry of Health, our team implemented a new process that integrated HERMES computational simulation modeling and on-the-ground implementers to evaluate and improve the Mozambique vaccine supply chain using a system-re-design that integrated new supply chain structures, information technology, equipment, personnel, and policies. The alternative system design raised vaccine availability (from 66% to 93% in Gaza; from 76% to 84% in Cabo Delgado) and reduced the logistics cost per dose administered (from $0.53 to $0.32 in Gaza; from $0.38 to $0.24 in Cabo Delgado) as compared to the multi-tiered system under the current EPI. The alternative system also produced higher availability at lower costs after new vaccine introductions. Since reviewing scenarios modeling deliveries every two months in the north of Gaza, the provincial directorate has decided to pilot this approach diverging from decades of policies dictating monthly deliveries. Re-design improved not only supply chain efficacy but also efficiency, important since resources to deliver vaccines are limited. The Mozambique experience and process can serve as a model for other countries during the Decade of Vaccines. For the Decade of Vaccines, getting vaccines at affordable prices to the market is not enough. Vaccines must reach the population to be successful. Copyright © 2016. Published by Elsevier Ltd.

Self-reported influenza vaccination and protective serum antibody titers in a cohort of COPD patients.

PubMed

Eagan, T M; Hardie, J A; Jul-Larsen, Å; Grydeland, T B; Bakke, P S; Cox, R J

2016-06-01

COPD patients are advised vaccination against seasonal influenza, yet few studies have evaluated the protective antibody titers obtained in this patient group. 1) To describe protective titers in COPD patients who self-reported influenza vaccination compared with vaccinated subjects without COPD and unvaccinated COPD patients, 2) analyze whether clinical parameters predicted influenza-specific antibody titers, and 3) whether antibody titers to influenza A at baseline could predict exacerbation risk or 5 years all-cause mortality. Influenza A (H1N1 and H3N2) titers were measured by haemagglutination inhibition assay in serum from 432 COPD patients and 77 controls in the Bergen COPD Cohort Study, at yearly visits between 2006/09. Titers of 40 or above were considered protective. We examined the variables sex, age, body composition, smoking, GOLD stage, yearly exacerbations, inhaled steroids, and Charlson score as predictive of titers, both univariately and in a multivariable model estimated by generalized estimating equations. The exacerbation incidence rate ratios and mortality hazard ratios were assessed by negative binominal and cox regression models respectively. At baseline, 59% of COPD patients reported influenza vaccination during the last season. Levels of predictive titers varied considerably each season, but trended lower in COPD patients compared with controls. Neither sex, age, body composition, smoking, comorbidities, GOLD stage nor use of inhaled steroids consistently predicted titers. Having high titers at baseline did not impact later risk for exacerbations, but seemed to be associated with higher all-cause mortality, even after adjustment for COPD disease characteristics. Vaccination coverage for influenza is imperfect for COPD patients in Norway, and there is a concern that immunization is suboptimal. Copyright © 2016 Elsevier Ltd. All rights reserved.

Leishmania cell surface prohibitin: role in host-parasite interaction.

PubMed

Jain, Rohit; Ghoshal, Angana; Mandal, Chitra; Shaha, Chandrima

2010-04-01

Proteins selectively upregulated in infective parasitic forms could be critical for disease pathogenesis. A mammalian prohibitin orthologue is upregulated in infective metacyclic promastigotes of Leishmania donovani, a parasite that causes visceral leishmaniasis. Leishmania donovani prohibitin shares 41% similarity with mammalian prohibitin and 95-100% within the genus. Prohibitin is concentrated at the surface of the flagellar and the aflagellar pole, the aflagellar pole being a region through which host-parasite interactions occur. Prohibitin is attached to the membrane through a GPI anchor. Overexpression of wild-type prohibitin increases protein surface density resulting in parasites with higher infectivity. However, parasites overexpressing a mutant prohibitin with an amino acid substitution at the GPI anchor site to prevent surface expression through GPI-link show lesser surface expression and lower infective abilities. Furthermore, the presence of anti-prohibitin antibodies during macrophage-Leishmania interaction in vitro reduces infection. The cognate binding partner for Leishmania prohibitin on the host cell appears to be macrophage surface HSP70, siRNA mediated downregulation of which abrogates the capability of the macrophage to bind to parasites. Leishmania prohibitin is able to generate a strong humoral response in visceral leishmaniasis patients. The above observations suggest that prohibitin plays an important role in events leading to Leishmania-host interaction.

Calcein+/PI- as an early apoptotic feature in Leishmania.

PubMed

Basmaciyan, Louise; Azas, Nadine; Casanova, Magali

2017-01-01

Although leishmaniases are responsible for high morbidity and mortality all over the world, no really satisfying treatment exists. Furthermore, the corresponding parasite Leishmania undergoes a very characteristic form of programmed cell death. Indeed, different stimuli can induce morphological and biochemical apoptotic-like features. However, the key proteins involved in mammal apoptosis, such as caspases and death receptors, are not encoded in the genome of this parasite. Currently, little is known about Leishmania apoptosis, notably owing to the lack of specific tools for programmed cell death analysis in these parasites. Furthermore, there is a need for a better understanding of Leishmania programmed cell death in order (i) to better understand the role of apoptosis in unicellular organisms, (ii) to better understand apoptosis in general through the study of an ancestral eukaryote, and (iii) to identify new therapeutic targets against leishmaniases. To advance understanding of apoptosis in Leishmania, in this study we developed a new tool based on the quantification of calcein and propidium iodide by flow cytometry. This double labeling can be employed to distinguish early apoptosis, late apoptosis and necrosis in Leishmania live cells with a very simple and rapid assay. This paper should, therefore, be of interest for people working on Leishmania and related parasites.

Immunization with Leishmania donovani protein disulfide isomerase DNA construct induces Th1 and Th17 dependent immune response and protection against experimental visceral leishmaniasis in Balb/c mice.

PubMed

Amit, Ajay; Vijayamahantesh; Dikhit, Manas R; Singh, Ashish Kumar; Kumar, Vikash; Suman, Shashi S; Singh, Ashu; Kumar, Akhilesh; Thakur, Ajit Kumar; Das, Vidyanand Ravi; Das, Pradeep; Bimal, Sanjiva

2017-02-01

In the present study, the efficacy of Leishmania donovani protein disulfide isomerase (LdPDI) as a DNA vaccine was evaluated in BALB/C mice. Mice immunized with the LdPDI-DNA construct were found to be the most immuno-reactive, as the construct induced higher T-cell proliferation. The increased T-cell proliferation was associated with a substantial rise in Th1 and Th17+ CD4 cell response and triggered a higher proportion of CD8+ T cells for the release of interferon-gamma along with a reduced splenic parasite load on Days20 and 60 post challenge (PC). Furthermore, the vaccine construct triggered increased interferon (IFN)-γ, interleukin(IL)-17A, and IL-22 release accompanied by decreased extracellular signal-regulated kinases (ERK) 1/2 signaling and increased mitogen-activated protein kinase (MAPK) signaling coinciding with an increase in the amount of nitrite and reactive oxygen species (ROS)in vaccinating the splenocyts. We summarize from our data that the PDI-DNA construct of Leishmania donovani has the potential to elicit protective immunity through the pro-inflammatory cytokines of CD8+ and CD4+(Th1 and Th17) following an intervention in the downstream signaling event of ERK1/2 (probably through p38MAPK signaling). Therefore, the study suggests a new control against visceral leishmaniasis in the future. Copyright © 2016 Elsevier Ltd. All rights reserved.

TRANSCRIPTIONAL INHIBITION OF INTERLEUKIN-12 PROMOTER ACTIVITY IN LEISHMANIA SPP.-INFECTED MACROPHAGES

PubMed Central

Jayakumar, Asha; Widenmaier, Robyn; Ma, Xiaojing; McDowell, Mary Ann

2009-01-01

To establish and persist within a host, Leishmania spp. parasites delay the onset of cell-mediated immunity by suppressing interleukin-12 (IL-12) production from host macrophages. Although it is established that Leishmania spp.-infected macrophages have impaired IL-12 production, the mechanisms that account for this suppression remain to be completely elucidated. Using a luciferase reporter assay assessing IL-12 transcription, we report here that Leishmania major, Leishmania donovani, and Leishmania chagasi inhibit IL-12 transcription in response to interferon-gamma, lipopolysaccharide, and CD40 ligand and that Leishmania spp. lipophosphoglycan, phosphoglycans, and major surface protein are not necessary for inhibition. In addition, all the Leishmania spp. strains and life-cycle stages tested inhibited IL-12 promoter activity. Our data further reveal that autocrine-acting host factors play no role in the inhibitory response and that phagocytosis signaling is necessary for inhibition of IL-12. PMID:18372625

Sand fly captures with Disney traps in area of occurrence of Leishmania (Leishmania) amazonensis in the state of Mato Grosso do Sul, mid-western Brazil.

PubMed

Dorval, Maria Elizabeth Cavalheiros; Alves, Tulia Peixoto; Cristaldo, Geucira; Rocha, Hilda Carlos da; Alves, Murilo Andrade; Oshiro, Elisa Teruya; Oliveira, Alessandra Gutierrez de; Brazil, Reginaldo Peçanha; Galati, Eunice Aparecida Bianchi; Cunha, Rivaldo Venancio da

2010-01-01

The work was conducted to study phlebotomine fauna (Diptera: Psychodidae) and aspects of American cutaneous leishmaniasis transmission in a forested area where Leishmania (Leishmania) amazonensis occurs, situated in the municipality of Bela Vista, State of Mato Grosso do Sul, Brazil. The captures were conducted with modified Disney traps, using hamster (Mesocricetus auratus) as bait, from May 2004 to January 2006. Ten species of phlebotomine sandflies were captured: Brumptomyia avellari, Brumptomyia brumpti, Bichromomyia flaviscutellata, Evandromyia bourrouli, Evandromyia lenti, Lutzomyia longipalpis, Psathyromyia campograndensis, Psathyromyia punctigeniculata, Psathyromyia shannoni and Sciopemyia sordellii. The two predominant species were Ev bourrouli (57.3%) and Bi flaviscutellata (41.4%), present at all sampling sites. Two of the 36 hamsters used as bait presented natural infection with Leishmania. The parasite was identified as Leishmania (Leishmania) amazonensis. Analysis of the results revealed the efficiency of Disney traps for capturing Bichromomyia flaviscutellata and the simultaneous presence of both vector and the Leishmania species transmitted by the same can be considered a predictive factor of the occurrence of leishmaniasis outbreaks for the human population that occupies the location.

Overview of dendritic cell-based vaccine development for leishmaniasis.

PubMed

Bagirova, M; Allahverdiyev, A M; Abamor, E S; Ullah, I; Cosar, G; Aydogdu, M; Senturk, H; Ergenoglu, B

2016-11-01

Leishmaniasis is one of the most serious vector-borne diseases in the world and is distributed over 98 countries. It is estimated that 350 million people are at risk for leishmaniasis. There are three different generation of vaccines that have been developed to provide immunity and protection against leishmaniasis. However, their use has been limited due to undesired side effects. These vaccines have also failed to provide effective and reliable protection and, as such, currently, there is no safe and effective vaccine for leishmaniasis. Dendritic cells (DCs) are a unique population of cells that come from bone marrow and become specialized to take up, process and present antigens to helper T cells in a mechanism similar to macrophages. By considering these significant features, DCs stimulated with different kinds of Leishmania antigens have been used in recent vaccine studies for leishmaniasis with promising results so far. In this review, we aim to review and combine the latest studies about this issue after defining potential problems in vaccine development for leishmaniasis and considering the importance of DCs in the immunopathogenesis of the disease. © 2016 John Wiley & Sons Ltd.

Re-designing the Mozambique vaccine supply chain to improve access to vaccines

PubMed Central

Lee, Bruce Y.; Haidari, Leila A.; Prosser, Wendy; Connor, Diana L.; Bechtel, Ruth; Dipuve, Amelia; Kassim, Hidayat; Khanlawia, Balbina; Brown, Shawn T.

2017-01-01

Introduction Populations and routine childhood vaccine regimens have changed substantially since supply chains were designed in the 1980s, and introducing new vaccines during the “Decade of Vaccine” may exacerbate existing bottlenecks, further inhibiting the flow of all vaccines. Methods Working with the Mozambique Ministry of Health, our team implemented a new process that integrated HERMES computational simulation modeling and on-the-ground implementers to evaluate and improve the Mozambique vaccine supply chain using a system-re-design that integrated new supply chain structures, information technology, equipment, personnel, and policies. Results The alternative system design raised vaccine availability (from 66% to 93% in Gaza; from 76% to 84% in Cabo Delgado) and reduced the logistics cost per dose administered (from $0.53 to $0.32 in Gaza; from $0.38 to $0.24 in Cabo Delgado) as compared to the multi-tiered system under the current EPI. The alternative system also produced higher availability at lower costs after new vaccine introductions. Since reviewing scenarios modeling deliveries every two months in the north of Gaza, the provincial directorate has decided to pilot this approach diverging from decades of policies dictating monthly deliveries. Discussion Re-design improved not only supply chain efficacy but also efficiency, important since resources to deliver vaccines are limited. The Mozambique experience and process can serve as a model for other countries during the Decade of Vaccines. For the Decade of Vaccines, getting vaccines at affordable prices to the market is not enough. Vaccines must reach the population to be successful. PMID:27576077

Comparative genomic analysis of three Leishmania species that cause diverse human disease

PubMed Central

Peacock, Christopher S; Seeger, Kathy; Harris, David; Murphy, Lee; Ruiz, Jeronimo C; Quail, Michael A; Peters, Nick; Adlem, Ellen; Tivey, Adrian; Aslett, Martin; Kerhornou, Arnaud; Ivens, Alasdair; Fraser, Audrey; Rajandream, Marie-Adele; Carver, Tim; Norbertczak, Halina; Chillingworth, Tracey; Hance, Zahra; Jagels, Kay; Moule, Sharon; Ormond, Doug; Rutter, Simon; Squares, Rob; Whitehead, Sally; Rabbinowitsch, Ester; Arrowsmith, Claire; White, Brian; Thurston, Scott; Bringaud, Frédéric; Baldauf, Sandra L; Faulconbridge, Adam; Jeffares, Daniel; Depledge, Daniel P; Oyola, Samuel O; Hilley, James D; Brito, Loislene O; Tosi, Luiz R O; Barrell, Barclay; Cruz, Angela K; Mottram, Jeremy C; Smith, Deborah F; Berriman, Matthew

2008-01-01

Leishmania parasites cause a broad spectrum of clinical disease. Here we report the sequencing of the genomes of two species of Leishmania: Leishmania infantum and Leishmania braziliensis. The comparison of these sequences with the published genome of Leishmania major reveals marked conservation of synteny and identifies only ∼200 genes with a differential distribution between the three species. L. braziliensis, contrary to Leishmania species examined so far, possesses components of a putative RNA-mediated interference pathway, telomere-associated transposable elements and spliced leader–associated SLACS retrotransposons. We show that pseudogene formation and gene loss are the principal forces shaping the different genomes. Genes that are differentially distributed between the species encode proteins implicated in host-pathogen interactions and parasite survival in the macrophage. PMID:17572675

Phylogenetic position of Leishmania isolates from Khyber Pakhtunkhwa province of Pakistan.

PubMed

Khan, Nazma Habib; Messenger, Louisa A; Wahid, Sobia; Sutherland, Colin J

2016-08-01

Several species of the genus Leishmania are causative agents of cutaneous leishmaniasis in Pakistan. This study aimed to determine phylogenetic placement of Leishmania species causing cutaneous leishmaniasis in Khyber Pakhtunkhwa province, Pakistan (34 Leishmania tropica, 3 Leishmania infantum), in-relation to species from other geographical areas using gene sequences encoding cytochrome b (cytb) and internal transcribed spacer 2 (its2). Based on cytochrome b sequence analysis, L. tropica strains from Pakistan and other geographical regions were differentiated into two genotype groups, A and B. Within the province, five distinct L. tropica genotypes were recognized; two in group A, three in group B. Two L. infantum isolates from the province were closely associated with both Afro-Eurasian and American species of the Leishmania donovani complex, including Leishmania chagasi, L. infantum and L. donovani from Sudan and Ethiopia; while a third L. infantum isolate could not be differentiated from visceralizing Kenyan and Indian L. donovani. We observed apposite phylogenetic placement of CL-causing L. tropica and L. infantum from Khyber Pakhtunkhwa. Affinities ascribed to Leishmania spp. From the region are valuable in tracing potential importation of leishmaniasis. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

Vaccination of dogs with six different candidate leishmaniasis vaccines composed of a chimerical recombinant protein containing ribosomal and histone protein epitopes in combination with different adjuvants.

PubMed

Poot, J; Janssen, L H M; van Kasteren-Westerneng, T J; van der Heijden-Liefkens, K H A; Schijns, V E J C; Heckeroth, A

2009-07-16

Chimerical protein "Q", composed of antigenic ribosomal and histone sequences, in combination with live BCG is a promising canine leishmaniasis vaccine candidate; one of the few vaccine candidates that have been tested successfully in dogs. Unfortunately, live BCG is not an appropriate adjuvant for commercial application due to safety problems in dogs. In order to find a safe adjuvant with similar efficacy to live BCG, muramyl dipeptide, aluminium hydroxide, Matrix C and killed Propionibacterium acnes in combination with either E. coli- or baculovirus-produced recombinant JPCM5_Q protein were tested. Groups of five or seven dogs were vaccinated with six different adjuvant-antigen combinations and challenged with a high dose intravenous injection of Leishmania infantum JPC strain promastigotes. All candidate vaccines proved to be safe, and both humoral and cellular responses to the recombinant proteins were detected at the end of the prime-boost vaccination scheme. However, clinical and parasitological data obtained during the 10 month follow-up period indicated that protection was not induced by either of the six candidate vaccines. Although no direct evidence was obtained, our data suggest that live BCG may have a significant protective effect against challenge with L. infantum in dogs.

Molecular Identification of Leishmania spp. in Sand Flies (Diptera: Psychodidae, Phlebotominae) From Ecuador.

PubMed

Quiroga, Cristina; Cevallos, Varsovia; Morales, Diego; Baldeón, Manuel E; Cárdenas, Paúl; Rojas-Silva, Patricio; Ponce, Patricio

2017-11-07

The detection and identification of natural infections in sand flies by Leishmania protozoan species in endemic areas is a key factor in assessing the risk of leishmaniasis and in designing prevention and control measures for this infectious disease. In this study, we analyzed the Leishmania DNA using nuclear ribosomal internal transcript spacer (ITS) sequences. Parasite DNA was extracted from naturally infected, blood-fed sand flies collected in nine localities considered leishmaniasis-endemic foci in Ecuador.The species of parasites identified in sand flies were Leishmania major-like, Leishmania naiffi, Leishmania mexicana, Leishmania lainsoni, and "Leishmania sp. siamensis". Sand fly specimens of Brumptomyia leopoldoi, Mycropigomyia cayennensis, Nyssomyia yuilli yuilli, Nyssomyia trapidoi, Pressatia triacantha, Pressatia dysponeta, Psychodopygus carrerai carrerai, Psychodopygus panamensis, and Trichophoromyia ubiquitalis were found positive for Leishmania parasite. These findings contribute to a better understanding of the epidemiology and transmission dynamics of the disease in high-risk areas of Ecuador. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America.

Molecular Identification of Leishmania spp. in Sand Flies (Diptera: Psychodidae, Phlebotominae) From Ecuador

PubMed Central

Cevallos, Varsovia; Morales, Diego; Baldeón, Manuel E; Cárdenas, Paúl; Rojas-Silva, Patricio; Ponce, Patricio

2017-01-01

Abstract The detection and identification of natural infections in sand flies by Leishmania protozoan species in endemic areas is a key factor in assessing the risk of leishmaniasis and in designing prevention and control measures for this infectious disease. In this study, we analyzed the Leishmania DNA using nuclear ribosomal internal transcript spacer (ITS) sequences. Parasite DNA was extracted from naturally infected, blood-fed sand flies collected in nine localities considered leishmaniasis-endemic foci in Ecuador. The species of parasites identified in sand flies were Leishmania major-like, Leishmania naiffi, Leishmania mexicana, Leishmania lainsoni, and “Leishmania sp. siamensis”. Sand fly specimens of Brumptomyia leopoldoi, Mycropigomyia cayennensis, Nyssomyia yuilli yuilli, Nyssomyia trapidoi, Pressatia triacantha, Pressatia dysponeta, Psychodopygus carrerai carrerai, Psychodopygus panamensis, and Trichophoromyia ubiquitalis were found positive for Leishmania parasite. These findings contribute to a better understanding of the epidemiology and transmission dynamics of the disease in high-risk areas of Ecuador. PMID:28981860

Monocyte-Derived Signals Activate Human Natural Killer Cells in Response to Leishmania Parasites

PubMed Central

Messlinger, Helena; Sebald, Heidi; Heger, Lukas; Dudziak, Diana; Bogdan, Christian; Schleicher, Ulrike

2018-01-01

Activated natural killer (NK) cells release interferon (IFN)-γ, which is crucial for the control of intracellular pathogens such as Leishmania. In contrast to experimental murine leishmaniasis, the human NK cell response to Leishmania is still poorly characterized. Here, we investigated the interaction of human blood NK cells with promastigotes of different Leishmania species (Leishmania major, Leishmania mexicana, Leishmania infantum, and Leishmania donovani). When peripheral blood mononuclear cells or purified NK cells and monocytes (all derived from healthy blood donors from Germany without a history of leishmaniasis) were exposed to promastigotes, NK cells showed increased surface expression of the activation marker CD69. The extent of this effect varied depending on the Leishmania species; differences between dermotropic and viscerotropic L. infantum strains were not observed. Upregulation of CD69 required direct contact between monocytes and Leishmania and was partly inhibitable by anti-interleukin (IL)-18. Unexpectedly, IL-18 was undetectable in most of the supernatants (SNs) of monocyte/parasite cocultures. Confocal fluorescence microscopy of non-permeabilized cells revealed that Leishmania-infected monocytes trans-presented IL-18 to NK cells. Native, but not heat-treated SNs of monocyte/Leishmania cocultures also induced CD69 on NK cells, indicating the involvement of a soluble heat-labile factor other than IL-18. A role for the NK cell-activating cytokines IL-1β, IL-2, IL-12, IL-15, IL-21, and IFN-α/β was excluded. The increase of CD69 was not paralleled by NK cell IFN-γ production or enhanced cytotoxicity. However, prior exposure of NK cells to Leishmania parasites synergistically increased their IFN-γ release in response to IL-12, which was dependent on endogenous IL-18. CD1c+ dendritic cells were identified as possible source of Leishmania-induced IL-12. Finally, we observed that direct contact between Leishmania and NK cells reduced the

Natural infection of Ctenodactylus gundi by Leishmania major in Tunisia.

PubMed

Ghawar, Wissem; Bettaieb, Jihène; Salem, Sadok; Snoussi, Mohammed-Ali; Jaouadi, Kaouther; Yazidi, Rihab; Ben-Salah, Afif

2018-01-01

Incriminating new rodent species, as reservoir hosts of Leishmania parasites is crucial for understanding the transmission cycle of cutaneous leishmaniasis in Tunisia. Ctenodactylus (C.) gundi was previously described as extremely abundant in all Tunisian Leishmania (L.) tropica foci in south Tunisia besides its presence in L. major endemic area. The aim of this study was to detect Leishmania species parasites among C. gundi in two endemic regions in Tunisia: Sidi Bouzid and Tataouine. Total DNA was isolated from the spleens and the livers of 92C. gundi. Leishmaniasis clinical manifestations were detected among 11 rodents (12%). Leishmania parasites were detected in 30 (32.6%) rodents using direct exam method. Leishmania DNA was detected in 40 (43.5%) C. gundi by combining results among spleens and livers using ITS1-PCR. Positive samples were confirmed to be L. major except for only one specimen which was L. tropica. These results demonstrated, for the first time, the high natural infection rate of C. gundi with L. major parasites in Tunisia. Hence, C. gundi should be considered as potential reservoir host of Leishmania parasites causing cutaneous leishmaniasis in Tunisia. Copyright © 2017. Published by Elsevier B.V.

Leishmania and the macrophage: a multifaceted interaction.

PubMed

Podinovskaia, Maria; Descoteaux, Albert

2015-01-01

Leishmania, the causative agent of leishmaniases, is an intracellular parasite of macrophages, transmitted to humans via the bite of its sand fly vector. This protozoan organism has evolved strategies for efficient uptake into macrophages and is able to regulate phagosome maturation in order to make the phagosome more hospitable for parasite growth and to avoid destruction. As a result, macrophage defenses such as oxidative damage, antigen presentation, immune activation and apoptosis are compromised whereas nutrient availability is improved. Many Leishmania survival factors are involved in shaping the phagosome and reprogramming the macrophage to promote infection. This review details the complexity of the host-parasite interactions and summarizes our latest understanding of key events that make Leishmania such a successful intracellular parasite.

Analysis of kinetoplast cytochrome b gene of 16 Leishmania isolates from different foci of China: different species of Leishmania in China and their phylogenetic inference

PubMed Central

2013-01-01

Background Leishmania species belong to the family Trypanosomatidae and cause leishmaniasis, a geographically widespread disease that infects humans and other vertebrates. This disease remains endemic in China. Due to the large geographic area and complex ecological environment, the taxonomic position and phylogenetic relationship of Chinese Leishmania isolates remain uncertain. A recent internal transcribed spacer 1 and cytochrome oxidase II phylogeny of Chinese Leishmania isolates has challenged some aspects of their traditional taxonomy as well as cladistics hypotheses of their phylogeny. The current study was designed to provide further disease background and sequence analysis. Methods We systematically analyzed 50 cytochrome b (cyt b) gene sequences of 19 isolates (16 from China, 3 from other countries) sequenced after polymerase chain reaction (PCR) using a special primer for cyt b as well as 31 sequences downloaded from GenBank. After alignment, the data were analyzed using the maximum parsimony, Bayesian and netwok methods. Results Sequences of six haplotypes representing 10 Chinese isolates formed a monophyletic group and clustered with Leishmania tarentolae. The isolates GS1, GS7, XJ771 of this study from China clustered with other isolates of Leishmania donovani complex. The isolate JS1 was a sister to Leishmania tropica, which represented an L. tropica complex instead of clustering with L. donovani complex or with the other 10 Chinese isolates. The isolates KXG-2 and GS-GER20 formed a monophyletic group with Leishmania turanica from central Asia. In the different phylogenetic trees, all of the Chinese isolates occurred in at least four groups regardless of geographic distribution. Conclusions The undescribed Leishmania species of China, which are clearly causative agents of canine leishmaniasis and human visceral leishmaniasis and are related to Sauroleishmania, may have evolved from a common ancestral parasite that came from the Americas and may have

Differentiation of Leishmania (Viannia) panamensis and Leishmania (V.) guyanensis using BccI for hsp70 PCR-RFLP.

PubMed

Montalvo Alvarez, Ana Margarita; Nodarse, Jorge Fraga; Goodridge, Ivón Montano; Fidalgo, Lianet Monzote; Marin, Marcel; Van Der Auwera, Gert; Dujardin, Jean-Claude; Bernal, Iván Darío Velez; Muskus, Carlos

2010-05-01

Leishmania panamensis and Leishmania guyanensis are two species of the subgenus Viannia that are genetically very similar. Both parasites are usually associated with cutaneous leishmaniasis, but also have the potential to cause the mucocutaneous form of the disease. In addition, the study of foci and consequently the identification of vectors and probable reservoirs involved in transmission require a correct differentiation between both species, which is important at epidemiological level. We explored the possibility of identifying these species by using restriction fragment length polymorphisms (RFLP) in the gene coding for heat-shock protein 70 (hsp70). Previously, an hsp70 PCR-RFLP assay proved to be very effective in differentiating other Leishmania species when HaeIII is used as restriction enzyme. Based on hsp70 sequences analysis, BccI was found to generate species-specific fragments that can easily be recognized by agarose gel electrophoresis. Using the analysis of biopsies, scrapings, and parasite isolates previously grouped in a cluster comprising both L. panamensis and L. guyanensis, we showed that our approach allowed differentiation of both entities. This offers the possibility not only for identification of parasites in biological samples, but also to apply molecular epidemiology in certain countries of the New World, where several Leishmania species could coexist. Copyright 2009 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved.

Distinct Leishmania Species Infecting Wild Caviomorph Rodents (Rodentia: Hystricognathi) from Brazil

PubMed Central

Cássia-Pires, Renata; Boité, Mariana C.; D'Andrea, Paulo S.; Herrera, Heitor M.; Cupolillo, Elisa; Jansen, Ana Maria; Roque, André Luiz R.

2014-01-01

Background Caviomorph rodents, some of the oldest Leishmania spp. hosts, are widely dispersed in Brazil. Despite both experimental and field studies having suggested that these rodents are potential reservoirs of Leishmania parasites, not more than 88 specimens were analyzed in the few studies of natural infection. Our hypothesis was that caviomorph rodents are inserted in the transmission cycles of Leishmania in different regions, more so than is currently recognized. Methodology We investigated the Leishmania infection in spleen fragments of 373 caviomorph rodents from 20 different species collected in five Brazilian biomes in a period of 13 years. PCR reactions targeting kDNA of Leishmania sp. were used to diagnose infection, while Leishmania species identification was performed by DNA sequencing of the amplified products obtained in the HSP70 (234) targeting. Serology by IFAT was performed on the available serum of these rodents. Principal findings In 13 caviomorph rodents, DNA sequencing analyses allowed the identification of 4 species of the subgenus L. (Viannia): L. shawi, L. guyanensis, L. naiffi, and L. braziliensis; and 1 species of the subgenus L. (Leishmania): L. infantum. These include the description of parasite species in areas not previously included in their known distribution: L. shawi in Thrichomys inermis from Northeastern Brazil and L. naiffi in T. fosteri from Western Brazil. From the four other positive rodents, two were positive for HSP70 (234) targeting but did not generate sequences that enabled the species identification, and another two were positive only in kDNA targeting. Conclusions/Significance The infection rate demonstrated by the serology (51.3%) points out that the natural Leishmania infection in caviomorph rodents is much higher than that observed in the molecular diagnosis (4.6%), highlighting that, in terms of the host species responsible for maintaining Leishmania species in the wild, our current knowledge represents only the

Evaluation of the immunoprophylactic potential of a killed vaccine candidate in combination with different adjuvants against murine visceral leishmaniasis.

PubMed

Thakur, Ankita; Kaur, Harpreet; Kaur, Sukhbir

2015-02-01

Despite a large number of field trials, till date no prophylactic antileishmanial vaccine exists for human use. Killed antigen formulations offer the advantage of being safe but they have limited immunogenicity. Recent research has documented that efforts to develop effective Leishmania vaccine have been limited due to the lack of an appropriate adjuvant. Addition of adjuvants to vaccines boosts and directs the immunogenicity of antigens. So, the present study was done to evaluate the effectiveness of four adjuvants i.e. alum, saponin, cationic liposomes and monophosphoryl lipid-A in combination with Autoclaved Leishmania donovani (ALD) antigen against murine visceral leishmaniasis (VL). BALB/c mice were immunized thrice with respective vaccine formulation. Two weeks after last booster, challenge infection was given. Mice were sacrificed 15 days after last immunization and on 30, 60 and 90 post infection/challenge days. A considerable protective efficacy was shown by all vaccine formulations. It was evident from significant reduction in parasite load, profound delayed type hypersensitivity responses (DTH), increased IgG2a titres and high levels of Th1 cytokines (IFN-γ, IL-12) as compared to the infected controls. However, level of protection varied with the type of adjuvant used. Maximum protection was achieved with the use of liposome encapsulated ALD antigen and it was closely followed by group immunized with ALD+MPL-A. Significant results were also obtained with ALD+saponin, ALD+alum and ALD antigen (alone) but the protective efficacy was reduced as compared to other immunized groups. The present study reveals greater efficacy of two vaccine formulations i.e. ALD+liposome and ALD+MPL-A against murine VL. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Mechanism of interaction of sitamaquine with Leishmania donovani.

PubMed

Coimbra, E S; Libong, D; Cojean, S; Saint-Pierre-Chazalet, M; Solgadi, A; Le Moyec, L; Duenas-Romero, A M; Chaminade, P; Loiseau, P M

2010-12-01

This study focuses on the mechanism of interaction of sitamaquine with Leishmania donovani membranes, and its accumulation within the parasites. A biomimetic model of the outer layer of a Leishmania plasma membrane was used to examine the interactions of sitamaquine with lipids. The plasma membranes of L. donovani promastigotes were depleted of sterol using cholesterol oxidase, in order to assess the importance of sterols in drug-membrane interactions. Sterols were quantified and sitamaquine susceptibility was assessed using the MTT test. Kinetics of sitamaquine accumulation and efflux were measured under different conditions. Sitamaquine interacts first with phospholipid anionic polar head groups and then with phospholipid acyl chains to insert within biological membranes and accumulates rapidly in the Leishmania cytosol according to a sterol-independent process. The rapid sitamaquine efflux observed was related to an energy-dependent mechanism since the intracellular amount of sitamaquine was enhanced three times in the absence of glucose and the efflux was inhibited in energy-depleted conditions. (1)H NMR analysis of motile lipid showed that sitamaquine did not affect lipid trafficking in Leishmania. We propose that sitamaquine rapidly accumulates in Leishmania by diffusion along an electrical gradient and is concentrated in the cytosol by an energy- and sterol-independent process. The affinity of sitamaquine for membranes was transitory and an energy-dependent efflux was demonstrated, suggesting the presence of an as yet uncharacterized transporter.

Vaccines for pandemic influenza. The history of our current vaccines, their limitations and the requirements to deal with a pandemic threat.

PubMed

Hampson, Alan W

2008-06-01

Fears of a potential pandemic due to A(H5N1) viruses have focussed new attention on our current vaccines, their shortcomings, and concerns regarding global vaccine supply in a pandemic. The bulk of current vaccines are inactivated split virus vaccines produced from egg-grown virus and have only modest improvements compared with those first introduced over 60 years ago. Splitting, which was introduced some years ago to reduce reactogenicity, also reduces the immunogenicity of vaccines in immunologically naïve recipients. The A(H5N1) viruses have been found poorly immunogenic and present other challenges for vaccine producers which further exacerbate an already limited global production capacity. There have been some recent improvements in vaccine production methods and improvements to immunogenicity by the development of new adjuvants, however, these still fall short of providing timely supplies of vaccine for all in the face of a pandemic. New approaches to influenza vaccines which might fulfil the demands of a pandemic situation are under evaluation, however, these remain some distance from clinical reality and face significant regulatory hurdles.

Adverse events of vaccines and the consequences of non-vaccination: a critical review

PubMed Central

Aps, Luana Raposo de Melo Moraes; Piantola, Marco Aurélio Floriano; Pereira, Sara Araujo; de Castro, Julia Tavares; Santos, Fernanda Ayane de Oliveira; Ferreira, Luís Carlos de Souza

2018-01-01

ABSTRACT OBJECTIVE: To analyze the risks related to vaccines and the impacts of non-vaccination on the world population. METHODS: This is a narrative review that has considered information present in the bibliographic databases NCBI-PubMed, Medline, Lilacs, and Scientific Electronic Library Online (SciELO), between November 2015 and November 2016. For the analysis of outbreaks caused by non-vaccination, we considered the work published between 2010 and 2016. RESULTS: We have described the main components of the vaccines offered by the Brazilian public health system and the adverse events associated with these elements. Except for local inflammatory reactions and rare events, such as exacerbation of autoimmune diseases and allergies, no causal relationship has been demonstrated between the administration of vaccines and autism, Alzheimer's disease, or narcolepsy. On the other hand, the lack of information and the dissemination of non-scientific information have contributed to the reemergence of infectious diseases in several countries in the world and they jeopardize global plans for the eradication of these diseases. CONCLUSIONS: The population should be well informed about the benefits of vaccination and health professionals should assume the role of disseminating truthful information with scientific support on the subject, as an ethical and professional commitment to society. PMID:29668817

DDX3 DEAD-box RNA helicase plays a central role in mitochondrial protein quality control in Leishmania

PubMed Central

Padmanabhan, Prasad Kottayil; Zghidi-Abouzid, Ouafa; Samant, Mukesh; Dumas, Carole; Aguiar, Bruno Guedes; Estaquier, Jerome; Papadopoulou, Barbara

2016-01-01

DDX3 is a highly conserved member of ATP-dependent DEAD-box RNA helicases with multiple functions in RNA metabolism and cellular signaling. Here, we describe a novel function for DDX3 in regulating the mitochondrial stress response in the parasitic protozoan Leishmania. We show that genetic inactivation of DDX3 leads to the accumulation of mitochondrial reactive oxygen species (ROS) associated with a defect in hydrogen peroxide detoxification. Upon stress, ROS production is greatly enhanced, causing mitochondrial membrane potential loss, mitochondrial fragmentation, and cell death. Importantly, this phenotype is exacerbated upon oxidative stress in parasites forced to use the mitochondrial oxidative respiratory machinery. Furthermore, we show that in the absence of DDX3, levels of major components of the unfolded protein response as well as of polyubiquitinated proteins increase in the parasite, particularly in the mitochondrion, as an indicator of mitochondrial protein damage. Consistent with these findings, immunoprecipitation and mass-spectrometry studies revealed potential interactions of DDX3 with key components of the cellular stress response, particularly the antioxidant response, the unfolded protein response, and the AAA-ATPase p97/VCP/Cdc48, which is essential in mitochondrial protein quality control by driving proteosomal degradation of polyubiquitinated proteins. Complementation studies using DDX3 deletion mutants lacking conserved motifs within the helicase core support that binding of DDX3 to ATP is essential for DDX3's function in mitochondrial proteostasis. As a result of the inability of DDX3-depleted Leishmania to recover from ROS damage and to survive various stresses in the host macrophage, parasite intracellular development was impaired. Collectively, these observations support a central role for the Leishmania DDX3 homolog in preventing ROS-mediated damage and in maintaining mitochondrial protein quality control. PMID:27735940

The use of kDNA minicircle subclass relative abundance to differentiate between Leishmania (L.) infantum and Leishmania (L.) amazonensis.

PubMed

Ceccarelli, Marcello; Galluzzi, Luca; Diotallevi, Aurora; Andreoni, Francesca; Fowler, Hailie; Petersen, Christine; Vitale, Fabrizio; Magnani, Mauro

2017-05-16

Leishmaniasis is a neglected disease caused by many Leishmania species, belonging to subgenera Leishmania (Leishmania) and Leishmania (Viannia). Several qPCR-based molecular diagnostic approaches have been reported for detection and quantification of Leishmania species. Many of these approaches use the kinetoplast DNA (kDNA) minicircles as the target sequence. These assays had potential cross-species amplification, due to sequence similarity between Leishmania species. Previous works demonstrated discrimination between L. (Leishmania) and L. (Viannia) by SYBR green-based qPCR assays designed on kDNA, followed by melting or high-resolution melt (HRM) analysis. Importantly, these approaches cannot fully distinguish L. (L.) infantum from L. (L.) amazonensis, which can coexist in the same geographical area. DNA from 18 strains/isolates of L. (L.) infantum, L. (L.) amazonensis, L. (V.) braziliensis, L. (V.) panamensis, L. (V.) guyanensis, and 62 clinical samples from L. (L.) infantum-infected dogs were amplified by a previously developed qPCR (qPCR-ML) and subjected to HRM analysis; selected PCR products were sequenced using an ABI PRISM 310 Genetic Analyzer. Based on the obtained sequences, a new SYBR-green qPCR assay (qPCR-ama) intended to amplify a minicircle subclass more abundant in L. (L.) amazonensis was designed. The qPCR-ML followed by HRM analysis did not allow discrimination between L. (L.) amazonensis and L. (L.) infantum in 53.4% of cases. Hence, the novel SYBR green-based qPCR (qPCR-ama) has been tested. This assay achieved a detection limit of 0.1 pg of parasite DNA in samples spiked with host DNA and did not show cross amplification with Trypanosoma cruzi or host DNA. Although the qPCR-ama also amplified L. (L.) infantum strains, the C q values were dramatically increased compared to qPCR-ML. Therefore, the combined analysis of C q values from qPCR-ML and qPCR-ama allowed to distinguish L. (L.) infantum and L. (L.) amazonensis in 100% of tested samples

Immunization with LJM11 salivary protein protects against infection with Leishmania braziliensis in the presence of Lutzomyia longipalpis saliva.

PubMed

Cunha, Jurema M; Abbehusen, Melissa; Suarez, Martha; Valenzuela, Jesus; Teixeira, Clarissa R; Brodskyn, Cláudia I

2018-01-01

Leishmania is transmitted in the presence of sand fly saliva. Protective immunity generated by saliva has encouraged identification of a vector salivary-based vaccine. Previous studies have shown that immunization with LJM11, a salivary protein from Lutzomyia longipalpis, is able to induce a Th1 immune response and protect mice against bites of Leishmania major-infected Lutzomyia longipalpis. Here, we further investigate if immunization with LJM11 recombinant protein is able to confer cross-protection against infection with Leishmania braziliensis associated with salivary gland sonicate (SGS) from Lutzomyia intermedia or Lu. longipalpis. Mice immunized with LJM11 protein exhibited an increased production of anti-LJM11 IgG, IgG1 and IgG2a and a DTH response characterized by an inflammatory infiltrate with the presence of CD4 + IFN-γ + T cells. LJM11-immunized mice were intradermally infected in the ear with L. braziliensis in the presence of Lu. longipalpis or Lu. intermedia SGS. A significant reduction of parasite numbers in the ear and lymph node in the group challenged with L. braziliensis plus Lu. longipalpis SGS was observed, but not when the challenge was performed with L. braziliensis plus Lu. intermedia SGS. A higher specific production of IFN-γ and absence of IL-10 by lymph node cells were only observed in LJM11 immunized mice after infection. After two weeks, a similar frequency of CD4 + IFN-γ + T cells was detected in LJM11 and BSA groups challenged with L. braziliensis plus Lu. longipalpis SGS, suggesting that early events possibly triggered by immunization are essential for protection against Leishmania infection. Our findings support the specificity of saliva-mediated immune responses and reinforce the importance of identifying cross-protective salivary antigens. Copyright © 2017 Elsevier B.V. All rights reserved.

First molecular detection of Leishmania tarentolae-like DNA in Sergentomyia minuta in Spain.

PubMed

Bravo-Barriga, Daniel; Parreira, Ricardo; Maia, Carla; Blanco-Ciudad, Juan; Afonso, Maria Odete; Frontera, Eva; Campino, Lenea; Pérez-Martín, Juan Enrique; Serrano Aguilera, Francisco Javier; Reina, David

2016-03-01

Phlebotomine sand flies (Diptera, Psychodidae) are vectors of multiple Leishmania species, among which Leishmania infantum stands out as a being frequently pathogenic to humans and dogs in Mediterranean countries. In this study, Sergentomyia minuta sand flies were collected using CDC miniature light traps in different 431 biotopes from Southwest Spain. A total of 114 females were tested for the presence of Leishmania DNA by targeting ITS-1 and cyt-B sequences by PCR. Leishmania DNA was detected in one S. minuta. Characterization of the obtained DNA sequences by phylogenetic analyses revealed close relatedness with Leishmania tarentolae Wenyon, 1921 as well as with both human and canine pathogenic strains of Asian origin (China), previously described as Leishmania sp. To our knowledge, this is the first report of phlebotomine sand flies naturally infected with L. tarentolae-like in Spain. The possible infection of sand flies with novel Leishmania species should be taken into consideration in epidemiological studies of vector species in areas where leishmaniosis is endemic.

Th1 stimulatory proteins of Leishmania donovani: comparative cellular and protective responses of rTriose phosphate isomerase, rProtein disulfide isomerase and rElongation factor-2 in combination with rHSP70 against visceral leishmaniasis.

PubMed

Jaiswal, Anil Kumar; Khare, Prashant; Joshi, Sumit; Kushawaha, Pramod Kumar; Sundar, Shyam; Dube, Anuradha

2014-01-01

In visceral leishmaniasis, the recovery from the disease is always associated with the generation of Th1-type of cellular responses. Based on this, we have previously identified several Th1-stimulatory proteins of Leishmania donovani -triose phosphate isomerase (TPI), protein disulfide isomerase (PDI) and elongation factor-2 (EL-2) etc. including heat shock protein 70 (HSP70) which induced Th1-type of cellular responses in both cured Leishmania patients/hamsters. Since, HSPs, being the logical targets for vaccines aimed at augmenting cellular immunity and can be early targets in the immune response against intracellular pathogens; they could be exploited as vaccine/adjuvant to induce long-term immunity more effectively. Therefore, in this study, we checked whether HSP70 can further enhance the immunogenicity and protective responses of the above said Th1-stimulatory proteins. Since, in most of the studies, immunogenicity of HSP70 of L. donovani was assessed in native condition, herein we generated recombinant HSP70 and tested its potential to stimulate immune responses in lymphocytes of cured Leishmania infected hamsters as well as in the peripheral blood mononuclear cells (PBMCs) of cured patients of VL either individually or in combination with above mentioned recombinant proteins. rLdHSP70 alone elicited strong cellular responses along with remarkable up-regulation of IFN-γ and IL-12 cytokines and extremely lower level of IL-4 and IL-10. Among the various combinations, rLdHSP70 + rLdPDI emerged as superior one augmenting improved cellular responses followed by rLdHSP70 + rLdEL-2. These combinations were further evaluated for its protective potential wherein rLdHSP70 + rLdPDI again conferred utmost protection (∼80%) followed by rLdHSP70 + rLdEL-2 (∼75%) and generated a strong cellular immune response with significant increase in the levels of iNOS transcript as well as IFN-γ and IL-12 cytokines which was further supported by the high level of IgG2 antibody

Th1 Stimulatory Proteins of Leishmania donovani: Comparative Cellular and Protective Responses of rTriose Phosphate Isomerase, rProtein Disulfide Isomerase and rElongation Factor-2 in Combination with rHSP70 against Visceral Leishmaniasis

PubMed Central

Jaiswal, Anil Kumar; Khare, Prashant; Joshi, Sumit; Kushawaha, Pramod Kumar; Sundar, Shyam; Dube, Anuradha

2014-01-01

In visceral leishmaniasis, the recovery from the disease is always associated with the generation of Th1-type of cellular responses. Based on this, we have previously identified several Th1-stimulatory proteins of Leishmania donovani -triose phosphate isomerase (TPI), protein disulfide isomerase (PDI) and elongation factor-2 (EL-2) etc. including heat shock protein 70 (HSP70) which induced Th1-type of cellular responses in both cured Leishmania patients/hamsters. Since, HSPs, being the logical targets for vaccines aimed at augmenting cellular immunity and can be early targets in the immune response against intracellular pathogens; they could be exploited as vaccine/adjuvant to induce long-term immunity more effectively. Therefore, in this study, we checked whether HSP70 can further enhance the immunogenicity and protective responses of the above said Th1-stimulatory proteins. Since, in most of the studies, immunogenicity of HSP70 of L. donovani was assessed in native condition, herein we generated recombinant HSP70 and tested its potential to stimulate immune responses in lymphocytes of cured Leishmania infected hamsters as well as in the peripheral blood mononuclear cells (PBMCs) of cured patients of VL either individually or in combination with above mentioned recombinant proteins. rLdHSP70 alone elicited strong cellular responses along with remarkable up-regulation of IFN-γ and IL-12 cytokines and extremely lower level of IL-4 and IL-10. Among the various combinations, rLdHSP70 + rLdPDI emerged as superior one augmenting improved cellular responses followed by rLdHSP70 + rLdEL-2. These combinations were further evaluated for its protective potential wherein rLdHSP70 + rLdPDI again conferred utmost protection (∼80%) followed by rLdHSP70 + rLdEL-2 (∼75%) and generated a strong cellular immune response with significant increase in the levels of iNOS transcript as well as IFN-γ and IL-12 cytokines which was further supported by the high level of IgG2 antibody

Performance of commercially available serological diagnostic tests to detect Leishmania infantum infection on experimentally infected dogs.

PubMed

Rodríguez-Cortés, Alhelí; Ojeda, Ana; Todolí, Felicitat; Alberola, Jordi

2013-01-31

Leishmania infantum (syn. Leishmania chagasi) is the etiological agent of a widespread serious zoonotic disease that affects both humans and dogs. Prevalence and incidence of the canine infection are important parameters to determine the risk and the ways to control this reemergent zoonosis. Unfortunately, there is not a gold standard test for Leishmania infection. Our aim was to assess the operative validity of commercial tests used to detect antibodies to Leishmania in serum samples from experimental infections. Three ELISA tests (LEISCAN(®) Leishmania ELISA Test, INGEZIM(®) LEISHMANIA, and INGEZIM(®) LEISHMANIA VET), three immunochromatographic tests (INGEZIM(®) LEISHMACROM, SNAP(®) Leishmania, and WITNESS(®) Leishmania), and one IFAT were evaluated. LEISCAN(®) Leishmania ELISA test achieved the highest sensitivity and accuracy (both 0.98). Specificity was 1 for all tests except for IFAT. All tests but IFAT obtained a positive predictive value of 1, while the maximum negative predictive value was achieved by LEISCAN(®) Leishmania ELISA Test (0.93). The best positive likelihood ratio was obtained by INGEZIM(®) LEISHMANIA VET (30.26), while the best negative likelihood ratio was obtained by LEISCAN(®) Leishmania ELISA Test (0.02). The highest diagnostic odds ratio was achieved by LEISCAN(®) Leishmania ELISA Test (729.00). The largest area under the ROC curve was obtained by LEISCAN(®) Leishmania ELISA Test (0.981). Quantitative ELISA based tests performmed better than qualitative tests ("Rapid Tests"), and the test best suited to detect Leishmania in infected dogs and to provide clinically useful information was LEISCAN(®) Leishmania ELISA Test. This and other results point also to the need of revising the status of IFAT as a gold standard for the diagnosis of leishmaniasis. Copyright © 2012 Elsevier B.V. All rights reserved.

A large-scale field randomized trial demonstrates safety and efficacy of the vaccine LetiFend® against canine leishmaniosis.

PubMed

Fernández Cotrina, Javier; Iniesta, Virginia; Monroy, Isabel; Baz, Victoria; Hugnet, Christophe; Marañon, Francisco; Fabra, Mercedes; Gómez-Nieto, Luis Carlos; Alonso, Carlos

2018-04-05

Canine leishmaniosis is a zoonotic disease caused by Leishmania infantum. Extensive research is currently ongoing to develop safe and effective vaccines to protect from disease development. The European Commission has granted a marketing authorization for LetiFend®, a new vaccine containing recombinant Protein Q. The efficacy of LetiFend® vaccination in a large-scale dog population of both sexes, different breeds and ages in endemic areas is reported in this multicenter, randomized, double-blind, placebo-controlled field trial. Dogs (n = 549) living in France and Spain were randomly selected to receive a single subcutaneous dose of LetiFend® or placebo per year, and were naturally exposed to two L. infantum transmission seasons. Clinical examinations, blood and lymphoid organ sampling to evaluate serological, parasitological and disease status of the dogs were performed at different time points during the study. LetiFend® was very well tolerated and clearly reduced the incidence of clinical signs related to leishmaniosis. The number of confirmed cases of leishmaniosis was statistically significantly lower in the vaccine group. The number of dogs with parasites was close to be significantly reduced in the vaccine group (p = 0.0564). Re-vaccination of seropositive dogs demonstrated to be safe and not to worsen the course of the disease. The likelihood that a dog vaccinated with LetiFend® develops a confirmed case or clinical signs of leishmaniosis in areas with high pressure is, respectively, 5 and 9.8 time less than that for an unvaccinated dog. Thus, the overall efficacy of the LetiFend® vaccine in the prevention of confirmed cases of leishmaniosis in endemic areas with high disease pressure was shown to be 72%. In conclusion, this field trial demonstrates that LetiFend® is a novel, safe and effective vaccine for the active immunization of non-infected dogs from 6 months of age in reducing the risk of developing clinical leishmaniosis after natural

Concentration of 2'C-methyladenosine triphosphate by Leishmania guyanensis enables specific inhibition of Leishmania RNA virus 1 via its RNA polymerase.

PubMed

Robinson, John I; Beverley, Stephen M

2018-04-27

Leishmania is a widespread trypanosomatid protozoan parasite causing significant morbidity and mortality in humans. The endobiont dsRNA virus Leishmania RNA virus 1 (LRV1) chronically infects some strains, where it increases parasite numbers and virulence in murine leishmaniasis models, and correlates with increased treatment failure in human disease. Previously, we reported that 2'-C-methyladenosine (2CMA) potently inhibited LRV1 in Leishmania guyanensis ( Lgy ) and Leishmania braziliensis , leading to viral eradication at concentrations above 10 μm Here we probed the cellular mechanisms of 2CMA inhibition, involving metabolism, accumulation, and inhibition of the viral RNA-dependent RNA polymerase (RDRP). Activation to 2CMA triphosphate (2CMA-TP) was required, as 2CMA showed no inhibition of RDRP activity from virions purified on cesium chloride gradients. In contrast, 2CMA-TP showed IC 50 values ranging from 150 to 910 μm, depending on the CsCl density of the virion (empty, ssRNA-, and dsRNA-containing). Lgy parasites incubated in vitro with 10 μm 2CMA accumulated 2CMA-TP to 410 μm, greater than the most sensitive RDRP IC 50 measured. Quantitative modeling showed good agreement between the degree of LRV1 RDRP inhibition and LRV1 levels. These results establish that 2CMA activity is due to its conversion to 2CMA-TP, which accumulates to levels that inhibit RDRP and cause LRV1 loss. This attests to the impact of the Leishmania purine uptake and metabolism pathways, which allow even a weak RDRP inhibitor to effectively eradicate LRV1 at micromolar concentrations. Future RDRP inhibitors with increased potency may have potential therapeutic applications for ameliorating the increased Leishmania pathogenicity conferred by LRV1. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Real-time PCR for Leishmania species identification: Evaluation and comparison with classical techniques.

PubMed

de Morais, Rayana Carla Silva; da Costa Oliveira, Cintia Nascimento; de Albuquerque, Suênia da Cunha Gonçalves; Mendonça Trajano Silva, Lays Adrianne; Pessoa-E-Silva, Rômulo; Alves da Cruz, Heidi Lacerda; de Brito, Maria Edileuza Felinto; de Paiva Cavalcanti, Milena

2016-06-01

Cutaneous leishmaniasis (CL) is a parasitic disease caused by various Leishmania species. Several studies have shown that real time quantitative PCR (qPCR) can be used for Leishmania spp. identification by analyzing the melting temperature (Tm). Thus, the aim of this study was to evaluate the viability of qPCR for differentiating eight closely related Leishmania species that cause the same clinical form of the disease and to compare the results with classical techniques. qPCR assays for standardizing the Tm using reference strains were performed. After the CL diagnosis on blood samples of domestic animals, positive samples were analyzed by their Tm and qPCR products were purified and sequenced. Ten human samples previously characterized by Multilocus Enzyme Electrophoresis (MLEE) were also analyzed by Tm. A Restriction Fragment Length Polymorphism (RFLP) assay, a reference test, was also standardized, by using the reference strains. Through standardization of Tm for Leishmania spp., two Tm ranges were created for analysis: 1 (Tm = 78-79.99 °C) included Leishmania (V.) braziliensis, Leishmania (V.) panamensis, Leishmania (V.) lainsoni, Leishmania (V.) guyanensis and Leishmania (V.) shawi; and 2 (Tm = 80-82.2 °C) included Leishmania (V.) naiffi, Leishmania (L.) amazonensis and Leishmania (L.) mexicana. A total of 223 positive blood samples were analyzed, with 58 included in range 1 and 165 in range 2. L. (V.) braziliensis, L. (V.) panamensis and L. (V.) guyanensis were identified by sequencing, while L. (V.) braziliensis, L. (L.) mexicana and L. (V.) panamensis were identified by RFLP analysis. Ten human samples previously characterized by Multilocus Enzyme Electrophoresis (MLEE) were also analyzed by qPCR Tm analysis; five were classified in range 1 and five in range 2. A concordance of 80% was calculated between qPCR and the gold-standard (MLEE) with no significant difference between the methods (p = 0.6499); a similar result was observed for sequencing

Distinct Macrophage Fates after in vitro Infection with Different Species of Leishmania: Induction of Apoptosis by Leishmania (Leishmania) amazonensis, but Not by Leishmania (Viannia) guyanensis

PubMed Central

DaMata, Jarina Pena; Mendes, Bárbara Pinheiro; Maciel-Lima, Kátia; Menezes, Cristiane Alves Silva; Dutra, Walderez Ornelas; Sousa, Lirlândia Pires; Horta, Maria Fátima

2015-01-01

Leishmania is an intracellular parasite in vertebrate hosts, including man. During infection, amastigotes replicate inside macrophages and are transmitted to healthy cells, leading to amplification of the infection. Although transfer of amastigotes from infected to healthy cells is a crucial step that may shape the outcome of the infection, it is not fully understood. Here we compare L. amazonensis and L. guyanensis infection in C57BL/6 and BALB/c mice and investigate the fate of macrophages when infected with these species of Leishmania in vitro. As previously shown, infection of mice results in distinct outcomes: L. amazonensis causes a chronic infection in both strains of mice (although milder in C57BL/6), whereas L. guyanensis does not cause them disease. In vitro, infection is persistent in L. amazonensis-infected macrophages whereas L. guyanensis growth is controlled by host cells from both strains of mice. We demonstrate that, in vitro, L. amazonensis induces apoptosis of both C57BL/6 and BALB/c macrophages, characterized by PS exposure, DNA cleavage into nucleosomal size fragments, and consequent hypodiploidy. None of these signs were seen in macrophages infected with L. guyanensis, which seem to die through necrosis, as indicated by increased PI-, but not Annexin V-, positive cells. L. amazonensis-induced macrophage apoptosis was associated to activation of caspases-3, -8 and -9 in both strains of mice. Considering these two species of Leishmania and strains of mice, macrophage apoptosis, induced at the initial moments of infection, correlates with chronic infection, regardless of its severity. We present evidence suggestive that macrophages phagocytize L. amazonensis-infected cells, which has not been verified so far. The ingestion of apoptotic infected macrophages by healthy macrophages could be a way of amastigote spreading, leading to the establishment of infection. PMID:26513474

Distinct Macrophage Fates after in vitro Infection with Different Species of Leishmania: Induction of Apoptosis by Leishmania (Leishmania) amazonensis, but Not by Leishmania (Viannia) guyanensis.

PubMed

DaMata, Jarina Pena; Mendes, Bárbara Pinheiro; Maciel-Lima, Kátia; Menezes, Cristiane Alves Silva; Dutra, Walderez Ornelas; Sousa, Lirlândia Pires; Horta, Maria Fátima

2015-01-01

Leishmania is an intracellular parasite in vertebrate hosts, including man. During infection, amastigotes replicate inside macrophages and are transmitted to healthy cells, leading to amplification of the infection. Although transfer of amastigotes from infected to healthy cells is a crucial step that may shape the outcome of the infection, it is not fully understood. Here we compare L. amazonensis and L. guyanensis infection in C57BL/6 and BALB/c mice and investigate the fate of macrophages when infected with these species of Leishmania in vitro. As previously shown, infection of mice results in distinct outcomes: L. amazonensis causes a chronic infection in both strains of mice (although milder in C57BL/6), whereas L. guyanensis does not cause them disease. In vitro, infection is persistent in L. amazonensis-infected macrophages whereas L. guyanensis growth is controlled by host cells from both strains of mice. We demonstrate that, in vitro, L. amazonensis induces apoptosis of both C57BL/6 and BALB/c macrophages, characterized by PS exposure, DNA cleavage into nucleosomal size fragments, and consequent hypodiploidy. None of these signs were seen in macrophages infected with L. guyanensis, which seem to die through necrosis, as indicated by increased PI-, but not Annexin V-, positive cells. L. amazonensis-induced macrophage apoptosis was associated to activation of caspases-3, -8 and -9 in both strains of mice. Considering these two species of Leishmania and strains of mice, macrophage apoptosis, induced at the initial moments of infection, correlates with chronic infection, regardless of its severity. We present evidence suggestive that macrophages phagocytize L. amazonensis-infected cells, which has not been verified so far. The ingestion of apoptotic infected macrophages by healthy macrophages could be a way of amastigote spreading, leading to the establishment of infection.

In vitro activity of Tridax procumbens against promastigotes of Leishmania mexicana.

PubMed

Martín-Quintal, Zhelmy; Moo-Puc, Rosa; González-Salazar, Francisco; Chan-Bacab, Manuel J; Torres-Tapia, Luis W; Peraza-Sánchez, Sergio R

2009-04-21

Tridax procumbens is an active herb against leishmaniasis. Leishmaniasis is a group of diseases caused by Leishmania protozoa. We investigated the antileishmanial activity of Tridax procumbens extracts and a pure compound against promastigotes of Leishmania mexicana, the causative agent of cutaneous leishmaniasis in the New World. Extracts and (3S)-16,17-didehydrofalcarinol (1) were obtained by chromatographic methods from Tridax procumbens, and the latter identified by spectroscopic analysis. The effect of these extracts and 1 on the growth inhibition of promastigotes of Leishmania mexicana was evaluated. In order to test the safety of extracts and 1, mammalian cells were treated with them, and cell viability was assessed using trypan blue and MTT. We demonstrated that extracts of Tridax procumbens and 1 showed a pronounced activity against Leishmania mexicana. The methanol extract inhibited promastigotes growth of Leishmania mexicana with a 50% inhibitory concentration (IC(50)) of 3 microg/ml, while oxylipin 1 exhibited the highest inhibition at IC(50)=0.478 microg/ml. In this study we report the biological activity of extracts and (3S)-16,17-didehydrofalcarinol (1), obtained from Tridax procumbens, on the promastigote form of Leishmania mexicana, with no effect upon mammalian cells.

Expansion of Vaccination Services and Strengthening Vaccine-Preventable Diseases Surveillance in Haiti, 2010–2016

PubMed Central

Tohme, Rania A.; Francois, Jeannot; Cavallaro, Kathleen F.; Paluku, Gilson; Yalcouye, Idrissa; Jackson, Ernsley; Wright, Tracie; Adrien, Paul; Katz, Mark A.; Hyde, Terri B.; Faye, Pape; Kimanuka, Francine; Dietz, Vance; Vertefeuille, John; Lowrance, David; Dahl, Benjamin; Patel, Roopal

2017-01-01

Abstract. Following the 2010 earthquake, Haiti was at heightened risk for vaccine-preventable diseases (VPDs) outbreaks due to the exacerbation of long-standing gaps in the vaccination program and subsequent risk of VPD importation from other countries. Therefore, partners supported the Haitian Ministry of Health and Population to improve vaccination services and VPD surveillance. During 2010–2016, three polio, measles, and rubella vaccination campaigns were implemented, achieving a coverage > 90% among children and maintaining Haiti free of those VPDs. Furthermore, Haiti is on course to eliminate maternal and neonatal tetanus, with 70% of communes achieving tetanus vaccine two-dose coverage > 80% among women of childbearing age. In addition, the vaccine cold chain storage capacity increased by 91% at the central level and 285% at the department level, enabling the introduction of three new vaccines (pentavalent, rotavirus, and pneumococcal conjugate vaccines) that could prevent an estimated 5,227 deaths annually. Haiti moved from the fourth worst performing country in the Americas in 2012 to the sixth best performing country in 2015 for adequate investigation of suspected measles/rubella cases. Sentinel surveillance sites for rotavirus diarrhea and meningococcal meningitis were established to estimate baseline rates of those diseases prior to vaccine introduction and to evaluate the impact of vaccination in the future. In conclusion, Haiti significantly improved vaccination services and VPD surveillance. However, high dependence on external funding and competing vaccination program priorities are potential threats to sustaining the improvements achieved thus far. Political commitment and favorable economic and legal environments are needed to maintain these gains. PMID:29064356

Proteases and phosphatases during Leishmania-macrophage interaction: paving the road for pathogenesis.

PubMed

Gómez, María Adelaida; Olivier, Martin

2010-01-01

The outcome of Leishmania infection depends both on host and pathogen factors. Macrophages, the specialized host cells for uptake and intracellular development of Leishmania, play a central role in the control of infection. Leishmania has evolved strategies to downregulate host cell functions, largely mediated by the parasite-induced activation of macrophage protein tyrosine phosphatases (PTPs). We have recently identified PTP1B and TCPTP as two additional PTPs engaged upon Leishmania infection and have unraveled an intimate interaction between the Leishmania surface protease GP63 and host PTPs, which mediates a mechanism of cleavage-dependent PTP activation. Here we discuss new perspectives for GP63-mediated parasite virulence and propose putative mechanisms of GP63 internalization into host macrophages and access to intracellular substrates.

Morinda citrifolia Linn. Reduces Parasite Load and Modulates Cytokines and Extracellular Matrix Proteins in C57BL/6 Mice Infected with Leishmania (Leishmania) amazonensis.

PubMed

Almeida-Souza, Fernando; Cardoso, Flávia de Oliveira; Souza, Bruno Vinicius da Conceição; do Valle, Tânia Zaverucha; de Sá, Joicy Cortez; Oliveira, Iara Dos Santos da Silva; de Souza, Celeste da Silva Freitas; Moragas Tellis, Carla Junqueira; Chagas, Maria do Socorro Dos Santos; Behrens, Maria Dutra; Abreu-Silva, Ana Lúcia; Calabrese, Kátia da Silva

2016-08-01

The absence of an effective vaccine and the debilitating chemotherapy for Leishmaniasis demonstrate the need for developing alternative treatments. Several studies conducted with Morinda citrifolia have shown various biological activities, including antileishmanial activity, however its mechanisms of action are unknown. This study aimed to analyze the in vivo activity of M. citrifolia fruit juice (Noni) against Leishmania (Leishmania) amazonensis in C57BL/6 mice. M. citrifolia fruit juice from the Brazilian Amazon has shown the same constitution of other juices produced around the world and liquid chromatography-mass spectrometry analysis identified five compounds: deacetylasperulosidic acid, asperulosidic acid, rutin, nonioside B and nonioside C. Daily intragastric treatment with Noni was carried out after 55 days of L. (L.) amazonensis infection in C57BL/6 mice. Parasitic loads, cytokine and extracellular protein matrix expressions of the lesion site were analyzed by qPCR. Histopathology of the lesion site, lymph nodes and liver were performed to evaluate the inflammatory processes. Cytokines and biochemical parameters of toxicity from sera were also evaluated. The Noni treatment at 500 mg.kg-1.day-1 for 60 days decreased the lesion size and parasitic load in the footpad infected with L. (L.) amazonensis. The site of infection also showed decreased inflammatory infiltrates and decreased cytokine expressions for IL-12, TNF-α, TGF-β and IL-10. On the other hand, Noni treatment enhanced the extracellular matrix protein expressions of collagen IV, fibronectin and laminin in the infected footpad as well collagen I and II, fibronectin and laminin in the mock-infected footpads. No toxicity was observed at the end of treatment. These data show the efficacy of Noni treatment.

Morinda citrifolia Linn. Reduces Parasite Load and Modulates Cytokines and Extracellular Matrix Proteins in C57BL/6 Mice Infected with Leishmania (Leishmania) amazonensis

PubMed Central

Almeida-Souza, Fernando; Cardoso, Flávia de Oliveira; Souza, Bruno Vinicius da Conceição; do Valle, Tânia Zaverucha; de Sá, Joicy Cortez; Oliveira, Iara dos Santos da Silva; de Souza, Celeste da Silva Freitas; Moragas Tellis, Carla Junqueira; Chagas, Maria do Socorro dos Santos; Behrens, Maria Dutra

2016-01-01

The absence of an effective vaccine and the debilitating chemotherapy for Leishmaniasis demonstrate the need for developing alternative treatments. Several studies conducted with Morinda citrifolia have shown various biological activities, including antileishmanial activity, however its mechanisms of action are unknown. This study aimed to analyze the in vivo activity of M. citrifolia fruit juice (Noni) against Leishmania (Leishmania) amazonensis in C57BL/6 mice. M. citrifolia fruit juice from the Brazilian Amazon has shown the same constitution of other juices produced around the world and liquid chromatography–mass spectrometry analysis identified five compounds: deacetylasperulosidic acid, asperulosidic acid, rutin, nonioside B and nonioside C. Daily intragastric treatment with Noni was carried out after 55 days of L. (L.) amazonensis infection in C57BL/6 mice. Parasitic loads, cytokine and extracellular protein matrix expressions of the lesion site were analyzed by qPCR. Histopathology of the lesion site, lymph nodes and liver were performed to evaluate the inflammatory processes. Cytokines and biochemical parameters of toxicity from sera were also evaluated. The Noni treatment at 500 mg.kg-1.day-1 for 60 days decreased the lesion size and parasitic load in the footpad infected with L. (L.) amazonensis. The site of infection also showed decreased inflammatory infiltrates and decreased cytokine expressions for IL-12, TNF-α, TGF-β and IL-10. On the other hand, Noni treatment enhanced the extracellular matrix protein expressions of collagen IV, fibronectin and laminin in the infected footpad as well collagen I and II, fibronectin and laminin in the mock-infected footpads. No toxicity was observed at the end of treatment. These data show the efficacy of Noni treatment. PMID:27579922

Vaccinating Patients With Inflammatory Bowel Disease

PubMed Central

Reich, Jason; Wasan, Sharmeel

2016-01-01

Patients with inflammatory bowel disease (IBD) are not vaccinated at the same rate as general medical patients. IBD places patients at increased risk for developing vaccine-preventable illnesses, and this risk is further exacerbated by immunosuppressive therapy. Therefore, gastroenterologists should familiarize themselves with health maintenance measures pertaining to patients with IBD. This article highlights the vaccinations required for patients with IBD, especially those who are immunosuppressed: influenza; pneumococcal pneumonia; hepatitis A and B viruses; human papilloma virus; meningococcal disease; tetanus, diphtheria, and pertussis; measles, mumps, and rubella; varicella zoster; and herpes zoster. This article also discusses issues regarding patients with IBD who travel outside of the United States, as well as highlights and provides suggestions for areas of quality improvement that are needed in the field. PMID:27917091

Naturally Occurring Culturable Aerobic Gut Flora of Adult Phlebotomus papatasi, Vector of Leishmania major in the Old World

PubMed Central

Mukhopadhyay, Jaba; Braig, Henk R.; Rowton, Edgar D.; Ghosh, Kashinath

2012-01-01

Background Cutaneous leishmaniasis is a neglected, vector-borne parasitic disease and is responsible for persistent, often disfiguring lesions and other associated complications. Leishmania, causing zoonotic cutaneous leishmaniasis (ZCL) in the Old World are mainly transmitted by the predominant sand fly vector, Phlebotomus papatasi. To date, there is no efficient control measure or vaccine available for this widespread insect-borne infectious disease. Methodology/Principal Findings A survey was carried out to study the abundance of different natural gut flora in P. papatasi, with the long-term goal of generating a paratransgenic sand fly that can potentially block the development of Leishmania in the sand fly gut, thereby preventing transmission of leishmania in endemic disease foci. Sand flies, in particular, P. papatasi were captured from different habitats of various parts of the world. Gut microbes were cultured and identified using 16S ribosomal DNA analysis and a phylogenetic tree was constructed. We found variation in the species and abundance of gut flora in flies collected from different habitats. However, a few Gram-positive, nonpathogenic bacteria including Bacillus flexus and B. pumilus were common in most of the sites examined. Conclusion/Significance Our results indicate that there is a wide range of variation of aerobic gut flora inhabiting sand fly guts, which possibly reflect the ecological condition of the habitat where the fly breeds. Also, some species of bacteria (B. pumilus, and B. flexus) were found from most of the habitats. Important from an applied perspective of dissemination, our results support a link between oviposition induction and adult gut flora. PMID:22629302

VACCINATION--COLLECTIVE RESPONSIBILITY OR VIOLATION OF RIGHTS?

PubMed

Florescu, Laura; Rugina, Aurica; Temneanu, Oana Raluca; Paduraru, Dana Teodora Anton; Matei, Mioara Calipsoana; Safta, Cosmin; Mindru, Dana Elena

2015-01-01

Vaccination is considered to be the most effective and the cheapest medical intervention through which individual and collective immunisation is achieved. Statistics show that, at present, immunisation annually saves 400 million lives and protects approximately 750,000 children against disabilities of varying degrees. Approximately 80% of worldwide children are vaccinated against diphtheria, tetanus, pertussis, polio, measles, etc.; these diseases used to be considered incurable in the past. Vaccines help the body to produce antibodies; they help the immune system to detect germs and inactivate their cells. The immunological protection is installed after a variable period of time following the inoculation and is long lasting. Immunisations can be achieved in several ways: through national immunisation campaigns with general recommendation--they may be compulsory, optional or prophylactic (for the diseases for which a vaccine is available); vaccinations not included in the compulsory immunisation programmes; they may also be targeted to the contagious infectious outbreaks or to groups of population in certain situations. There is no guarantee that a vaccine will provide 100% protection. However, it will significantly reduce the risk of getting an infection. Vaccines have side effects which can be divided into reactions triggered by the vaccine or reactions exacerbated by it, without a causal relationship to the vaccine.

Anti-Leishmania activity of new ruthenium(II) complexes: Effect on parasite-host interaction.

PubMed

Costa, Mônica S; Gonçalves, Yasmim G; Nunes, Débora C O; Napolitano, Danielle R; Maia, Pedro I S; Rodrigues, Renata S; Rodrigues, Veridiana M; Von Poelhsitz, Gustavo; Yoneyama, Kelly A G

2017-10-01

Leishmaniasis is a parasitic disease caused by protozoa of the genus Leishmania. The many complications presented by the current treatment - including high toxicity, high cost and parasite resistance - make the development of new therapeutic agents indispensable. The present study aims to evaluate the anti-Leishmania potential of new ruthenium(II) complexes, cis‑[Ru II (η 2 -O 2 CR)(dppm) 2 ]PF 6 , with dppm=bis(diphenylphosphino)methane and R=4-butylbenzoate (bbato) 1, 4-(methylthio)benzoate (mtbato) 2 and 3-hydroxy-4-methoxybenzoate (hmxbato) 3, in promastigote cytotoxicity and their effect on parasite-host interaction. The cytotoxicity of complexes was analyzed by MTT assay against Leishmania (Leishmania) amazonensis, Leishmania (Viannia) braziliensis, Leishmania (Leishmania) infantum promastigotes and the murine macrophage (RAW 264.7). The effect of complexes on parasite-host interaction was evaluated by in vitro infectivity assay performed in the presence of two different concentrations of each complex: the promastigote IC 50 value and the concentration nontoxic to 90% of RAW 264.7 macrophages. Complexes 1-3 exhibited potent cytotoxic activity against all Leishmania species assayed. The IC 50 values ranged from 7.52-12.59μM (complex 1); 0.70-3.28μM (complex 2) and 0.52-1.75μM (complex 3). All complexes significantly inhibited the infectivity index at both tested concentrations. The infectivity inhibitions ranged from 37 to 85%. Interestingly, the infectivity inhibitions due to complex action did not differ significantly at either of the tested concentrations, except for the complex 1 against Leishmania (Leishmania) infantum. The infectivity inhibitions resulted from reductions in both percentage of infected macrophages and number of parasites per macrophage. Taken together the results suggest remarkable leishmanicidal activity in vitro by these new ruthenium(II) complexes. Copyright © 2017 Elsevier Inc. All rights reserved.

Vaccination and Health Maintenance Issues to Consider in Patients With Inflammatory Bowel Disease

PubMed Central

Wasan, Sharmeel K.; Farraye, Francis A.

2017-01-01

Patients with inflammatory bowel disease (IBD) do not receive routine preventive care at the same rate as the general population. IBD places patients at increased risk for developing vaccine-preventable illnesses. This risk is further exacerbated by immunosuppressive therapy. This article highlights the necessary vaccinations for IBD patients and the timing of vaccination for immunosuppressed patients, and discusses the health maintenance needs and preventive care issues related to heart disease, smoking, osteoporosis, mental health, cervical cancer, and skin cancer. PMID:29339947

Modeled structure of trypanothione reductase of Leishmania infantum.

PubMed

Singh, Bishal K; Sarkar, Nandini; Jagannadham, M V; Dubey, Vikash K

2008-06-30

Trypanothione reductase is an important target enzyme for structure-based drug design against Leishmania. We used homology modeling to construct a three-dimensional structure of the trypanothione reductase (TR) of Leishmania infantum. The structure shows acceptable Ramachandran statistics and a remarkably different active site from glutathione reductase(GR). Thus, a specific inhibitor against TR can be designed without interfering with host (human) GR activity.

RIPK1 and PGAM5 Control Leishmania Replication through Distinct Mechanisms.

PubMed

Farias Luz, Nivea; Balaji, Sakthi; Okuda, Kendi; Barreto, Aline Silva; Bertin, John; Gough, Peter J; Gazzinelli, Ricardo; Almeida, Roque P; Bozza, Marcelo T; Borges, Valeria M; Chan, Francis Ka-Ming

2016-06-15

Leishmaniasis is an important parasitic disease found in the tropics and subtropics. Cutaneous and visceral leishmaniasis affect an estimated 1.5 million people worldwide. Despite its human health relevance, relatively little is known about the cell death pathways that control Leishmania replication in the host. Necroptosis is a recently identified form of cell death with potent antiviral effects. Receptor interacting protein kinase 1 (RIPK1) is a critical kinase that mediates necroptosis downstream of death receptors and TLRs. Heme, a product of hemoglobin catabolism during certain intracellular pathogen infections, is also a potent inducer of macrophage necroptosis. We found that human visceral leishmaniasis patients exhibit elevated serum levels of heme. Therefore, we examined the impact of heme and necroptosis on Leishmania replication. Indeed, heme potently inhibited Leishmania replication in bone marrow-derived macrophages. Moreover, we found that inhibition of RIPK1 kinase activity also enhanced parasite replication in the absence of heme. We further found that the mitochondrial phosphatase phosphoglycerate mutase family member 5 (PGAM5), a putative downstream effector of RIPK1, was also required for inhibition of Leishmania replication. In mouse infection, both PGAM5 and RIPK1 kinase activity are required for IL-1β expression in response to Leishmania However, PGAM5, but not RIPK1 kinase activity, was directly responsible for Leishmania-induced IL-1β secretion and NO production in bone marrow-derived macrophages. Collectively, these results revealed that RIPK1 and PGAM5 function independently to exert optimal control of Leishmania replication in the host. Copyright © 2016 by The American Association of Immunologists, Inc.

A Historical Overview of the Classification, Evolution, and Dispersion of Leishmania Parasites and Sandflies

PubMed Central

Akhoundi, Mohammad; Kuhls, Katrin; Cannet, Arnaud; Votýpka, Jan; Marty, Pierre; Delaunay, Pascal; Sereno, Denis

2016-01-01

Background The aim of this study is to describe the major evolutionary historical events among Leishmania, sandflies, and the associated animal reservoirs in detail, in accordance with the geographical evolution of the Earth, which has not been previously discussed on a large scale. Methodology and Principal Findings Leishmania and sandfly classification has always been a controversial matter, and the increasing number of species currently described further complicates this issue. Despite several hypotheses on the origin, evolution, and distribution of Leishmania and sandflies in the Old and New World, no consistent agreement exists regarding dissemination of the actors that play roles in leishmaniasis. For this purpose, we present here three centuries of research on sandflies and Leishmania descriptions, as well as a complete description of Leishmania and sandfly fossils and the emergence date of each Leishmania and sandfly group during different geographical periods, from 550 million years ago until now. We discuss critically the different approaches that were used for Leishmana and sandfly classification and their synonymies, proposing an updated classification for each species of Leishmania and sandfly. We update information on the current distribution and dispersion of different species of Leishmania (53), sandflies (more than 800 at genus or subgenus level), and animal reservoirs in each of the following geographical ecozones: Palearctic, Nearctic, Neotropic, Afrotropical, Oriental, Malagasy, and Australian. We propose an updated list of the potential and proven sandfly vectors for each Leishmania species in the Old and New World. Finally, we address a classical question about digenetic Leishmania evolution: which was the first host, a vertebrate or an invertebrate? Conclusions and Significance We propose an updated view of events that have played important roles in the geographical dispersion of sandflies, in relation to both the Leishmania species they

A Historical Overview of the Classification, Evolution, and Dispersion of Leishmania Parasites and Sandflies.

PubMed

Akhoundi, Mohammad; Kuhls, Katrin; Cannet, Arnaud; Votýpka, Jan; Marty, Pierre; Delaunay, Pascal; Sereno, Denis

2016-03-01

The aim of this study is to describe the major evolutionary historical events among Leishmania, sandflies, and the associated animal reservoirs in detail, in accordance with the geographical evolution of the Earth, which has not been previously discussed on a large scale. Leishmania and sandfly classification has always been a controversial matter, and the increasing number of species currently described further complicates this issue. Despite several hypotheses on the origin, evolution, and distribution of Leishmania and sandflies in the Old and New World, no consistent agreement exists regarding dissemination of the actors that play roles in leishmaniasis. For this purpose, we present here three centuries of research on sandflies and Leishmania descriptions, as well as a complete description of Leishmania and sandfly fossils and the emergence date of each Leishmania and sandfly group during different geographical periods, from 550 million years ago until now. We discuss critically the different approaches that were used for Leishmana and sandfly classification and their synonymies, proposing an updated classification for each species of Leishmania and sandfly. We update information on the current distribution and dispersion of different species of Leishmania (53), sandflies (more than 800 at genus or subgenus level), and animal reservoirs in each of the following geographical ecozones: Palearctic, Nearctic, Neotropic, Afrotropical, Oriental, Malagasy, and Australian. We propose an updated list of the potential and proven sandfly vectors for each Leishmania species in the Old and New World. Finally, we address a classical question about digenetic Leishmania evolution: which was the first host, a vertebrate or an invertebrate? We propose an updated view of events that have played important roles in the geographical dispersion of sandflies, in relation to both the Leishmania species they transmit and the animal reservoirs of the parasites.

First report of natural infection of a bush dog (Speothos venaticus) with Leishmania (Leishmania) chagasi in Brazil.

PubMed

Figueiredo, F B; Gremião, I D F; Pereira, S A; Fedulo, L P; Menezes, R C; Balthazar, D A; Schubach, T M P; Madeira, M F

2008-02-01

We report here the first known case of natural infection of a bush dog with Leishmania (Leishmania) chagasi in Brazil. The specimen was captured in the wild in the State of Mato Grosso and is currently being held in captivity at Fundação Jardim Zoológico, Rio de Janeiro, Brazil. The leishmaniasis was diagnosed by culture of promastigote forms in intact skin fragments and their characterization by isoenzyme electrophoresis. This report calls attention to the parasitological and etiological control of certain zoonoses, such as leishmaniasis, in wild animals kept in captivity, especially when animals are exchanged between zoos in Brazil.

Immunogenicity and efficacy of a bivalent DNA vaccine containing LeIF and TSA genes against murine cutaneous leishmaniasis.

PubMed

Maspi, Nahid; Ghaffarifar, Fatemeh; Sharifi, Zohreh; Dalimi, Abdolhossein; Dayer, Mohammad Saaid

2017-03-01

There is no effective vaccine for the prevention and elimination of leishmaniasis. For this reason, we assessed the protective effects of DNA vaccines containing LeIF, TSA genes alone, or LeIF-TSA fusion against cutaneous leishmaniasis pEGFP-N1 plasmid (empty vector) and phosphate buffer saline (PBS) were used as control groups. Therefore, cellular and humoral immune responses were evaluated before and after the challenge with Leishmania major. Lesion diameter was also measured 3-12 weeks after challenge. All immunized mice with plasmid DNA encoding Leishmania antigens induced the partial immunity characterized by increased IFN-γ and IgG2a levels compared with control groups (p < 0.001). Furthermore, the immunized mice showed significant reduction in mean lesion sizes compared with mice in empty vector and PBS groups (p < 0.05). The reduction in lesion diameter was 29.3%, 34.1%, and 46.2% less in groups vaccinated with LeIF, TSA, and LeIF-TSA, respectively, than in PBS group at 12th week post infection. IFN/IL-4 and IgG2a/IgG1 ratios indicated that group receiving LeIF-TSA fusion had the highest IFN-γ and IgG2a levels. In this study, DNA immunization promoted Th1 immune response characterized by higher IFN-γ and IgG2a levels and also reduction in lesion size. These results showed that a bivalent vaccine containing two distinct antigens may induce more potent immune responses against leishmaniasis. © 2017 APMIS. Published by John Wiley & Sons Ltd.

Functional Transcriptomics of Wild-Caught Lutzomyia intermedia Salivary Glands: Identification of a Protective Salivary Protein against Leishmania braziliensis Infection

PubMed Central

Carneiro, Marcia W.; Miranda, José Carlos; Clarêncio, Jorge; Barral-Netto, Manoel; Brodskyn, Cláudia; Barral, Aldina; Ribeiro, José M. C.; Valenzuela, Jesus G.; de Oliveira, Camila I.

2013-01-01

Background Leishmania parasites are transmitted in the presence of sand fly saliva. Together with the parasite, the sand fly injects salivary components that change the environment at the feeding site. Mice immunized with Phlebotomus papatasi salivary gland (SG) homogenate are protected against Leishmania major infection, while immunity to Lutzomyia intermedia SG homogenate exacerbated experimental Leishmania braziliensis infection. In humans, antibodies to Lu. intermedia saliva are associated with risk of acquiring L. braziliensis infection. Despite these important findings, there is no information regarding the repertoire of Lu. intermedia salivary proteins. Methods and Findings A cDNA library from the Salivary Glands (SGs) of wild-caught Lu. intermedia was constructed, sequenced, and complemented by a proteomic approach based on 1D SDS PAGE and mass/mass spectrometry to validate the transcripts present in this cDNA library. We identified the most abundant transcripts and proteins reported in other sand fly species as well as novel proteins such as neurotoxin-like proteins, peptides with ML domain, and three small peptides found so far only in this sand fly species. DNA plasmids coding for ten selected transcripts were constructed and used to immunize BALB/c mice to study their immunogenicity. Plasmid Linb-11—coding for a 4.5-kDa protein—induced a cellular immune response and conferred protection against L. braziliensis infection. This protection correlated with a decreased parasite load and an increased frequency of IFN-γ-producing cells. Conclusions We identified the most abundant and novel proteins present in the SGs of Lu. intermedia, a vector of cutaneous leishmaniasis in the Americas. We also show for the first time that immunity to a single salivary protein from Lu. intermedia can protect against cutaneous leishmaniasis caused by L. braziliensis. PMID:23717705

Functional transcriptomics of wild-caught Lutzomyia intermedia salivary glands: identification of a protective salivary protein against Leishmania braziliensis infection.

PubMed

de Moura, Tatiana R; Oliveira, Fabiano; Carneiro, Marcia W; Miranda, José Carlos; Clarêncio, Jorge; Barral-Netto, Manoel; Brodskyn, Cláudia; Barral, Aldina; Ribeiro, José M C; Valenzuela, Jesus G; de Oliveira, Camila I

2013-01-01

Leishmania parasites are transmitted in the presence of sand fly saliva. Together with the parasite, the sand fly injects salivary components that change the environment at the feeding site. Mice immunized with Phlebotomus papatasi salivary gland (SG) homogenate are protected against Leishmania major infection, while immunity to Lutzomyia intermedia SG homogenate exacerbated experimental Leishmania braziliensis infection. In humans, antibodies to Lu. intermedia saliva are associated with risk of acquiring L. braziliensis infection. Despite these important findings, there is no information regarding the repertoire of Lu. intermedia salivary proteins. A cDNA library from the Salivary Glands (SGs) of wild-caught Lu. intermedia was constructed, sequenced, and complemented by a proteomic approach based on 1D SDS PAGE and mass/mass spectrometry to validate the transcripts present in this cDNA library. We identified the most abundant transcripts and proteins reported in other sand fly species as well as novel proteins such as neurotoxin-like proteins, peptides with ML domain, and three small peptides found so far only in this sand fly species. DNA plasmids coding for ten selected transcripts were constructed and used to immunize BALB/c mice to study their immunogenicity. Plasmid Linb-11--coding for a 4.5-kDa protein--induced a cellular immune response and conferred protection against L. braziliensis infection. This protection correlated with a decreased parasite load and an increased frequency of IFN-γ-producing cells. We identified the most abundant and novel proteins present in the SGs of Lu. intermedia, a vector of cutaneous leishmaniasis in the Americas. We also show for the first time that immunity to a single salivary protein from Lu. intermedia can protect against cutaneous leishmaniasis caused by L. braziliensis.

Leishmania donovani chaperonin 10 regulates parasite internalization and intracellular survival in human macrophages.

PubMed

Colineau, Lucie; Clos, Joachim; Moon, Kyung-Mee; Foster, Leonard J; Reiner, Neil E

2017-06-01

Protozoa of the genus Leishmania infect macrophages in their mammalian hosts causing a spectrum of diseases known as the leishmaniases. The search for leishmania effectors that support macrophage infection is a focus of significant interest. One such candidate is leishmania chaperonin 10 (CPN10) which is secreted in exosomes and may have immunosuppressive properties. Here, we report for the first time that leishmania CPN10 localizes to the cytosol of infected macrophages. Next, we generated two genetically modified strains of Leishmania donovani (Ld): one strain overexpressing CPN10 (CPN10+++) and the second, a CPN10 single allele knockdown (CPN10+/-), as the null mutant was lethal. When compared with the wild-type (WT) parental strain, CPN10+/- Ld showed higher infection rates and parasite loads in human macrophages after 24 h of infection. Conversely, CPN10+++ Ld was associated with lower initial infection rates. This unexpected apparent gain-of-function for the knockdown could have been explained either by enhanced parasite internalization or by enhanced intracellular survival. Paradoxically, we found that CPN10+/- leishmania were more readily internalized than WT Ld, but also displayed significantly impaired intracellular survival. This suggests that leishmania CPN10 negatively regulates the rate of parasite uptake by macrophages while being required for intracellular survival. Finally, quantitative proteomics identified an array of leishmania proteins whose expression was positively regulated by CPN10. In contrast, many macrophage proteins involved in innate immunity were negatively regulated by CPN10. Taken together, these findings identify leishmania CPN10 as a novel effector with broad based effects on macrophage cell regulation and parasite survival.

Genetically Validated Drug Targets in Leishmania: Current Knowledge and Future Prospects.

PubMed

Jones, Nathaniel G; Catta-Preta, Carolina M C; Lima, Ana Paula C A; Mottram, Jeremy C

2018-04-13

There has been a very limited number of high-throughput screening campaigns carried out with Leishmania drug targets. In part, this is due to the small number of suitable target genes that have been shown by genetic or chemical methods to be essential for the parasite. In this perspective, we discuss the state of genetic target validation in the field of Leishmania research and review the 200 Leishmania genes and 36 Trypanosoma cruzi genes for which gene deletion attempts have been made since the first published case in 1990. We define a quality score for the different genetic deletion techniques that can be used to identify potential drug targets. We also discuss how the advances in genome-scale gene disruption techniques have been used to assist target-based and phenotypic-based drug development in other parasitic protozoa and why Leishmania has lacked a similar approach so far. The prospects for this scale of work are considered in the context of the application of CRISPR/Cas9 gene editing as a useful tool in Leishmania.

Leishmania, microbiota and sand fly immunity.

PubMed

Telleria, Erich Loza; Martins-da-Silva, Andrea; Tempone, Antonio Jorge; Traub-Csekö, Yara Maria

2018-06-20

In this review, we explore the state-of-the-art of sand fly relationships with microbiota, viruses and Leishmania, with particular emphasis on the vector immune responses. Insect-borne diseases are a major public health problem in the world. Phlebotomine sand flies are proven vectors of several aetiological agents including viruses, bacteria and the trypanosomatid Leishmania, which are responsible for diseases such as viral encephalitis, bartonellosis and leishmaniasis, respectively. All metazoans in nature coexist intimately with a community of commensal microorganisms known as microbiota. The microbiota has a fundamental role in the induction, maturation and function of the host immune system, which can modulate host protection from pathogens and infectious diseases. We briefly review viruses of public health importance present in sand flies and revisit studies done on bacterial and fungal gut contents of these vectors. We bring this information into the context of sand fly development and immune responses. We highlight the immunity mechanisms that the insect utilizes to survive the potential threats involved in these interactions and discuss the recently discovered complex interactions among microbiota, sand fly, Leishmania and virus. Additionally, some of the alternative control strategies that could benefit from the current knowledge are considered.

Leishmania (Viannia) naiffi: rare enough to be neglected?

PubMed

Fagundes-Silva, Giselle Aparecida; Romero, Gustavo Adolfo Sierra; Cupolillo, Elisa; Yamashita, Ellen Priscila Gadelha; Gomes-Silva, Adriano; Guerra, Jorge Augusto de Oliveira; Da-Cruz, Alda Maria

2015-09-01

In the Brazilian Amazon, American tegumentary leishmaniasis (ATL) is endemic and presents a wide spectrum of clinical manifestations due, in part, to the circulation of at least seven Leishmania species. Few reports of Leishmania (Viannia) naiffi infection suggest that its occurrence is uncommon and the reported cases present a benign clinical course and a good response to treatment. This study aimed to strengthen the clinical and epidemiological importance of L. (V.) naiffi in the Amazon Region (Manaus, state of Amazonas) and to report therapeutic failure in patients infected with this species. Thirty Leishmania spp samples isolated from cutaneous lesions were characterised by multilocus enzyme electrophoresis. As expected, the most common species was Leishmania (V.) guyanensis (20 cases). However, a relevant number of L. (V.) naiffi patients (8 cases) was observed, thus demonstrating that this species is not uncommon in the region. No patient infected with L. (V.) naiffi evolved to spontaneous cure until the start of treatment, which indicated that this species may not have a self-limiting nature. In addition, two of the patients experienced a poor response to antimonial or pentamidine therapy. Thus, either ATL cases due to L. (V.) naiffi cannot be as uncommon as previously thought or this species is currently expanding in this region.

Genome-wide mapping reveals single-origin chromosome replication in Leishmania, a eukaryotic microbe.

PubMed

Marques, Catarina A; Dickens, Nicholas J; Paape, Daniel; Campbell, Samantha J; McCulloch, Richard

2015-10-19

DNA replication initiates on defined genome sites, termed origins. Origin usage appears to follow common rules in the eukaryotic organisms examined to date: all chromosomes are replicated from multiple origins, which display variations in firing efficiency and are selected from a larger pool of potential origins. To ask if these features of DNA replication are true of all eukaryotes, we describe genome-wide origin mapping in the parasite Leishmania. Origin mapping in Leishmania suggests a striking divergence in origin usage relative to characterized eukaryotes, since each chromosome appears to be replicated from a single origin. By comparing two species of Leishmania, we find evidence that such origin singularity is maintained in the face of chromosome fusion or fission events during evolution. Mapping Leishmania origins suggests that all origins fire with equal efficiency, and that the genomic sites occupied by origins differ from related non-origins sites. Finally, we provide evidence that origin location in Leishmania displays striking conservation with Trypanosoma brucei, despite the latter parasite replicating its chromosomes from multiple, variable strength origins. The demonstration of chromosome replication for a single origin in Leishmania, a microbial eukaryote, has implications for the evolution of origin multiplicity and associated controls, and may explain the pervasive aneuploidy that characterizes Leishmania chromosome architecture.

Natural Products: Insights into Leishmaniasis Inflammatory Response

PubMed Central

Rodrigues, Igor A.; Mazotto, Ana Maria; Cardoso, Verônica; Alves, Renan L.; Amaral, Ana Claudia F.; Silva, Jefferson Rocha de Andrade; Pinheiro, Anderson S.; Vermelho, Alane B.

2015-01-01

Leishmaniasis is a vector-borne disease that affects several populations worldwide, against which there are no vaccines available and the chemotherapy is highly toxic. Depending on the species causing the infection, the disease is characterized by commitment of tissues, including the skin, mucous membranes, and internal organs. Despite the relevance of host inflammatory mediators on parasite burden control, Leishmania and host immune cells interaction may generate an exacerbated proinflammatory response that plays an important role in the development of leishmaniasis clinical manifestations. Plant-derived natural products have been recognized as bioactive agents with several properties, including anti-protozoal and anti-inflammatory activities. The present review focuses on the antileishmanial activity of plant-derived natural products that are able to modulate the inflammatory response in vitro and in vivo. The capability of crude extracts and some isolated substances in promoting an anti-inflammatory response during Leishmania infection may be used as part of an effective strategy to fight the disease. PMID:26538837

Therapeutic immunization with radio-attenuated Leishmania parasites through i.m. route revealed protection against the experimental murine visceral leishmaniasis.

PubMed

Datta, Sanchita; Manna, Madhumita; Khanra, Supriya; Ghosh, Moumita; Bhar, Radhaballav; Chakraborty, Anindita; Roy, Syamal

2012-07-01

After our promising results from prophylactic and therapeutic study (i.p. route) with the radio-attenuated Leishmania donovani parasites against experimental murine visceral leishmaniasis, we prompted to check their therapeutic efficacy through i.m route. BALB/c mice were infected with highly virulent L. donovani parasites. After 75 days, mice were treated with gamma (γ)-irradiated parasites. A second therapeutic immunization was given after 15 days of first immunization. The protection against kala-azar was estimated with the reduction of Leishman-Donovan unit from spleen and liver that scored up to 80% and 93%, respectively, while a twofold increase in nitric oxide (NO) and reactive oxygen species (ROS) productions has been observed in the immunized groups of animals. These groups of mice also showed disease regression by skewing Th2 cytokines (IL-10) towards Th1 cytokine (IFN-γ) bias along with the increased generation of NO and ROS, while the infected control group of mice without such treatment surrendered to the disease. Establishment of Th1 ambience in the treated groups has also been supported from the measured antileishmanial antibody IgG subsets (IgG2a and IgG1) with higher anti-soluble Leishmania antigen-specific IgG2a titer. As seen in our previous studies, doses of attenuation by γ-radiation should be taken into serious consideration. Attenuation of parasites at 50 Gy of absorbed dose of gamma rays has not worked well. Thus, therapeutic use of L. donovani parasites radio-attenuated at particular doses can be exploited as a promising vaccine agent. Absence of any adjuvant may increase its acceptability as vaccine candidate further.

In vivo and in vitro phagocytosis of Leishmania (Leishmania) amazonensis promastigotes by B-1 cells.

PubMed

Geraldo, M M; Costa, C R; Barbosa, F M C; Vivanco, B C; Gonzaga, W F K M; Novaes E Brito, R R; Popi, A F; Lopes, J D; Xander, P

2016-06-01

Leishmaniasis is caused by Leishmania parasites that infect several cell types. The promastigote stage of Leishmania is internalized by phagocytic cells and transformed into the obligate intracellular amastigote form. B-1 cells are a subpopulation of B cells that are able to differentiate in vitro and in vivo into mononuclear phagocyte-like cells with phagocytic properties. B-1 cells use several receptors for phagocytosis, such as the mannose receptor and third complement receptor. Leishmania binds to the same receptors on macrophages. In this study, we demonstrated that phagocytes derived from B-1 cells (B-1 CDP) were able to internalize promastigotes of L. (L.) amazonensis in vitro. The internalized promastigotes differentiated into amastigotes. Our results showed that the phagocytic index was higher in B-1 CDP compared to peritoneal macrophages and bone marrow-derived macrophages. The in vivo phagocytic ability of B-1 cells was also demonstrated. Parasites were detected inside purified B-1 cells after intraperitoneal infection with L. (L.) amazonensis promastigotes. Intraperitoneal stimulation with the parasites led to an increase in both IL-10 and TNF-α. These results highlight the importance of studying B-1 CDP cells as phagocytic cells that can participate and contribute to immunity to parasites. © 2016 John Wiley & Sons Ltd.

Biodistribution of meglumine antimoniate in healthy and Leishmania (Leishmania) infantum chagasi-infected BALB/c mice.

PubMed

Borborema, Samanta Etel Treiger; Osso, João Alberto; Andrade, Heitor Franco de; Nascimento, Nanci do

2013-08-01

Pentavalent antimonials such as meglumine antimoniate (MA) are the primary treatments for leishmaniasis, a complex disease caused by protozoan parasites of the genus Leishmania . Despite over 70 years of clinical use, their mechanisms of action, toxicity and pharmacokinetics have not been fully elucidated. Radiotracer studies performed on animals have the potential to play a major role in pharmaceutical development. The aims of this study were to prepare an antimony radiotracer by neutron irradiation of MA and to determine the biodistribution of MA in healthy and Leishmania (Leishmania) infantum chagasi-infected mice. MA (Glucantime®) was neutron irradiated inside the IEA-R1 nuclear reactor, producing two radioisotopes, ¹²²Sb and ¹²⁴Sb, with high radionuclidic purity and good specific activity. This irradiated compound presented anti-leishmanial activity similar to that of non-irradiated MA in both in vitro and in vivo evaluations. In the biodistribution studies, healthy mice showed higher uptake of antimony in the liver than infected mice and elimination occurred primarily through biliary excretion, with a small proportion of the drug excreted by the kidneys. The serum kinetic curve was bi-exponential, with two compartments: the central compartment and another compartment associated with drug excretion. Radiotracers, which can be easily produced by neutron irradiation, were demonstrated to be an interesting tool for answering several questions regarding antimonial pharmacokinetics and chemotherapy.

Functional Characterization of Monomeric GTPase Rab1 in the Secretory Pathway of Leishmania.

PubMed

Bahl, Surbhi; Parashar, Smriti; Malhotra, Himanshu; Raje, Manoj; Mukhopadhyay, Amitabha

2015-12-11

Leishmania secretes a large number of its effectors to the extracellular milieu. However, regulation of the secretory pathway in Leishmania is not well characterized. Here, we report the cloning, expression, and characterization of the Rab1 homologue from Leishmania. We have found that LdRab1 localizes in Golgi in Leishmania. To understand the role of LdRab1 in the secretory pathway of Leishmania, we have generated transgenic parasites overexpressing GFP-LdRab1:WT, GFP-LdRab1:Q67L (a GTPase-deficient dominant positive mutant of Rab1), and GFP-LdRab1:S22N (a GDP-locked dominant negative mutant of Rab1). Surprisingly, our results have shown that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N does not disrupt the trafficking and localization of hemoglobin receptor in Leishmania. To determine whether the Rab1-dependent secretory pathway is conserved in parasites, we have analyzed the role of LdRab1 in the secretion of secretory acid phosphatase and Ldgp63 in Leishmania. Our results have shown that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N significantly inhibits the secretion of secretory acid phosphatase by Leishmania. We have also found that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N retains RFP-Ldgp63 in Golgi and blocks the secretion of Ldgp63, whereas the trafficking of RFP-Ldgp63 in GFP-LdRab1:WT-expressing cells is unaltered in comparison with control cells. Taken together, our results have shown that the Rab1-regulated secretory pathway is well conserved, and hemoglobin receptor trafficking follows an Rab1-independent secretory pathway in Leishmania. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Functional Characterization of Monomeric GTPase Rab1 in the Secretory Pathway of Leishmania*

PubMed Central

Bahl, Surbhi; Parashar, Smriti; Malhotra, Himanshu; Raje, Manoj; Mukhopadhyay, Amitabha

2015-01-01

Leishmania secretes a large number of its effectors to the extracellular milieu. However, regulation of the secretory pathway in Leishmania is not well characterized. Here, we report the cloning, expression, and characterization of the Rab1 homologue from Leishmania. We have found that LdRab1 localizes in Golgi in Leishmania. To understand the role of LdRab1 in the secretory pathway of Leishmania, we have generated transgenic parasites overexpressing GFP-LdRab1:WT, GFP-LdRab1:Q67L (a GTPase-deficient dominant positive mutant of Rab1), and GFP-LdRab1:S22N (a GDP-locked dominant negative mutant of Rab1). Surprisingly, our results have shown that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N does not disrupt the trafficking and localization of hemoglobin receptor in Leishmania. To determine whether the Rab1-dependent secretory pathway is conserved in parasites, we have analyzed the role of LdRab1 in the secretion of secretory acid phosphatase and Ldgp63 in Leishmania. Our results have shown that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N significantly inhibits the secretion of secretory acid phosphatase by Leishmania. We have also found that overexpression of GFP-LdRab1:Q67L or GFP-LdRab1:S22N retains RFP-Ldgp63 in Golgi and blocks the secretion of Ldgp63, whereas the trafficking of RFP-Ldgp63 in GFP-LdRab1:WT-expressing cells is unaltered in comparison with control cells. Taken together, our results have shown that the Rab1-regulated secretory pathway is well conserved, and hemoglobin receptor trafficking follows an Rab1-independent secretory pathway in Leishmania. PMID:26499792

Micro/nanoparticle adjuvants for antileishmanial vaccines: present and future trends.

PubMed

Badiee, Ali; Heravi Shargh, Vahid; Khamesipour, Ali; Jaafari, Mahmoud Reza

2013-01-21

Leishmania infection continues to have a major impact on public health inducing significant morbidity and mortality mostly in the poorest populations. Drug resistance, toxicity and side effects associated with expensive chemotherapeutic treatments and difficult reservoir control emphasize the need for a safe and effective vaccine which is not available yet. Although, Leishmanization (LZ) was shown to be effective against cutaneous leishmaniasis, standardization and safety are the main problems of LZ. First generation killed parasites demonstrated limited efficacy in phase 3 trials and moreover well defined molecules have not reached to phase 3 yet. Limited efficacy in vaccines against leishmaniasis is partly due to lack of an appropriate adjuvant. Hence, the use of particulate delivery systems as carriers for antigen and/or immunostimulatory adjuvants for effective delivery to the antigen-presenting cells (APCs) is a valuable strategy to enhance vaccine efficacies. Particle-based delivery systems such as emulsions, liposomes, virosomes, and polymeric microspheres have the potential for successfully delivering antigens, which can then be further improved via incorporation of additional antigenic or immustimulatory adjuvant components in or onto the particle carrier system. In this review, we have attempted to provide a list of particulate vaccine delivery systems involved in the production of candidate leishmaniasis vaccines and introduced some potentially useful vaccine delivery systems for leishmaniasis in future experiments. In conclusion, combination vaccines (adjuvant systems) composed of candidate antigens and more importantly well-developed particulate delivery systems, such as lipid-based particles containing immunostimulatory adjuvants, have a chance to succeed as antileishmanial vaccines. Copyright © 2012 Elsevier Ltd. All rights reserved.

[Health economic evaluation of a 23 value pneumococcal polysaccharide vaccination pilot programme among elderly chronic obstructive pulmonary disease patients in China].

PubMed

Qiu, Y P; Zhao, K; Li, X; Shi, L W; Guo, W D; Qi, X R; Sui, B Y; Zhou, R M

2016-12-06

Objective: From the perspective of health economics, to evaluate 23 pneumococcal polysaccharide vaccination programme among chronic obstructive pulmonary disease (COPD) patient. Methods: In the pilot counties of the project of integrated care pathway for COPD patient (Hanbin district of Hanzhong city in Shanxi Province, Qianjian district of Qingqing city, Huandao district of Qindao city in Shangdong Province, Wen county of Jiaozuo city in Henan Province), information of insurance participants of New Rural Cooperative Medical System (NRCS) was collected by local NRCM information system, which included general information as well as records of medical care and medical fee. Nonprobability sampling method was applied to select a total of 860 objects, who were over 60 years old with local household registration, hospitalized within one recent year due to COPD acute exacerbation, and without vaccination of 23 voluntary pneumococcal polysaccharide vaccine within 3 years. A quasi-experimental design without control group was adopted. Objects were vaccinated with 23-valent pneumococcal polysaccharide vaccine from January to December in 2013, then were followed up from January in 2014 for one year. Data of effectiveness and medical cost was collected by self-designed questionnaire and (Chinese version). Paired rank sum test applied to test the difference of quality of life, number and direct medical cost of treatment (including outpatient treatment and hospitalization) due to COPD acute exacerbation, one year before and after intervention. The incremental cost-effectiveness ratio (ICER) and cost-benefit ratio (CBR) of the programme were calculated. Results: By January 2014, eight hundred sixty objects were vaccinated. By January 2015, seven hundred eighty eight objects were followed up, with 72 cases withdrawed (8.4%). On average, COPD patients reduced 1.12±2.51 treatments due to acute exacerbation, including 0.28±2.09 outpatient

Verbascoside Inhibits Promastigote Growth and Arginase Activity of Leishmania amazonensis.

PubMed

Maquiaveli, Claudia C; Lucon-Júnior, João F; Brogi, Simone; Campiani, Giuseppe; Gemma, Sandra; Vieira, Paulo C; Silva, Edson R

2016-05-27

Verbascoside (1) is a phenylethanoid glycoside that has antileishmanial activity against Leishmania infantum and Leishmania donovani. In this study, we verified the activity of 1 on Leishmania amazonensis and arginase inhibition. Compound 1 showed an EC50 of 19 μM against L. amazonensis promastigotes and is a competitive arginase inhibitor (Ki = 0.7 μM). Docking studies were performed to assess the interaction of 1 with arginase at the molecular level. Arginase is an enzyme of the polyamine biosynthesis pathway that is important to parasite infectivity, and the results of our study suggest that 1 could be useful to develop new approaches for treating leishmaniasis.

Dissecting Leishmania infantum Energy Metabolism - A Systems Perspective

PubMed Central

Subramanian, Abhishek; Jhawar, Jitesh; Sarkar, Ram Rup

2015-01-01

Leishmania infantum, causative agent of visceral leishmaniasis in humans, illustrates a complex lifecycle pertaining to two extreme environments, namely, the gut of the sandfly vector and human macrophages. Leishmania is capable of dynamically adapting and tactically switching between these critically hostile situations. The possible metabolic routes ventured by the parasite to achieve this exceptional adaptation to its varying environments are still poorly understood. In this study, we present an extensively reconstructed energy metabolism network of Leishmania infantum as an attempt to identify certain strategic metabolic routes preferred by the parasite to optimize its survival in such dynamic environments. The reconstructed network consists of 142 genes encoding for enzymes performing 237 reactions distributed across five distinct model compartments. We annotated the subcellular locations of different enzymes and their reactions on the basis of strong literature evidence and sequence-based detection of cellular localization signal within a protein sequence. To explore the diverse features of parasite metabolism the metabolic network was implemented and analyzed as a constraint-based model. Using a systems-based approach, we also put forth an extensive set of lethal reaction knockouts; some of which were validated using published data on Leishmania species. Performing a robustness analysis, the model was rigorously validated and tested for the secretion of overflow metabolites specific to Leishmania under varying extracellular oxygen uptake rate. Further, the fate of important non-essential amino acids in L. infantum metabolism was investigated. Stage-specific scenarios of L. infantum energy metabolism were incorporated in the model and key metabolic differences were outlined. Analysis of the model revealed the essentiality of glucose uptake, succinate fermentation, glutamate biosynthesis and an active TCA cycle as driving forces for parasite energy metabolism

In silico studies on tryparedoxin peroxidase of Leishmania infantum: structural aspects.

PubMed

Singh, Bishal Kumar; Dubey, Vikash Kumar

2009-09-01

Tryparedoxin peroxidase (TryP) is a key enzyme of the trypanothione-dependent metabolism for removal of oxidative stress in leishmania. These enzymes function as antioxidants through their peroxidase and peroxynitrite reductase activities. Inhibitors of this enzyme are presumed to be antilesihmania drugs and structural studies are prerequisite of rational drug design. We have constructed three dimensional structure of TryP of Leishmania infantum using comparative modeling. Structural analysis reveals several interesting features. Moreover, it shows remarkable structural difference with human host glutathione peroxidase, an enzyme involved in similar function and TryP from Leishmania major.

Use of a LiESP/QA-21 Vaccine (CaniLeish) Stimulates an Appropriate Th1-Dominated Cell-Mediated Immune Response in Dogs

PubMed Central

Moreno, Javier; Vouldoukis, Ioannis; Martin, Virginie; McGahie, David; Cuisinier, Anne-Marie; Gueguen, Sylvie

2012-01-01

Canine leishmaniasis is an important zoonotic disease of dogs. The clinical outcome of infection is variable, with the efficiency of the immune response being the key determining factor. There is now a general consensus that a predominant Th1 immune profile in an overall mixed Th1/Th2 response is associated with resistance in dogs, and the absence of a strong Th1 influence is associated with a progression to clinical disease. As a result, there has been a growing demand for vaccines that can induce a specific, strong Th1 response. In this study, we measured the impact of a primary course of a newly available LiESP/QA-21 vaccine on selected humoral and cellular markers of the canine immune response during the onset of immunity. All vaccinated dogs developed a humoral response characterised by IgG2 production. More importantly, vaccinated dogs developed significantly stronger cell-mediated immunity responses than did control dogs. Vaccination induced specific cellular reactivity to soluble Leishmania antigens, with a Leishmania-specific lymphoproliferation (p = 0.0072), characterised by an increased population of T lymphocytes producing IFN-γ (p = 0.0021) and a significant ability of macrophages to reduce intracellular parasite burdens in vitro after co-culture with autologous lymphocytes (p = 0.0014). These responses were correlated with induction of the NOS pathway and production of NO derivatives, which has been shown to be an important leishmanicidal mechanism. These results confirm that vaccination with LiESP/QA-21 induces an appropriate Th1-profile cell-mediated response within three weeks of completing the primary course, and that this response effectively reduces the parasite load in pre-infected macrophages in vitro. PMID:22724031

Predicting asthma exacerbations in children.

PubMed

Forno, Erick; Celedón, Juan C

2012-01-01

This review critically assesses recently published literature on predicting asthma exacerbations in children, while also providing general recommendations for future research in this field. Current evidence suggests that every effort should be made to provide optimal treatment to achieve adequate asthma control, as this will significantly reduce the risk of severe disease exacerbations. Children who have had at least one asthma exacerbation in the previous year are at highest risk for subsequent exacerbations, regardless of disease severity and/or control. Although several tools and biomarkers to predict asthma exacerbations have been recently developed, these approaches need further validation and/or have only had partial success in identifying children at risk. Although considerable progress has been made, much remains to be done. Future studies should clearly differentiate severe asthma exacerbations due to inadequate asthma control from those occurring in children whose asthma is well controlled, utilize standardized definitions of asthma exacerbations, and use a systematic approach to identify the best predictors after accounting for the multiple dimensions of the problem. Our ability to correctly predict the development of severe asthma exacerbations in an individual child should improve in parallel with increased knowledge and/or understanding of the complex interactions among genetic, environmental (e.g. viral infections) and lifestyle (e.g. adherence to treatment) factors underlying these events.

Molecular detection of Leishmania infantum and Leishmania tropica in rodent species from endemic cutaneous leishmaniasis areas in Morocco.

PubMed

Echchakery, Mohamed; Chicharro, Carmen; Boussaa, Samia; Nieto, Javier; Carrillo, Eugenia; Sheila, Ortega; Moreno, Javier; Boumezzough, Ali

2017-10-02

Leishmaniasis remains a major public health problem in African nations, including Morocco, where little is known about the vertebrate reservoirs involved in the causal parasites' transmission cycles. The present study investigates the role of rodent species as potential reservoirs of Leishmania spp. in central Morocco, where both L. tropica and L. infantum have been reported. Rodents were caught from 22 sites in central Morocco, by using Sherman metal traps, and identified morphologically. For each specimen, genomic DNA was extracted from different tissues using the Speed Tools DNA extraction Kit. Then, samples were PCR-analyzed, targeting the SSU rRNA gene to detect Leishmania spp. DNA, followed by amplification of the internal transcribed spacer 1 (ITS1) and its sequencing to identify the species. A total of 197 rodents belonging to ten species were captured and identified: Rattus rattus (40.61%), Mus musculus (25.38%), Apodemus sylvaticus (8.63%), Mus spretus (7.11%), Meriones shawi (5.58%), Rattus norvegicus (4.57%), Meriones libycus (3.05%), Mastomys erythroleucus (2.03%), Gerbillus campestris (2.03%) and Lemniscomys barbarus (1.01%). Molecular analysis revealed the presence of Leishmania species in 18 specimens: six R. rattus (out of 80 captured; 7.5%), 11 M. musculus (out of 50 captured; 22%), and one R. norvegicus (out of 9 captured; 11.11%). To the best of our knowledge, L. infantum and L. tropica were identified in rodent species for the first time in Morocco. These findings suggest that rodent species may be involved in L. infantum and L. tropica transmission cycles in this country but that further studies are needed to confirm their role as reservoirs of Leishmania species in Morocco.

Serological and molecular survey of Leishmania parasites in apparently healthy dogs in the West Bank, Palestine

PubMed Central

2012-01-01

Background Canine visceral leishmaniasis (CVL) is caused by Leishmania infantum in all Mediterranean countries. The Leishmania parasite is transmitted by the bite of a corresponding sand fly vector and primarily maintained in nature by wild and domestic reservoirs, including dogs, foxes and jackals. Infected dogs are the primary reservoir host in endemic regions and are the most significant risk disposing humans to infection. The present study aimed at assessing the prevalence of infection with Leishmania and identification of Leishmania infantum in domestic dogs in the West Bank, Palestine. Methods The infection rate among domestic dogs collected from seven districts in the Palestinian West Bank was investigated by examination of parasites in culture from the buffy coat using serological and molecular methods; based on ELISA, internal transcribed spacer 1 (ITS1) and cysteine protease (CPB) PCR. Results Out of 215 dogs examined for Leishmania, 36 (16.7%) were positive in at least one method. Twenty three animals (11.5%) were positive for Leishmania DNA, whereas, ELISA and culture revealed 16 (7.5%), and 4 (1.5%) respectively. CPB-PCR on one of three culture-positive isolates revealed Leishmania infantum as the causative agent for Leishmania infection in dogs. Conclusions Our study showed that canine leishmania infection is prevalent with varying degrees in all the seven studied districts in Palestine despite the absence of human VL cases in 4 of these districts. The causative agent was confirmed to be Leishmania infantum. PMID:22937916

Leishmania-derived trimannose modulates inflammatory response to significantly reduce Leishmania (L.) major-induced lesions.

PubMed

Grinnage-Pulley, Tara L; Kabotso, Daniel E K; Rintelmann, Chelsea L; Roychoudhury, Rajarshi; Schaut, Robert G; Toepp, Angela J; Gibson-Corley, Katherine N; Parrish, Molly; Pohl, Nicola L B; Petersen, Christine A

2017-10-23

Leishmania lipophosphoglycan (LPG) is a key virulence factor, initiating inflammation resulting in cutaneous lesions. LPG is capped by various oligosaccharides. How these glycans are recognized and how they alter the course of Leishmania infection is poorly understood. Previous studies synthesized α-1,2-trimannose cap sugars on latex beads demonstrated that C57BL/6 mice co-inoculated with L. major and trimannose-coated beads produced significantly higher levels of IL-12 p40 and other pro-inflammatory, type 1 cytokines compared L. major infection alone within the first 48 h of infection. However, as L. major infection typically progress over weeks to months, the role of trimannose in altering disease progression over the course of infection was unknown. Wild-type mice were inoculated with either trimannose or carrier (uncoated) beads, infected with L. major alone, co-inoculated with carrier beads and L. major, or co-inoculated with trimannose beads and L. major Trimannose treatment of L. major- infected mice decreased parasite load and significantly decreased lesion size at 14 days post infection (pi) compared to non-treated, infected mice. Infected, trimannose-treated mice had decreased IL-12p40 and IL-10 secretion and increased IFN-γ at 14 days pi. Mice lacking the ability to detect trimannose, mannose-receptor deleted mice (MR -/- ), when treated with trimannose beads and infected with L. major did not have decreased lesion size. Leishmania -derived trimannose represents a novel immunomodulator that provides early type 1-skewed cytokine production to control parasite load and alter the course of cutaneous leishmaniasis. Copyright © 2017 American Society for Microbiology.

Non-conventional expression systems for the production of vaccine proteins and immunotherapeutic molecules

PubMed Central

Legastelois, Isabelle; Buffin, Sophie; Peubez, Isabelle; Mignon, Charlotte; Sodoyer, Régis; Werle, Bettina

2017-01-01

ABSTRACT The increasing demand for recombinant vaccine antigens or immunotherapeutic molecules calls into question the universality of current protein expression systems. Vaccine production can require relatively low amounts of expressed materials, but represents an extremely diverse category consisting of different target antigens with marked structural differences. In contrast, monoclonal antibodies, by definition share key molecular characteristics and require a production system capable of very large outputs, which drives the quest for highly efficient and cost-effective systems. In discussing expression systems, the primary assumption is that a universal production platform for vaccines and immunotherapeutics will unlikely exist. This review provides an overview of the evolution of traditional expression systems, including mammalian cells, yeast and E.coli, but also alternative systems such as other bacteria than E. coli, transgenic animals, insect cells, plants and microalgae, Tetrahymena thermophila, Leishmania tarentolae, filamentous fungi, cell free systems, and the incorporation of non-natural amino acids. PMID:27905833

Leishmania (L.) mexicana Infected Bats in Mexico: Novel Potential Reservoirs

PubMed Central

Berzunza-Cruz, Miriam; Rodríguez-Moreno, Ángel; Gutiérrez-Granados, Gabriel; González-Salazar, Constantino; Stephens, Christopher R.; Hidalgo-Mihart, Mircea; Marina, Carlos F.; Rebollar-Téllez, Eduardo A.; Bailón-Martínez, Dulce; Balcells, Cristina Domingo; Ibarra-Cerdeña, Carlos N.; Sánchez-Cordero, Víctor; Becker, Ingeborg

2015-01-01

Leishmania (Leishmania) mexicana causes cutaneous leishmaniasis, an endemic zoonosis affecting a growing number of patients in the southeastern states of Mexico. Some foci are found in shade-grown cocoa and coffee plantations, or near perennial forests that provide rich breeding grounds for the sand fly vectors, but also harbor a variety of bat species that live off the abundant fruits provided by these shade-giving trees. The close proximity between sand flies and bats makes their interaction feasible, yet bats infected with Leishmania (L.) mexicana have not been reported. Here we analyzed 420 bats from six states of Mexico that had reported patients with leishmaniasis. Tissues of bats, including skin, heart, liver and/or spleen were screened by PCR for Leishmania (L.) mexicana DNA. We found that 41 bats (9.77%), belonging to 13 species, showed positive PCR results in various tissues. The infected tissues showed no evidence of macroscopic lesions. Of the infected bats, 12 species were frugivorous, insectivorous or nectarivorous, and only one species was sanguivorous (Desmodus rotundus), and most of them belonged to the family Phyllostomidae. The eco-region where most of the infected bats were caught is the Gulf Coastal Plain of Chiapas and Tabasco. Through experimental infections of two Tadarida brasiliensis bats in captivity, we show that this species can harbor viable, infective Leishmania (L.) mexicana parasites that are capable of infecting BALB/c mice. We conclude that various species of bats belonging to the family Phyllostomidae are possible reservoir hosts for Leishmania (L.) mexicana, if it can be shown that such bats are infective for the sand fly vector. Further studies are needed to determine how these bats become infected, how long the parasite remains viable inside these potential hosts and whether they are infective to sand flies to fully evaluate their impact on disease epidemiology. PMID:25629729

Leishmania (L.) mexicana infected bats in Mexico: novel potential reservoirs.

PubMed

Berzunza-Cruz, Miriam; Rodríguez-Moreno, Ángel; Gutiérrez-Granados, Gabriel; González-Salazar, Constantino; Stephens, Christopher R; Hidalgo-Mihart, Mircea; Marina, Carlos F; Rebollar-Téllez, Eduardo A; Bailón-Martínez, Dulce; Balcells, Cristina Domingo; Ibarra-Cerdeña, Carlos N; Sánchez-Cordero, Víctor; Becker, Ingeborg

2015-01-01

Leishmania (Leishmania) mexicana causes cutaneous leishmaniasis, an endemic zoonosis affecting a growing number of patients in the southeastern states of Mexico. Some foci are found in shade-grown cocoa and coffee plantations, or near perennial forests that provide rich breeding grounds for the sand fly vectors, but also harbor a variety of bat species that live off the abundant fruits provided by these shade-giving trees. The close proximity between sand flies and bats makes their interaction feasible, yet bats infected with Leishmania (L.) mexicana have not been reported. Here we analyzed 420 bats from six states of Mexico that had reported patients with leishmaniasis. Tissues of bats, including skin, heart, liver and/or spleen were screened by PCR for Leishmania (L.) mexicana DNA. We found that 41 bats (9.77%), belonging to 13 species, showed positive PCR results in various tissues. The infected tissues showed no evidence of macroscopic lesions. Of the infected bats, 12 species were frugivorous, insectivorous or nectarivorous, and only one species was sanguivorous (Desmodus rotundus), and most of them belonged to the family Phyllostomidae. The eco-region where most of the infected bats were caught is the Gulf Coastal Plain of Chiapas and Tabasco. Through experimental infections of two Tadarida brasiliensis bats in captivity, we show that this species can harbor viable, infective Leishmania (L.) mexicana parasites that are capable of infecting BALB/c mice. We conclude that various species of bats belonging to the family Phyllostomidae are possible reservoir hosts for Leishmania (L.) mexicana, if it can be shown that such bats are infective for the sand fly vector. Further studies are needed to determine how these bats become infected, how long the parasite remains viable inside these potential hosts and whether they are infective to sand flies to fully evaluate their impact on disease epidemiology.

Immunization against leishmaniasis by PLGA nanospheres loaded with an experimental autoclaved Leishmania major (ALM) and Quillaja saponins.

PubMed

Tafaghodi, M; Eskandari, M; Kharazizadeh, M; Khamesipour, A; Jaafari, M R

2010-12-01

Immune responses against the Leishmania antigens are not sufficient to protect against a leishmania challenge. Therefore these antigens need to be potentiated by various adjuvants and delivery systems. In this study, Poly (d,l-lactide-co-glycolide (PLGA) nanospheres as antigen delivery system and Quillaja saponins (QS) as immunoadjuvant have been used to enhance the immune response against autoclaved Leishmania major (ALM). PLGA nanospheres were prepared by a double-emulsion (W/O/W) technique. Particulate characteristics were studied by scanning electron microscopy and particle size analysis. Mean diameter for nanospheres loaded with ALM+QS was 294 ± 106 nm. BALB/c mice were immunized three times in 3-weeks intervals using ALM plus QS loaded nanospheres [(ALM+QS)PLGA], ALM encapsulated with PLGA nanospheres [(ALM)PLGA], (ALM)PLGA + QS, ALM + QS, ALM alone or PBS. The intensity of infection induced by L. major challenge was assessed by measuring size of footpad swelling. The strongest protection, showed by significantly (P < 0.05) smaller footpad, were observed in mice immunized with (ALM)PLGA. The (ALM+QS)PLGA group showed the least protection and highest swelling, while the (ALM)PLGA+QS, ALM+QS and ALM showed an intermediate protection with no significant difference. The mice immunized with ALM and ALM+QS showed the highest IgG2a/IgG1 ratio (P < 0.01), followed by (ALM)PLGA+QS. The highest IFN-γ and lowest IL-4 production was seen in (ALM)PLGA+QS, ALM+QS groups. The highest parasite burden was observed in (ALM)PLGA+QS and (ALM+QS)PLGA groups. It is concluded that PLGA nanospheres as a vaccine delivery system could increase the protective immune responses, but QS adjuvant has a reverse effect on protective immune responses and the least protective responses were seen in the presence of this adjuvant.

Melatonin attenuates Leishmania (L.) amazonensis infection by modulating arginine metabolism.

PubMed

Laranjeira-Silva, Maria Fernanda; Zampieri, Ricardo A; Muxel, Sandra M; Floeter-Winter, Lucile Maria; Markus, Regina P

2015-11-01

Acute inflammatory responses induced by bacteria or fungi block nocturnal melatonin synthesis by rodent pineal glands. Here, we show Leishmania infection does not impair daily melatonin rhythm in hamsters. Remarkably, the attenuated parasite burden and lesion progression in hamsters infected at nighttime was impaired by blockage of melatonin receptors with luzindole, whereas melatonin treatment during the light phase attenuated Leishmania infection. In vitro studies corroborated in vivo observations. Melatonin treatment reduced macrophage expression of Cat-2b, Cat1, and ArgI, genes involved in arginine uptake and polyamine synthesis. Indeed, melatonin reduced macrophage arginine uptake by 40%. Putrescine supplementation reverted the attenuation of infectivity by melatonin indicating that its effect was due to the arrest of parasite replication. This study shows that the Leishmania/host interaction varies in a circadian manner according to nocturnal melatonin pineal synthesis. Our results provide new data regarding Leishmania infectiveness and show new approaches for applying agonists of melatonin receptors in Leishmaniasis therapy. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The contribution of the DNA microarray technology to gene expression profiling in Leishmania spp.: a retrospective.

PubMed

Alonso, Ana; Larraga, Vicente; Alcolea, Pedro J

2018-05-07

The first genome project of any living organism excluding viruses, the gammaproteobacteria Haemophilus influenzae, was completed in 1995. Until the last decade, genome sequencing was very tedious because genome survey sequences (GSS) and/or expressed sequence tags (ESTs) belonging to plasmid, cosmid and artificial chromosome genome libraries had to be sequenced and assembled in silico. Nowadays, no genome is completely assembled actually, because gaps and unassembled contigs are always remaining. However, most represent the whole genome of the organism of origin from a practical point of view. The first genome sequencing projects of trypanosomatid parasites were completed in 2005 following those strategies, and belong to Leishmania major, Trypanosoma cruzi and T. brucei. The functional genomics era rapidly developed on the basis of the microarray technology and has been evolving. In the case of the genus Leishmania, substantial biological information about differentiation in the digenetic life cycle of the parasite has been obtained. Later on, next generation sequencing has revolutionized genome sequencing and functional genomics, leading to more sensitive, accurate results by using much less resources. This new technology is more advantageous, but does not invalidate microarray results. In fact, promising vaccine candidates and drug targets have been found on the basis of microarray-based screening and preliminary proof-of-concept tests. Copyright © 2018. Published by Elsevier B.V.

Vector-transmitted disease vaccines: targeting salivary proteins in transmission (SPIT).

PubMed

McDowell, Mary Ann

2015-08-01

More than half the population of the world is at risk for morbidity and mortality from vector-transmitted diseases, and emerging vector-transmitted infections are threatening new populations. Rising insecticide resistance and lack of efficacious vaccines highlight the need for novel control measures. One such approach is targeting the vector-host interface by incorporating vector salivary proteins in anti-pathogen vaccines. Debate remains about whether vector saliva exposure exacerbates or protects against more severe clinical manifestations, induces immunity through natural exposure or extends to all vector species and associated pathogens. Nevertheless, exploiting this unique biology holds promise as a viable strategy for the development of vaccines against vector-transmitted diseases. Copyright © 2015 Elsevier Ltd. All rights reserved.

Viability and Burden of Leishmania in Extralesional Sites during Human Dermal Leishmaniasis

PubMed Central

Romero, Ibeth; Téllez, Jair; Suárez, Yazmín; Cardona, Maria; Figueroa, Roger; Zelazny, Adrian; Gore Saravia, Nancy

2010-01-01

Background The clinical and epidemiological significance of Leishmania DNA in extralesional sites is obscured by uncertainty of whether the DNA derives from viable parasites. To examine dissemination of Leishmania during active disease and the potential participation of human infection in transmission, Leishmania 7SLRNA was exploited to establish viability and estimate parasite burden in extralesional sites of dermal leishmaniasis patients. Methods The feasibility of discriminating parasite viability by PCR of Leishmania 7SLRNA was evaluated in relation with luciferase activity of luc transfected intracellular amastigotes in dose-response assays of Glucantime cytotoxicity. Monocytes, tonsil swabs, aspirates of normal skin and lesions of 28 cutaneous and 2 mucocutaneous leishmaniasis patients were screened by kDNA amplification/Southern blot. Positive samples were analyzed by quantitative PCR of Leishmania 7SLRNA genes and transcripts. Results 7SLRNA amplification coincided with luciferase activity, confirming discrimination of parasite viability. Of 22 patients presenting kDNA in extralesional samples, Leishmania 7SLRNA genes or transcripts were detected in one or more kDNA positive samples in 100% and 73% of patients, respectively. Gene and transcript copy number amplified from extralesional tissues were comparable to lesions. 7SLRNA transcripts were detected in 13/19 (68%) monocyte samples, 5/12 (42%) tonsil swabs, 4/11 (36%) normal skin aspirates, and 22/25 (88%) lesions; genes were quantifiable in 15/19 (79%) monocyte samples, 12/13 (92%) tonsil swabs, 8/11 (73%) normal skin aspirates. Conclusion Viable parasites are present in extralesional sites, including blood monocytes, tonsils and normal skin of dermal leishmaniasis patients. Leishmania 7SLRNA is an informative target for clinical and epidemiologic investigations of human leishmaniasis. PMID:20856851

Mitigating an undesirable immune response of inherent susceptibility to cutaneous leishmaniosis in a mouse model: the role of the pathoantigenic HISA70 DNA vaccine.

PubMed

Domínguez-Bernal, Gustavo; Horcajo, Pilar; Orden, José A; De La Fuente, Ricardo; Herrero-Gil, Aldara; Ordóñez-Gutiérrez, Lara; Carrión, Javier

2012-08-09

Leishmania major is the major cause of cutaneous leishmaniosis (CL) outside of the Americas. In the present study we have cloned six Leishmania genes (H2A, H2B, H3, H4, A2 and HSP70) into the eukaryotic expression vector pCMVβ-m2a, resulting in pCMV-HISA70m2A, which encodes all six pathoantigenic proteins as a single polyprotein. This expression plasmid has been evaluated as a novel vaccine candidate in the BALB/c mouse model of CL. The DNA vaccine shifted the immune response normally induced by L. major infection away from a Th2-specific pathway to one of basal susceptibility. Immunization with pCMV-HISA70m2A dramatically reduced footpad lesions and lymph node parasite burdens relative to infected control mice. Complete absence of visceral parasite burden was observed in all 12 immunized animals but not in any of the 24 control mice. Moreover, vaccinated mice produced large amounts of IFN-γ, IL-17 and NO at 7 weeks post-infection (pi), and they showed lower arginase activity at the site of infection, lower IL-4 production and a weaker humoral immune response than infected control mice. Taken together, these results demonstrate the ability of the HISA70 vaccine to shift the murine immune response to L. major infection away from an undesirable, Th2-specific pathway to a less susceptible-like pathway involving Th1 and Th17 cytokine profiles.

Uptake of the antileishmania drug tafenoquine follows a sterol-dependent diffusion process in Leishmania.

PubMed

Manzano, José Ignacio; Carvalho, Luis; García-Hernández, Raquel; Poveda, José Antonio; Ferragut, José Antonio; Castanys, Santiago; Gamarro, Francisco

2011-11-01

The present study was designed to elucidate the mechanism of tafenoquine uptake in Leishmania and its sterol dependence. Because tafenoquine is a fluorescent compound, spectrofluorimetric analysis allowed us to monitor its uptake by Leishmania promastigotes and intracellular amastigotes, and to evaluate the effect of temperature, energy and H+ gradient on drug entry. The influence of sterols on tafenoquine uptake in Leishmania parasites was determined in experiments using sterol-depleting agents such as methyl-β-cyclodextrin or cholesterol oxidase. Tafenoquine exhibited fast entry kinetics into Leishmania in an energy-independent, but pH- and temperature-dependent, non-saturable process. Furthermore, sterol depletion decreased tafenoquine uptake. These findings suggest that Leishmania takes up tafenoquine by a diffusion process and that decreases in membrane sterol content may induce a decrease in drug uptake.

Anti-Leishmania and cytotoxic activities of perillaldehyde epoxide synthetic positional isomers.

PubMed

Keesen, Tatjana Souza Lima; da Silva, Larisse Virgolino; da Câmara Rocha, Juliana; Andrade, Luciana Nalone; Lima, Tamires Cardoso; de Sousa, Damião Pergentino

2018-03-13

Leishmaniasis belongs to a complex of zoonotic disease caused by protozoa of the genus Leishmania and is considered a major public health problem. Several essential oil chemical components have inhibitory effect against protozoa, including Leishmania donovani. Thus, the aim of this study was to evaluate for the first time the anti-Leishmania activity of two p-menthane monoterpene isomers (EPER-1: perillaldehyde 1,2-epoxide and EPER-2: perillaldehyde 8,9-epoxide) against L. donovani promastigotes as well as evaluating cytotoxic effect on mononuclear peripheral blood cells. Results of anti-Leishmania assay revealed that EPER-2 (IC 50  = 3.8 μg.mL -1 ) was 16-fold more potent than its isomer EPER-1 (IC 50  = 64.6 μg.mL -1 ). In contrast to PBMC cells, EPER-2 was not cytotoxic (IC 50  > 400 μg.mL -1 ) when compared to positive control. These data suggest that the disposition of epoxide group into the p-menthane skeleton affects the anti-Leishmania activity, being that the presence of the exocyclic epoxide group considerably increased potency. Thus, it was possible to observe that the location of the epoxide group into the p-menthane skeleton resulted in different potencies.

The Dynamics of Lateral Gene Transfer in Genus Leishmania - A Route for Adaptation and Species Diversification.

PubMed

Vikeved, Elisabet; Backlund, Anders; Alsmark, Cecilia

2016-01-01

The genome of Leishmania major harbours a comparably high proportion of genes of prokaryote origin, acquired by lateral gene transfer (LGT). Some of these are present in closely related trypanosomatids, while some are detected in Leishmania only. We have evaluated the impact and destiny of LGT in genus Leishmania. To study the dynamics and fate of LGTs we have performed phylogenetic, as well as nucleotide and amino acid composition analyses within orthologous groups of LGTs detected in Leishmania. A set of universal trypanosomatid LGTs was added as a reference group. Both groups of LGTs have, to some extent, ameliorated to resemble the recipient genomes. However, while virtually all of the universal trypanosomatid LGTs are distributed and conserved in the entire genus Leishmania, the LGTs uniquely present in genus Leishmania are more prone to gene loss and display faster rates of evolution. Furthermore, a PCR based approach has been employed to ascertain the presence of a set of twenty LGTs uniquely present in genus Leishmania, and three universal trypanosomatid LGTs, in ten additional strains of Leishmania. Evolutionary rates and predicted expression levels of these LGTs have also been estimated. Ten of the twenty LGTs are distributed and conserved in all species investigated, while the remainder have been subjected to modifications, or undergone pseudogenization, degradation or loss in one or more species. LGTs unique to the genus Leishmania have been acquired after the divergence of Leishmania from the other trypanosomatids, and are evolving faster than their recipient genomes. This implies that LGT in genus Leishmania is a continuous and dynamic process contributing to species differentiation and speciation. This study also highlights the importance of carefully evaluating these dynamic genes, e.g. as LGTs have been suggested as potential drug targets.

The Dynamics of Lateral Gene Transfer in Genus Leishmania - A Route for Adaptation and Species Diversification

PubMed Central

Vikeved, Elisabet; Backlund, Anders; Alsmark, Cecilia

2016-01-01

Background The genome of Leishmania major harbours a comparably high proportion of genes of prokaryote origin, acquired by lateral gene transfer (LGT). Some of these are present in closely related trypanosomatids, while some are detected in Leishmania only. We have evaluated the impact and destiny of LGT in genus Leishmania. Methodology/Principal Findings To study the dynamics and fate of LGTs we have performed phylogenetic, as well as nucleotide and amino acid composition analyses within orthologous groups of LGTs detected in Leishmania. A set of universal trypanosomatid LGTs was added as a reference group. Both groups of LGTs have, to some extent, ameliorated to resemble the recipient genomes. However, while virtually all of the universal trypanosomatid LGTs are distributed and conserved in the entire genus Leishmania, the LGTs uniquely present in genus Leishmania are more prone to gene loss and display faster rates of evolution. Furthermore, a PCR based approach has been employed to ascertain the presence of a set of twenty LGTs uniquely present in genus Leishmania, and three universal trypanosomatid LGTs, in ten additional strains of Leishmania. Evolutionary rates and predicted expression levels of these LGTs have also been estimated. Ten of the twenty LGTs are distributed and conserved in all species investigated, while the remainder have been subjected to modifications, or undergone pseudogenization, degradation or loss in one or more species. Conclusions/Significance LGTs unique to the genus Leishmania have been acquired after the divergence of Leishmania from the other trypanosomatids, and are evolving faster than their recipient genomes. This implies that LGT in genus Leishmania is a continuous and dynamic process contributing to species differentiation and speciation. This study also highlights the importance of carefully evaluating these dynamic genes, e.g. as LGTs have been suggested as potential drug targets. PMID:26730948

Vaccination against yellow fever in French Guiana: The impact of educational level, negative beliefs and attitude towards vaccination.

PubMed

Koïvogui, Akoï; Carbunar, Aurel; Imounga, Laure-Manuella; Laruade, Christelle; Laube, Sylvaine

Analyze the impact of educational level, negative beliefs and negative attitudes on the yellow fever vaccination coverage (YFVC). This analytical study involved a sample of 2763 people from 866 households. Educational status was described in six levels: No level (Respondent had never attended school), level-1 (respondent left before intermediate school), level-2 (Respondent attended intermediate school), level-3 (respondent attended high school), level-4 (Respondent attended university), Other level (When the level could not be determined). The Attitude towards vaccination was described in terms of person's availability to recommend vaccination to third. The relationships were analyzed by multivariate mixed logistic regression. Among the 2763 peoples, 2039 (73.8%) were vaccinated against yellow fever. People who left high school with or without the French baccalaureate were more likely to be vaccinated against YF than people without any diploma (OR = 1.4; p < 0.05). The probability of being vaccinated among people with negative attitudes was reduced by 40% (OR = 0.6; p < 0.05). Low level of education, negative beliefs and negative attitudes have significant impacts on YFVC. Negatives beliefs and attitudes result often from a major lack of information about the benefits of vaccination. This deficit is exacerbated in persons with low educational level. Copyright © 2016 Elsevier Ltd. All rights reserved.

The protective immune response produced in dogs after primary vaccination with the LiESP/QA-21 vaccine (CaniLeish®) remains effective against an experimental challenge one year later

PubMed Central

2014-01-01

Control of canine leishmaniasis is an important objective for the benefit of dogs living in or visiting endemic areas and for public health because of the zoonotic nature of this disease. Resistance or susceptibility to developing canine leishmaniasis after exposure to Leishmania infantum is primarily determined by the ability of the immune system to develop an appropriate Th1-dominated specific response to the parasite. For this reason there is a need for effective canine vaccines that can decrease the number of dogs developing progressive infections. In this study, we followed the impact of the LiESP/QA-21 canine vaccine (composed of excreted-secreted proteins of L. infantum and the QA-21 saponin adjuvant), recently launched commercially in Europe, on selected humoral and cellular immune parameters following an infectious intravenous challenge with L. infantum promastigotes administered one year after the primary vaccine course. We also followed parasitological parameters to determine the parasitological status of the challenged dogs. In contrast to controls, vaccinated dogs retained significantly stronger cell-mediated immune responses against the parasite despite a virulent challenge and had significantly lower mean parasite burdens at the end of the study, associated with a lower probability of developing active infections. These results confirm that the immune responses generated by vaccination with LiESP/QA-21 are still effective against an intravenous challenge one year after the primary vaccine course. PMID:24964736

The safety and immunogenicity of influenza vaccine in children with asthma in Mexico.

PubMed

Pedroza, Alvaro; Huerta, José G; Garcia, Maria de la Luz; Rojas, Arsheli; López-Martínez, Irma; Penagos, Martín; Franco-Paredes, Carlos; Deroche, Christele; Mascareñas, Cesar

2009-07-01

The morbidity and mortality associated with influenza is substantial in children with asthma. There are no available data on the safety and immunogenicity of influenza vaccine in children with asthma in Latin America. Furthermore, it is unclear if influenza vaccination may cause asthma exacerbations. We conducted a placebo-controlled trial to investigate the safety and immunogenicity of an inactivated trivalent split virus influenza vaccine in children with asthma in Mexico. We also measured the impact of influenza vaccination on pulmonary function tests in this population. The inactivated influenza vaccine was immunogenic and safe in terms of local and systemic side effects compared to placebo. We observed no significant impact on pulmonary function tests among vaccine recipients. Given the significant morbidity associated with influenza in children, strategies to promote increased influenza vaccination coverage in this high-risk group in Latin America and elsewhere are urgently needed.

Mixed infection of Leishmania infantum and Leishmania braziliensis in rodents from endemic urban area of the New World.

PubMed

Ferreira, Eduardo de Castro; Cruz, Israel; Cañavate, Carmen; de Melo, Lutiana Amaral; Pereira, Agnes Antônia Sampaio; Madeira, Filipe A M; Valério, Sofia Alves Nogueira; Cunha, Heitor Morais; Paglia, Adriano Pereira; Gontijo, Célia Maria Ferreira

2015-03-20

In Brazil Leishmania braziliensis and L. infantum are the principal species responsible for cutaneous and visceral leishmaniases, respectively. Domestic dogs are the main reservoirs of visceral leishmaniasis, while rodents and marsupials are the main reservoirs for cutaneous leishmaniasis. It has also been suggested that dogs could play a role in transmission of cutaneous leishmaniasis. The identification of the species of Leishmania, the reservoirs, and the vectors involved in each particular transmission cycle is critical for the establishment of control activities. Belo Horizonte has emerged as an endemic region for leishmaniases, however, epidemiological studies assessing the contribution of wild reservoirs to transmission are scarce in the area. The aim of this study was to investigate Leishmania spp. infection in possible reservoirs of an urbanized area. A high rate of infection was found in small mammals (64.9%) and dogs (DG1 30.4% and DG2 48.6%). The presence of L. infantum and L. braziliensis was detected in small mammals and dogs, and mixed infections by both species were detected in rodents which, to the best of our knowledge, is the first description of this phenomenon in an urban area. Additionally, L. amazonensis was detected in the canine samples. The possible role of these animals as a source of infection of the vector of each species of Leishmania identified should not be overlooked and should be taken into account in future control activities. The results of mixed infection by L. braziliensis and L. infantum in cosmopolitan rodents as M. musculus and R. rattus, may have important implications in the context of the control of leishmaniasis in urban areas, especially when considering that these rodents live in close relationship with human dwellings, especially those in more precarious conditions.

Integrity of the actin cytoskeleton of host macrophages is essential for Leishmania donovani infection.

PubMed

Roy, Saptarshi; Kumar, G Aditya; Jafurulla, Md; Mandal, Chitra; Chattopadhyay, Amitabha

2014-08-01

Visceral leishmaniasis is a vector-borne disease caused by an obligate intracellular protozoan parasite Leishmania donovani. The molecular mechanism involved in internalization of Leishmania is poorly understood. The entry of Leishmania involves interaction with the plasma membrane of host cells. We have previously demonstrated the requirement of host membrane cholesterol in the binding and internalization of L. donovani into macrophages. In the present work, we explored the role of the host actin cytoskeleton in leishmanial infection. We observed a dose-dependent reduction in the attachment of Leishmania promastigotes to host macrophages upon destabilization of the actin cytoskeleton by cytochalasin D. This is accompanied by a concomitant reduction in the intracellular amastigote load. We utilized a recently developed high resolution microscopy-based method to quantitate cellular F-actin content upon treatment with cytochalasin D. A striking feature of our results is that binding of Leishmania promastigotes and intracellular amastigote load show close correlation with cellular F-actin level. Importantly, the binding of Escherichia coli remained invariant upon actin destabilization of host cells, thereby implying specific involvement of the actin cytoskeleton in Leishmania infection. To the best of our knowledge, these novel results constitute the first comprehensive demonstration on the specific role of the host actin cytoskeleton in Leishmania infection. Our results could be significant in developing future therapeutic strategies to tackle leishmaniasis. Copyright © 2014 Elsevier B.V. All rights reserved.

Redirection of sphingolipid metabolism toward de novo synthesis of ethanolamine in Leishmania

PubMed Central

Zhang, Kai; Pompey, Justine M; Hsu, Fong-Fu; Key, Phillip; Bandhuvula, Padmavathi; Saba, Julie D; Turk, John; Beverley, Stephen M

2007-01-01

In most eukaryotes, sphingolipids (SLs) are critical membrane components and signaling molecules. However, mutants of the trypanosomatid protozoan Leishmania lacking serine palmitoyltransferase (spt2−) and SLs grow well, although they are defective in stationary phase differentiation and virulence. Similar phenotypes were observed in sphingolipid (SL) mutant lacking the degradatory enzyme sphingosine 1-phosphate lyase (spl−). This epistatic interaction suggested that a metabolite downstream of SLs was responsible. Here we show that unlike other organisms, the Leishmania SL pathway has evolved to be the major route for ethanolamine (EtN) synthesis, as EtN supplementation completely reversed the viability and differentiation defects of both mutants. Thus Leishmania has undergone two major metabolic shifts: first in de-emphasizing the metabolic roles of SLs themselves in growth, signaling, and maintenance of membrane microdomains, which may arise from the unique combination of abundant parasite lipids; Second, freed of typical SL functional constraints and a lack of alternative routes to produce EtN, Leishmania redirected SL metabolism toward bulk EtN synthesis. Our results thus reveal a striking example of remodeling of the SL metabolic pathway in Leishmania. PMID:17290222

Th1-stimulatory polyproteins of soluble Leishmania donovani promastigotes ranging from 89.9 to 97.1 kDa offers long-lasting protection against experimental visceral leishmaniasis.

PubMed

Kumari, Shraddha; Samant, Mukesh; Misra, Pragya; Khare, Prashant; Sisodia, Brijesh; Shasany, Ajit K; Dube, Anuradha

2008-10-23

Our earlier studies identified a fraction (F2) of Leishmania donovani soluble promastigote antigen belonging to 97.4-68 kDa for its ability to stimulate Th1-type cellular responses in cured visceral leishmaniasis (VL) patients as well as in cured hamsters. A further fractionation of F2-fraction into seven subfractions (F2.1-F2.7) and re-assessment for their immunostimulatory responses revealed that out of these, only four (F2.4-F2.7) belonging to 89.9-97.1 kDa, stimulated remarkable Th1-type cellular responses either individually or in a pooled form (P4-7). In this study these potential subfractions were further assessed for their prophylactic potential in combination with BCG against L. donovani challenge in hamsters. Optimum parasite inhibition ( approximately 99%) was obtained in hamsters vaccinated with pooled subfractions and they survived for 1 year. The protection was further supported by remarkable lymphoproliferative, IFN-gamma and IL-12 responses along with profound delayed type hypersensitivity and increased levels of Leishmania-specific IgG2 antibody as observed on days 45, 90 and 120 post-challenge suggesting that a successful subunit vaccine against VL may require multiple Th1-immunostimulatory proteins. MALDI-TOF-MS/MS analysis of these subfractions further revealed that of the 19 identified immunostimulatory proteins, Elongation factor-2, p45, Heat shock protein-70/83, Aldolase, Enolase, Triosephosphate isomerase, Disulfideisomerase and Calreticulin were the major ones in these subfractions.

Large-scale investigation of Leishmania interaction networks with host extracellular matrix by surface plasmon resonance imaging.

PubMed

Fatoux-Ardore, Marie; Peysselon, Franck; Weiss, Anthony; Bastien, Patrick; Pratlong, Francine; Ricard-Blum, Sylvie

2014-02-01

We have set up an assay to study the interactions of live pathogens with their hosts by using protein and glycosaminoglycan arrays probed by surface plasmon resonance imaging. We have used this assay to characterize the interactions of Leishmania promastigotes with ~70 mammalian host biomolecules (extracellular proteins, glycosaminoglycans, growth factors, cell surface receptors). We have identified, in total, 27 new partners (23 proteins, 4 glycosaminoglycans) of procyclic promastigotes of six Leishmania species and 18 partners (15 proteins, 3 glycosaminoglycans) of three species of stationary-phase promastigotes for all the strains tested. The diversity of the interaction repertoires of Leishmania parasites reflects their dynamic and complex interplay with their mammalian hosts, which depends mostly on the species and strains of Leishmania. Stationary-phase Leishmania parasites target extracellular matrix proteins and glycosaminoglycans, which are highly connected in the extracellular interaction network. Heparin and heparan sulfate bind to most Leishmania strains tested, and 6-O-sulfate groups play a crucial role in these interactions. Numerous Leishmania strains bind to tropoelastin, and some strains are even able to degrade it. Several strains interact with collagen VI, which is expressed by macrophages. Most Leishmania promastigotes interact with several regulators of angiogenesis, including antiangiogenic factors (endostatin, anastellin) and proangiogenic factors (ECM-1, VEGF, and TEM8 [also known as anthrax toxin receptor 1]), which are regulated by hypoxia. Since hypoxia modulates the infection of macrophages by the parasites, these interactions might influence the infection of host cells by Leishmania.

Large-Scale Investigation of Leishmania Interaction Networks with Host Extracellular Matrix by Surface Plasmon Resonance Imaging

PubMed Central

Fatoux-Ardore, Marie; Peysselon, Franck; Weiss, Anthony; Bastien, Patrick; Pratlong, Francine

2014-01-01

We have set up an assay to study the interactions of live pathogens with their hosts by using protein and glycosaminoglycan arrays probed by surface plasmon resonance imaging. We have used this assay to characterize the interactions of Leishmania promastigotes with ∼70 mammalian host biomolecules (extracellular proteins, glycosaminoglycans, growth factors, cell surface receptors). We have identified, in total, 27 new partners (23 proteins, 4 glycosaminoglycans) of procyclic promastigotes of six Leishmania species and 18 partners (15 proteins, 3 glycosaminoglycans) of three species of stationary-phase promastigotes for all the strains tested. The diversity of the interaction repertoires of Leishmania parasites reflects their dynamic and complex interplay with their mammalian hosts, which depends mostly on the species and strains of Leishmania. Stationary-phase Leishmania parasites target extracellular matrix proteins and glycosaminoglycans, which are highly connected in the extracellular interaction network. Heparin and heparan sulfate bind to most Leishmania strains tested, and 6-O-sulfate groups play a crucial role in these interactions. Numerous Leishmania strains bind to tropoelastin, and some strains are even able to degrade it. Several strains interact with collagen VI, which is expressed by macrophages. Most Leishmania promastigotes interact with several regulators of angiogenesis, including antiangiogenic factors (endostatin, anastellin) and proangiogenic factors (ECM-1, VEGF, and TEM8 [also known as anthrax toxin receptor 1]), which are regulated by hypoxia. Since hypoxia modulates the infection of macrophages by the parasites, these interactions might influence the infection of host cells by Leishmania. PMID:24478075

Vaxfectin (registered trademark) Enhances Both Antibody and In Vitro T Cell Responses to Each Component of a 5-gene Plasmodium falciparum Plasmid DNA Vaccine Mixture Administered at Low Doses

DTIC Science & Technology

2010-01-01

received intramus- cular injections in the tibialis anterior muscle pDNA formu lated in PBS or Vaxfectin® using insulin syringes (0.3 ml: Becton Dick... phosphatidylcholine (DSPC) liposomes in Leishmania vaccine stud- ies and report long-term immunity in mice when the adjuvant was added with the

Targeting Leishmania major parasite with peptides derived from a combinatorial phage display library.

PubMed

Rhaiem, Rafik Ben; Houimel, Mehdi

2016-07-01

Cutaneous leishmaniasis (CL) is a global problem caused by intracellular protozoan pathogens of the genus Leishmania for which there are no suitable vaccine or chemotherapy options. Thus, de novo identification of small molecules binding to the Leishmania parasites by direct screening is a promising and appropriate alternative strategy for the development of new drugs. In this study, we used a random linear hexapeptide library fused to the gene III protein of M13 filamentous bacteriophage to select binding peptides to metacyclic promastigotes from a highly virulent strain of Leishmania major (Zymodeme MON-25; MHOM/TN/94/GLC94). After four rounds of stringent selection and amplification, polyclonal and monoclonal phage-peptides directed against L. major metacyclic promastigotes were assessed by ELISA, and the optimal phage-peptides were grown individually and characterized for binding to L. major by monoclonal phage ELISA. The DNA of 42 phage-peptides clones was amplified by PCR, sequenced, and their amino acid sequences deduced. Six different peptide sequences were obtained with frequencies of occurrence ranging from 2.3% to 85.7%. The biological effect of the peptides was assessed in vitro on human monocytes infected with L. major metacyclic promastigotes, and in vivo on susceptible parasite-infected BALB/c mice. The development of cutaneous lesions in the right hind footpads of infected mice after 13 weeks post-infection showed a protection rate of 81.94% with the injected peptide P2. Moreover, Western blots revealed that the P2 peptide interacted with the major surface protease gp63, a protein of 63kDa molecular weight. Moreover, bioinformatics were used to predict the interaction between peptides and the major surface molecule of the L. major. The molecular docking showed that the P2 peptide has the minimum interaction energy and maximum shape complimentarity with the L. major gp63 active site. Our study demonstrated that the P2 peptide occurs at high frequency

The Likelihood of Preventing Respiratory Exacerbations in Children and Adolescents with either Chronic Suppurative Lung Disease or Bronchiectasis

PubMed Central

O’Grady, Kerry-Ann F; Grimwood, Keith

2017-01-01

Chronic suppurative lung disease (CSLD) and bronchiectasis in children and adolescents are important causes of respiratory morbidity and reduced quality of life (QoL), also leading to subsequent premature death during adulthood. Acute respiratory exacerbations in pediatric CSLD and bronchiectasis are important markers of disease control clinically, given that they impact upon QoL and increase health-care-associated costs and can adversely affect future lung functioning. Preventing exacerbations in this population is, therefore, likely to have significant individual, familial, societal, and health-sector benefits. In this review, we focus on therapeutic interventions, such as drugs (antibiotics, mucolytics, hyperosmolar agents, bronchodilators, corticosteroids, non-steroidal anti-inflammatory agents), vaccines and physiotherapy, and care-planning, such as post-hospitalization management and health promotion strategies, including exercise, diet, and reducing exposure to environmental toxicants. The review identified a conspicuous lack of moderate or high-quality evidence for preventing respiratory exacerbations in children and adolescents with CSLD or bronchiectasis. Given the short- and long-term impact of exacerbations upon individuals, their families, and society as a whole, large studies addressing interventions at the primary and tertiary prevention phases are required. This research must include children and adolescents in both developing and developed countries and address long-term health outcomes. PMID:28393062

Immunoliposomes containing Soluble Leishmania Antigens (SLA) as a novel antigen delivery system in murine model of leishmaniasis.

PubMed

Eskandari, Faeze; Talesh, Ghazal Alipour; Parooie, Maryam; Jaafari, Mahmoud Reza; Khamesipour, Ali; Saberi, Zahra; Abbasi, Azam; Badiee, Ali

2014-11-01

Development of new generation of vaccines against leishmaniasis requires adjuvants to elicit the type and intensity of immune response needed for protection. The coupling of target-specific antibodies to the liposomal surface to create immunoliposomes has appeared as a promising way in achieving a liposome active targeting. In this study, immunoliposomes were prepared by grafting non-immune mouse IgG onto the liposomal surface. The influence of active targeted immunoliposomes on the type and intensity of generated immune response against Leishmania was then investigated and compared with that of liposomes and control groups which received either SLA or HEPES buffer alone. All formulations contained SLA and were used to immunize the mice in the left hind footpad three times in 3-week intervals. Evaluation of lesion development and parasite burden in the foot and spleen after challenge with Leishmania major, evaluation of Th1 cytokine (IFN-γ), and titration of IgG isotypes were carried out to assess the type of generated immune response and the extent of protection. The results indicated that liposomes might be effective adjuvant systems to induce protection against L. major challenge in BALB/c mice, but stronger cell mediated immune responses were induced when immunoliposomes were utilized. Thus, immune modulation using immunoliposomes might be a practical approach to improve the immunization against L. major. Copyright © 2014 Elsevier Inc. All rights reserved.

Species-Specific Antimonial Sensitivity in Leishmania Is Driven by Post-Transcriptional Regulation of AQP1

PubMed Central

Mandal, Goutam; Mandal, Srotoswati; Sharma, Mansi; Charret, Karen Santos; Papadopoulou, Barbara; Bhattacharjee, Hiranmoy; Mukhopadhyay, Rita

2015-01-01

Leishmania is a digenetic protozoan parasite causing leishmaniasis in humans. The different clinical forms of leishmaniasis are caused by more than twenty species of Leishmania that are transmitted by nearly thirty species of phlebotomine sand flies. Pentavalent antimonials (such as Pentostam or Glucantime) are the first line drugs for treating leishmaniasis. Recent studies suggest that pentavalent antimony (Sb(V)) acts as a pro-drug, which is converted to the more active trivalent form (Sb(III)). However, sensitivity to trivalent antimony varies among different Leishmania species. In general, Leishmania species causing cutaneous leishmaniasis (CL) are more sensitive to Sb(III) than the species responsible for visceral leishmaniasis (VL). Leishmania aquaglyceroporin (AQP1) facilitates the adventitious passage of antimonite down a concentration gradient. In this study, we show that Leishmania species causing CL accumulate more antimonite, and therefore exhibit higher sensitivity to antimonials, than the species responsible for VL. This species-specific differential sensitivity to antimonite is directly proportional to the expression levels of AQP1 mRNA. We show that the stability of AQP1 mRNA in different Leishmania species is regulated by their respective 3’-untranslated regions. The differential regulation of AQP1 mRNA explains the distinct antimonial sensitivity of each species. PMID:25714343

Leishmaniasis in Texas: Isolation of Leishmania mexicana from Neotoma Micropus

DTIC Science & Technology

1990-01-01

leishmaniasis cases in Texas, 2) Lutzomyia anthophora, a sand fly which has transmitted Leishmania mexicana under laboratory conditions (Endris et al., 1984...Addis, 1945). Other vertebrates such as opossums, hispid cotton rats, and armadillos and other sand flies such as Lutzomyia diabolica and Lutzomyia texana...Leishmania mexicana by a North American sand fly, Lutzomyia anthophora (Diptera: Psychodidae). Journal of Medical Entomology 24: 243- 247. GRIMALDI, G. Jr

Natural infection of Didelphis aurita (Mammalia: Marsupialia) with Leishmania infantum in Brazil

PubMed Central

2012-01-01

Background The opossum Didelphis have been considered as natural hosts of Leishmania parasites in the New World, suggesting an important role in the epidemiology of Visceral Leishmaniasis (VL). Among six extant species that belong to the genus Didelphis, only two (D. marsupialis and D. albiventris), have been mentioned as natural hosts of Leishmania infantum in Brazil and Colombia. In the present paper, it is reported for the first time, the observation of intracellular parasites (amastigotes) in tissues of Didelphis aurita naturally infected with Leishmania infantum in Brazil. We also discuss some aspects associated to the relationship between L. infantum and the geographical distribution of some species of the genus Didelphis. Methods The opossums studied were caught by wire traps (Tomahawk) in Barra de Guaratiba, a peri-urban area in Rio de Janeiro. The opossums were killed with an overdose of Thiopental sodium.At necropsy, macroscopic alterations were examined and samples from liver, spleen, lymph nodes, ear, abdominal skin, scent glands and bone marrow were collected for parasitological and molecular diagnoses. Results Forty-eight opossums were captured in an AVL endemic region, 30 being caught in a mangrove area and eighteen animals in a forest area near to some residential-yards. Among the thirty opossums trapped in the mangrove area, all of them were negative by both imprint and sera samples assayed on Dipstick Tests, that is a test based on a combination of protein-A colloidal gold conjugate and rk39 Leishmania antigen to detect anti-Leishmania antibody in serum or plasma. At the macroscopic examination one out of eighteen opossums, caught close to the forest, presented alterations compatible with spleen hypertrophy and three were positive by Dipstick Tests (16.6%) and presented amastigotes in the spleen and in one of them, the parasites were also observed in a submandibular lymph node. Leishmania infantum infections were confirmed through dot blot

Natural infection of Didelphis aurita (Mammalia: Marsupialia) with Leishmania infantum in Brazil.

PubMed

Carreira, João Carlos Araujo; da Silva, Alba Valéria Machado; de Pita Pereira, Daniela; Brazil, Reginaldo Peçanha

2012-06-07

The opossum Didelphis have been considered as natural hosts of Leishmania parasites in the New World, suggesting an important role in the epidemiology of Visceral Leishmaniasis (VL). Among six extant species that belong to the genus Didelphis, only two (D. marsupialis and D. albiventris), have been mentioned as natural hosts of Leishmania infantum in Brazil and Colombia. In the present paper, it is reported for the first time, the observation of intracellular parasites (amastigotes) in tissues of Didelphis aurita naturally infected with Leishmania infantum in Brazil. We also discuss some aspects associated to the relationship between L. infantum and the geographical distribution of some species of the genus Didelphis. The opossums studied were caught by wire traps (Tomahawk) in Barra de Guaratiba, a peri-urban area in Rio de Janeiro. The opossums were killed with an overdose of Thiopental sodium.At necropsy, macroscopic alterations were examined and samples from liver, spleen, lymph nodes, ear, abdominal skin, scent glands and bone marrow were collected for parasitological and molecular diagnoses. Forty-eight opossums were captured in an AVL endemic region, 30 being caught in a mangrove area and eighteen animals in a forest area near to some residential-yards. Among the thirty opossums trapped in the mangrove area, all of them were negative by both imprint and sera samples assayed on Dipstick Tests, that is a test based on a combination of protein-A colloidal gold conjugate and rk39 Leishmania antigen to detect anti-Leishmania antibody in serum or plasma. At the macroscopic examination one out of eighteen opossums, caught close to the forest, presented alterations compatible with spleen hypertrophy and three were positive by Dipstick Tests (16.6%) and presented amastigotes in the spleen and in one of them, the parasites were also observed in a submandibular lymph node. Leishmania infantum infections were confirmed through dot blot hybridization using a L. infantum

Preparation of highly infective Leishmania promastigotes by cultivation on SNB-9 biphasic medium.

PubMed

Grekov, Igor; Svobodová, Milena; Nohýnková, Eva; Lipoldová, Marie

2011-12-01

Protozoan hemoflagellates Leishmania are causative agents of leishmaniases and an important biological model for study of host-pathogen interaction. A wide range of methods of Leishmania cultivation on both biphasic and liquid media is available. Biphasic media are considered to be superior for initial isolation of the parasites and obtaining high promastigote infectivity; however, liquid media are more suitable for large-scale experiments. The aim of the present study was the adaptation and optimization of the cultivation of Leishmania promastigotes on a biphasic SNB-9 (saline-neopeptone-blood 9) medium that was originally developed for Trypanosoma cultivation and combines the advantages of biphasic and liquid media. SNB-9 medium is characterized with a large volume of the liquid phase, which facilitates the manipulation with the culture and provides parasite yields comparable to parasite yields on such liquid medium as Schneider's Insect Medium. We demonstrate that SNB-9 very considerably surpasses Schneider's Insect Medium in in vitro infectivity of the parasites. Additionally, we show that the ratio of apoptotic parasites, which are important for the infectivity of the inoculum, in Leishmania culture in SNB-9 is higher than in Leishmania culture in Schneider's Insect Medium. Thus, we demonstrate that the cultivation of Leishmania on SNB-9 reliably yields highly infective promastigotes suitable for experimental infection. Copyright © 2011 Elsevier B.V. All rights reserved.

Comparative cost effectiveness of varicella, hepatitis A, and pneumococcal conjugate vaccines.

PubMed

Jacobs, R J; Meyerhoff, A S

2001-12-01

Several state and local U.S. governments are considering making varicella, hepatitis A, and/or pneumococcal conjugate vaccination conditions of day care or school entry. These requirements will likely be issued sequentially, because simultaneous mandates exacerbate budget constraints and complicate communication with parents and providers. Cost-effectiveness assessments should aid the establishment of vaccination priorities, but comparing results of published studies is confounded by their dissimilar methods. We reviewed U.S. cost-effectiveness studies of childhood varicella, hepatitis A, and pneumococcal conjugate vaccines and identified four providing data required to standardize methods. Vaccination, disease treatment, and work-loss costs were estimated from original study results and current prices. Estimated life-years saved were derived from original study results, epidemiological evidence, and alternative procedures for discounting to present values. Hepatitis A vaccine would have the lowest health system costs per life-year saved. Varicella vaccine would provide the greatest reduction in societal costs, mainly through reduced parent work loss. Pneumococcal conjugate vaccine would cost twice the amount of varicella and hepatitis A vaccines combined and be less cost effective than the other vaccines. Hepatitis A and varicella vaccines, but not pneumococcal conjugate vaccine, meet or exceed conventional standards of cost effectiveness. Copyright 2001 American Health Foundation and Elsevier Science.

Serological Evidence of Infection by Leishmania (Leishmania) infantum (Synonym: Leishmania (Leishmania) chagasi) in Free-Ranging Wild Mammals in a Nonendemic Region of the State of São Paulo, Brazil.

PubMed

Paiz, Laís Moraes; Fornazari, Felipe; Menozzi, Benedito Donizete; Oliveira, Gabriela Capriogli; Coiro, Carla Janeiro; Teixeira, Carlos Roberto; da Silva, Valdinei Moraes Campanucci; Donalisio, Maria Rita; Langoni, Helio

2015-11-01

Concerns about the interface between wildlife, domestic animals, and humans in the transmission of visceral leishmaniasis (VL) have been growing due to natural or anthropogenic environmental changes. In this context, investigations of the infection in wild mammals are important to assess their exposure to the vector and the parasite. A study of anti-Leishmania (Leishmania) infantum antibodies was carried out using the direct agglutination test (DAT) on 528 free-ranging wild mammals of 38 species from the region of Botucatu, state of São Paulo, Brazil, a municipality that has no records of the vector or of human or canine autochthony. Antibodies were detected, with a cutoff of 1:320, in 9/528 (1.7%; 95% confidence interval [CI] 0.6-2.8%) mammals of the species Callithrix jacchus, Lepus europaeus, Sphiggurus villosus, Nasua nasua, Eira barbara, and Galictis cuja, with high titers (≥1280) for the last three. These three are little-studied species, and previous records of the detection of anti-Leishmania spp. antibodies in Brazil exist only for coatis (N. nasua), whereas worldwide, infection by L. (L.) infantum has been confirmed only in hares (Le. europaeus). On the other hand, opossums and canids, the species most commonly reported to be naturally infected by L. (L.) infantum, were not seropositive. Fifty-eight (58/528; 10.9%) mammals were found to have antibody titers ranging from 20 to 160 and were not included among the seropositive animals due to the adopted cutoff. However, the possibility of infection in these animals should not be discarded, because there is no standard cutoff point for the different wild species. Our findings indicate the need for investigations into the exact role of the seropositive species in the epidemiology of VL and for effective epidemiological surveillance to prevent its expansion, because even in regions where there are no records of canine or human autochthonous cases, there may be parasite circulation among wild mammals.

Structurally optimized analogs of the retrograde trafficking inhibitor Retro-2cycl limit Leishmania infections.

PubMed

Craig, Evan; Huyghues-Despointes, Charles-Eugene; Yu, Chun; Handy, Emma L; Sello, Jason K; Kima, Peter E

2017-05-01

In infected mammalian cells, Leishmania parasites reside within specialized compartments called parasitophorous vacuoles (LPVs). We have previously shown that Retro-2, a member of a novel class of small retrograde pathway inhibitors caused reduced LPV sizes and lower parasite numbers during experimental L. mexicana sp. infections. The purpose of this study was to determine if structural analogs of Retro-2cycl reported to have superior potency in the inhibition of retrograde pathway-dependent phenomena (i.e., polyomavirus cellular infection by polyomavrius and Shiga toxin trafficking in cells) are also more effective than the parent compound at controlling Leishmania infections. In addition to their effects on LPV development, we show that two optimized analogs of Retro-2cycl, DHQZ 36 and DHQZ 36.1 limit Leishmania amazonensis infection in macrophages at EC50 of 13.63+/-2.58μM and10.57+/-2.66μM, respectively, which is significantly lower than 40.15μM the EC50 of Retro-2cycl. In addition, these analogs caused a reversal in Leishmania induced suppression of IL-6 release by infected cells after LPS activation. Moreover, we show that in contrast to Retro-2cycl that is Leishmania static, the analogs can kill Leishmania parasites in axenic cultures, which is a desirable attribute for any drug to treat Leishmania infections. Together, these studies validate and extend the published structure-activity relationship analyses of Retro-2cycl.

First report of naturally infected Sergentomyia minuta with Leishmania major in Tunisia.

PubMed

Jaouadi, Kaouther; Ghawar, Wissem; Salem, Sadok; Gharbi, Mohamed; Bettaieb, Jihene; Yazidi, Rihab; Harrabi, Mariem; Hamarsheh, Omar; Ben Salah, Afif

2015-12-21

Many sand fly species are implicated in the transmission cycle of Leishmania parasites around the world. Incriminating new sand flies species, as vectors of Leishmania is crucial to understanding the parasite-vector transmission cycle in different areas in Tunisia and surrounding countries. Seventy-four unfed females belonging to the genera Sergentomyia and Phlebotomus were collected in South Tunisia between June and November 2014, using sticky papers. PCR-RFLP (Restriction Fragment Length Polymorphism) analysis of the internal transcribed spacer 1 (ITS1) was used for Leishmania parasites detection and identification. Leishmania (L.) major (Yakimoff & Shokkor, 1914) was identified within two Sergentomyia (S.) minuta (Rondani, 1843) and one Phlebotomus papatasi (Scopoli, 1786). This is the first report of L. major identified from S. minuta in Tunisia. This novel finding enhances the understanding of the transmission cycle of L. major parasites of cutaneous leishmaniasis in an endemic area in South Tunisia.

The flagellar protein FLAG1/SMP1 is a candidate for Leishmania-sand fly interaction.

PubMed

Di-Blasi, Tatiana; Lobo, Amanda R; Nascimento, Luanda M; Córdova-Rojas, Jose L; Pestana, Karen; Marín-Villa, Marcel; Tempone, Antonio J; Telleria, Erich L; Ramalho-Ortigão, Marcelo; McMahon-Pratt, Diane; Traub-Csekö, Yara M

2015-03-01

Leishmaniasis is a serious problem that affects mostly poor countries. Various species of Leishmania are the agents of the disease, which take different clinical manifestations. The parasite is transmitted by sandflies, predominantly from the Phlebotomus genus in the Old World and Lutzomyia in the New World. During development in the gut, Leishmania must survive various challenges, which include avoiding being expelled with blood remnants after digestion. It is believed that attachment to the gut epithelium is a necessary step for vector infection, and molecules from parasites and sand flies have been implicated in this attachment. In previous work, monoclonal antibodies were produced against Leishmania. Among these an antibody was obtained against Leishmania braziliensis flagella, which blocked the attachment of Leishmania panamensis flagella to Phlebotomus papatasi guts. The protein recognized by this antibody was identified and named FLAG1, and the complete FLAG1 gene sequence was obtained. This protein was later independently identified as a small, myristoylated protein and called SMP1, so from now on it will be denominated FLAG1/SMP1. The FLAG1/SMP1 gene is expressed in all developmental stages of the parasite, but has higher expression in promastigotes. The anti-FLAG1/SMP1 antibody recognized the flagellum of all Leishmania species tested and generated the expected band by western blots. This antibody was used in attachment and infection blocking experiments. Using the New World vector Lutzomyia longipalpis and Leishmania infantum chagasi, no inhibition of attachment ex vivo or infection in vivo was seen. On the other hand, when the Old World vectors P. papatasi and Leishmania major were used, a significant decrease of both attachment and infection were seen in the presence of the antibody. We propose that FLAG1/SMP1 is involved in the attachment/infection of Leishmania in the strict vector P. papatasi and not the permissive vector L. longipalpis.

Field randomized trial to evaluate the efficacy of the Leish-Tec® vaccine against canine visceral leishmaniasis in an endemic area of Brazil.

PubMed

Regina-Silva, Shara; Feres, Ana Maria Leonardi Tibúrcio; França-Silva, João Carlos; Dias, Edelberto Santos; Michalsky, Érika Monteiro; de Andrade, Hélida Monteiro; Coelho, Eduardo Antonio Ferraz; Ribeiro, Gustavo Meirelles; Fernandes, Ana Paula; Machado-Coelho, George Luiz Lins

2016-04-27

A canine vaccine remains a promising approach for effective control of visceral leishmaniasis (VL), given its complex epidemiology in areas where zoonotic VL is prevalent. Leish-Tec(®) is a recombinant vaccine, based on the Leishmania A2 antigen, against canine VL (CVL). It is, since 2014, the single commercial vaccine licensed in Brazil. Here, Leish-Tec(®) efficacy was estimated through a randomized field trial (RFT), in a highly VL endemic area. The RFT was conducted from 2008 to 2010 in an endemic area of southeastern Brazil, presenting a CVL seroprevalence of 41.9%. Eight hundred forty-seven seronegative dogs were randomly selected to receive Leish-Tec(®) (n=429) or placebo (n=418). Animals were followed up by clinical, serological, and parasitological exams for 18 months. The CVL incidence in both groups was compared through proportion analysis. A significant reduction in the number of cases of CVL was observed in the vaccine group, as compared with the placebo group, whether efficacy was estimated according to parasitological results (71.4%; 95% CI: 34.9-87.3%; p=0.001; risk ratio=0.287), by adding results of xenodiagnosis and parasitological exams (58.1%; 95% CI: 26.0-76.3%; p=0.002; risk ratio=0.419). Among the animals that converted to a positive anti-A2 serology, efficacy reached 80.8% (95% CI: 37.6-94.1%, p=0.001; risk ratio=0.192). Xenodiagnosis has detected a reduction of 46.6% (p=0.05) in transmission to sand flies from vaccinated animals presenting anti-A2 positive serology. The Leish-Tec(®) vaccine proved significantly effective for prophylaxis of CVL, after natural challenge assured by transmission of Leishmania parasites, in a highly endemic area. Noteworthy, this report has unveiled the complexity of performing a RFT for anti-CVL vaccines in Brazil, which may be helpful for designing of future studies. Copyright © 2016 Elsevier Ltd. All rights reserved.

A third generation vaccine for human visceral leishmaniasis and post kala azar dermal leishmaniasis: First-in-human trial of ChAd63-KH

PubMed Central

Osman, Mohamed; Mistry, Anoop; Keding, Ada; Cook, Elizabeth; Wiggins, Rebecca; Di Marco, Stefania; Colloca, Stefano; Siani, Loredana; Smith, Deborah F.; Aebischer, Toni; Lacey, Charles J.

2017-01-01

Background Visceral leishmaniasis (VL or kala azar) is the most serious form of human leishmaniasis, responsible for over 20,000 deaths annually, and post kala azar dermal leishmaniasis (PKDL) is a stigmatizing skin condition that often occurs in patients after successful treatment for VL. Lack of effective or appropriately targeted cell mediated immunity, including CD8+ T cell responses, underlies the progression of VL and progression to PKDL, and can limit the therapeutic efficacy of anti-leishmanial drugs. Hence, in addition to the need for prophylactic vaccines against leishmaniasis, the development of therapeutic vaccines for use alone or in combined immuno-chemotherapy has been identified as an unmet clinical need. Here, we report the first clinical trial of a third-generation leishmaniasis vaccine, developed intentionally to induce Leishmania-specific CD8+ T cells. Methods We conducted a first-in-human dose escalation Phase I trial in 20 healthy volunteers to assess the safety, tolerability and immunogenicity of a prime-only adenoviral vaccine for human VL and PKDL. ChAd63-KH is a replication defective simian adenovirus expressing a novel synthetic gene (KH) encoding two Leishmania proteins KMP-11 and HASPB. Uniquely, the latter was engineered to reflect repeat domain polymorphisms and arrangements identified from clinical isolates. We monitored innate immune responses by whole blood RNA-Seq and antigen specific CD8+ T cell responses by IFNγ ELISPOT and intracellular flow cytometry. Findings ChAd63-KH was safe at intramuscular doses of 1x1010 and 7.5x1010 vp. Whole blood transcriptomic profiling indicated that ChAd63-KH induced innate immune responses characterized by an interferon signature and the presence of activated dendritic cells. Broad and quantitatively robust CD8+ T cell responses were induced by vaccination in 100% (20/20) of vaccinated subjects. Conclusion The results of this study support the further development of ChAd63-KH as a novel third

A third generation vaccine for human visceral leishmaniasis and post kala azar dermal leishmaniasis: First-in-human trial of ChAd63-KH.

PubMed

Osman, Mohamed; Mistry, Anoop; Keding, Ada; Gabe, Rhian; Cook, Elizabeth; Forrester, Sarah; Wiggins, Rebecca; Di Marco, Stefania; Colloca, Stefano; Siani, Loredana; Cortese, Riccardo; Smith, Deborah F; Aebischer, Toni; Kaye, Paul M; Lacey, Charles J

2017-05-01

Visceral leishmaniasis (VL or kala azar) is the most serious form of human leishmaniasis, responsible for over 20,000 deaths annually, and post kala azar dermal leishmaniasis (PKDL) is a stigmatizing skin condition that often occurs in patients after successful treatment for VL. Lack of effective or appropriately targeted cell mediated immunity, including CD8+ T cell responses, underlies the progression of VL and progression to PKDL, and can limit the therapeutic efficacy of anti-leishmanial drugs. Hence, in addition to the need for prophylactic vaccines against leishmaniasis, the development of therapeutic vaccines for use alone or in combined immuno-chemotherapy has been identified as an unmet clinical need. Here, we report the first clinical trial of a third-generation leishmaniasis vaccine, developed intentionally to induce Leishmania-specific CD8+ T cells. We conducted a first-in-human dose escalation Phase I trial in 20 healthy volunteers to assess the safety, tolerability and immunogenicity of a prime-only adenoviral vaccine for human VL and PKDL. ChAd63-KH is a replication defective simian adenovirus expressing a novel synthetic gene (KH) encoding two Leishmania proteins KMP-11 and HASPB. Uniquely, the latter was engineered to reflect repeat domain polymorphisms and arrangements identified from clinical isolates. We monitored innate immune responses by whole blood RNA-Seq and antigen specific CD8+ T cell responses by IFNγ ELISPOT and intracellular flow cytometry. ChAd63-KH was safe at intramuscular doses of 1x1010 and 7.5x1010 vp. Whole blood transcriptomic profiling indicated that ChAd63-KH induced innate immune responses characterized by an interferon signature and the presence of activated dendritic cells. Broad and quantitatively robust CD8+ T cell responses were induced by vaccination in 100% (20/20) of vaccinated subjects. The results of this study support the further development of ChAd63-KH as a novel third generation vaccine for VL and PKDL. This

Evaluation of four molecular methods to detect Leishmania infection in dogs.

PubMed

Albuquerque, Andreia; Campino, Lenea; Cardoso, Luís; Cortes, Sofia

2017-03-13

Canine leishmaniasis, a zoonotic disease caused by Leishmania infantum vectored by phlebotomine sand flies, is considered a relevant veterinary and public health problem in various countries, namely in the Mediterranean basin and Brazil, where dogs are considered the main reservoir hosts. Not only diseased dogs but also those subclinically infected play a relevant role in the transmission of L. infantum to vectors; therefore, early diagnosis is essential, under both a clinical and an epidemiological perspective. Molecular tools can be a more accurate and sensitive approach for diagnosis, with a wide range of protocols currently in use. The aim of the present report was to compare four PCR based protocols for the diagnosis of canine Leishmania infection in a cohort of dogs from the Douro region, Portugal. A total of 229 bone marrow samples were collected from dogs living in the Douro region, an endemic region for leishmaniasis. Four PCR protocols were evaluated for Leishmania DNA detection in canine samples, three single (ITS1-PCR, MC-PCR and Uni21/Lmj4-PCR) and one nested (nested SSU rRNA-PCR). Two of the protocols were based on nuclear targets and the other two on kinetoplastid targets. The higher overall percentage of infected dogs was detected with the nested SSU rRNA-PCR (37.6%), which also was able to detect Leishmania DNA in a higher number of samples from apparently healthy dogs (25.3%). The ITS1-PCR presented the lowest level of Leishmania detection. Nested SSU rRNA-PCR is an appropriate method to detect Leishmania infection in dogs. Accurate and early diagnosis in clinically suspect as well as apparently healthy dogs is essential, in order to treat and protect animals and public health and contribute to the control and awareness of the disease.

Leishmania isoenzyme polymorphisms in Ecuador: Relationships with geographic distribution and clinical presentation

PubMed Central

Calvopina, Manuel; Armijos, Rodrigo X; Marco, Jorge D; Uezato, Hiroshi; Kato, Hirotomo; Gomez, Eduardo A; Korenaga, Masataka; Barroso, Paola A; Mimori, Tatsuyuki; Cooper, Philip J; Nonaka, Shigeo; Hashiguchi, Yoshihisa

2006-01-01

Background Determinants of the clinical presentation of the leishmaniases are poorly understood but Leishmania species and strain differences are important. To examine the relationship between clinical presentation, species and isoenzyme polymorphisms, 56 Leishmania isolates from distinct presentations of American tegumentary leishmaniasis (ATL) from Ecuador were analyzed. Methods Isolates were characterized by multilocus enzyme electrophoresis for polymorphisms of 11 isoenzymes. Patients were infected in four different ecologic regions: highland and lowland jungle of the Pacific coast, Amazonian lowlands and Andean highlands. Results Six Leishmania species constituting 21 zymodemes were identified: L. (Viannia) panamensis (21 isolates, 7 zymodemes), L. (V.) guyanensis (7 isolates, 4 zymodemes), L. (V.) braziliensis (5 isolates, 3 zymodemes), L. (Leishmania) mexicana (11 isolates, 4 zymodemes), L. (L.) amazonensis (10 isolates, 2 zymodemes) and L. (L.) major (2 isolates, 1 zymodeme). L. panamensis was the species most frequently identified in the Pacific region and was associated with several clinical variants of cutaneous disease (CL); eight cases of leishmaniasis recidiva cutis (LRC) found in the Pacific highlands were associated with 3 zymodemes of this species. Mucocutaneous leishmaniasis found only in the Amazonian focus was associated with 3 zymodemes of L. braziliensis. The papular variant of CL, Uta, found in the Andean highlands was related predominantly with a single zymodeme of L. mexicana. Conclusion Our data show a high degree of phenotypic variation within species, and some evidence for associations between specific variants of ATL (i.e. Uta and LRC) and specific Leishmania zymodemes. This study further defines the geographic distribution of Leishmania species and clinical variants of ATL in Ecuador. PMID:16968553

Evaluation of Leishmania species reactivity in human serologic diagnosis of leishmaniasis.

PubMed

Silvestre, Ricardo; Santarém, Nuno; Teixeira, Lúcia; Cunha, Joana; Schallig, Henk; Cordeiro-da-Silva, Anabela

2009-08-01

The sensitivities and specificities of IgG-ELISA and IgG flow cytometry based techniques using different Leishmania species were determined using sera collected from 40 cutaneous or visceral leishmaniasis patients. The flow cytometry technique, using promastigote parasite forms, performed better than total soluble extract IgG-ELISA. At the species level, the use of Leishmania amazonensis and Leishmania major as antigens in enzyme linked immunosorbent assay (ELISA) decreased the overall sensitivity. To assess the specificity of these tests, sera from malaria, toxoplasmosis, amoebiasis, schistosomiasis, and leprosy patients were used. We also included sera from Leishmania non-infected endemic individuals. The cutaneous species displayed a decreased specificity in both assays. Although more sensitive, flow cytometry using promastigote parasite forms generally presented lower levels of specificity when compared with total extract of IgG-ELISA. Overall, the results of the study show the potential of IgG flow cytometry for the diagnosis of leishmaniasis. Although highly sensitive, a refinement of the flow cytometry method should be performed to improve the overall specificity.

Statin-induced chronic cholesterol depletion inhibits Leishmania donovani infection: Relevance of optimum host membrane cholesterol.

PubMed

Kumar, G Aditya; Roy, Saptarshi; Jafurulla, Md; Mandal, Chitra; Chattopadhyay, Amitabha

2016-09-01

Leishmania are obligate intracellular protozoan parasites that invade and survive within host macrophages leading to leishmaniasis, a major cause of mortality and morbidity worldwide, particularly among economically weaker sections in tropical and subtropical regions. Visceral leishmaniasis is a potent disease caused by Leishmania donovani. The detailed mechanism of internalization of Leishmania is poorly understood. A basic step in the entry of Leishmania involves interaction of the parasite with the host plasma membrane. In this work, we have explored the effect of chronic metabolic cholesterol depletion using lovastatin on the entry and survival of Leishmania donovani in host macrophages. We show here that chronic cholesterol depletion of host macrophages results in reduction in the attachment of Leishmania promastigotes, along with a concomitant reduction in the intracellular amastigote load. These results assume further relevance since chronic cholesterol depletion is believed to mimic physiological cholesterol modulation. Interestingly, the reduction in the ability of Leishmania to enter host macrophages could be reversed upon metabolic replenishment of cholesterol. Importantly, enrichment of host membrane cholesterol resulted in reduction in the entry and survival of Leishmania in host macrophages. As a control, the binding of Escherichia coli to host macrophages remained invariant under these conditions, thereby implying specificity of cholesterol requirement for effective leishmanial infection. To the best of our knowledge, these results constitute the first comprehensive demonstration that an optimum content of host membrane cholesterol is necessary for leishmanial infection. Our results assume relevance in the context of developing novel therapeutic strategies targeting cholesterol-mediated leishmanial infection. Copyright © 2016 Elsevier B.V. All rights reserved.

The characterization by isoenzyme electrophoresis of Leishmania isolated in the People's Republic of China.

PubMed

Xu, Z B; Le Blancq, S; Evans, D A; Peters, W

1984-01-01

Seven isolates of Leishmania from mainland China were characterized on the basis of their isoenzyme profiles for 10 enzymes. Five isolates were from human visceral leishmaniasis patients, and four of these showed isoenzyme patterns similar to the marker strain of Leishmania infantum, while one was similar to L. donovani sensu lato. One isolate was from a presumed reservoir host of human visceral leishmaniasis, the racoon dog Nyctereutes procyanoides, and was isoenzymically indistinguishable from L. infantum. An isolate of L. gerbilli from the great gerbil Rhombomys opimus was readily distinguishable from Old World marker strains and other Chinese leishmanias. This is the first report of the biochemical characterization of Chinese isolates of Leishmania.

Impact of Leishmania metalloprotease GP63 on macrophage signaling

PubMed Central

Isnard, Amandine; Shio, Marina T.; Olivier, Martin

2012-01-01

The intramacrophage protozoan parasites of Leishmania genus have developed sophisticated ways to subvert the innate immune response permitting their infection and propagation within the macrophages of the mammalian host. Several Leishmania virulence factors have been identified and found to be of importance for the development of leishmaniasis. However, recent findings are now further reinforcing the critical role played by the zinc-metalloprotease GP63 as a virulence factor that greatly influence host cell signaling mechanisms and related functions. GP63 has been found to be involved not only in the cleavage and degradation of various kinases and transcription factors, but also to be the major molecule modulating host negative regulatory mechanisms involving for instance protein tyrosine phosphatases (PTPs). Those latter being well recognized for their pivotal role in the regulation of a great number of signaling pathways. In this review article, we are providing a complete overview about the role of Leishmania GP63 in the mechanisms underlying the subversion of macrophage signaling and functions. PMID:22919663

Impact of Leishmania metalloprotease GP63 on macrophage signaling.

PubMed

Isnard, Amandine; Shio, Marina T; Olivier, Martin

2012-01-01

The intramacrophage protozoan parasites of Leishmania genus have developed sophisticated ways to subvert the innate immune response permitting their infection and propagation within the macrophages of the mammalian host. Several Leishmania virulence factors have been identified and found to be of importance for the development of leishmaniasis. However, recent findings are now further reinforcing the critical role played by the zinc-metalloprotease GP63 as a virulence factor that greatly influence host cell signaling mechanisms and related functions. GP63 has been found to be involved not only in the cleavage and degradation of various kinases and transcription factors, but also to be the major molecule modulating host negative regulatory mechanisms involving for instance protein tyrosine phosphatases (PTPs). Those latter being well recognized for their pivotal role in the regulation of a great number of signaling pathways. In this review article, we are providing a complete overview about the role of Leishmania GP63 in the mechanisms underlying the subversion of macrophage signaling and functions.

A Poly(Lactic-co-Glycolic) Acid Nanovaccine Based on Chimeric Peptides from Different Leishmania infantum Proteins Induces Dendritic Cells Maturation and Promotes Peptide-Specific IFNγ-Producing CD8+ T Cells Essential for the Protection against Experimental Visceral Leishmaniasis.

PubMed

Athanasiou, Evita; Agallou, Maria; Tastsoglou, Spyros; Kammona, Olga; Hatzigeorgiou, Artemis; Kiparissides, Costas; Karagouni, Evdokia

2017-01-01

Visceral leishmaniasis, caused by Leishmania ( L .) donovani and L. infantum protozoan parasites, can provoke overwhelming and protracted epidemics, with high case-fatality rates. An effective vaccine against the disease must rely on the generation of a strong and long-lasting T cell immunity, mediated by CD4 + T H1 and CD8 + T cells. Multi-epitope peptide-based vaccine development is manifesting as the new era of vaccination strategies against Leishmania infection. In this study, we designed chimeric peptides containing HLA-restricted epitopes from three immunogenic L. infantum proteins (cysteine peptidase A, histone H1, and kinetoplastid membrane protein 11), in order to be encapsulated in poly(lactic- co -glycolic) acid nanoparticles with or without the adjuvant monophosphoryl lipid A (MPLA) or surface modification with an octapeptide targeting the tumor necrosis factor receptor II. We aimed to construct differentially functionalized peptide-based nanovaccine candidates and investigate their capacity to stimulate the immunomodulatory properties of dendritic cells (DCs), which are critical regulators of adaptive immunity generated upon vaccination. According to our results, DCs stimulation with the peptide-based nanovaccine candidates with MPLA incorporation or surface modification induced an enhanced maturation profile with prominent IL-12 production, promoting allogeneic T cell proliferation and intracellular production of IFNγ by CD4 + and CD8 + T cell subsets. In addition, DCs stimulated with the peptide-based nanovaccine candidate with MPLA incorporation exhibited a robust transcriptional activation, characterized by upregulated genes indicative of vaccine-driven DCs differentiation toward type 1 phenotype. Immunization of HLA A2.1 transgenic mice with this peptide-based nanovaccine candidate induced peptide-specific IFNγ-producing CD8 + T cells and conferred significant protection against L. infantum infection. Concluding, our findings supported that

A Poly(Lactic-co-Glycolic) Acid Nanovaccine Based on Chimeric Peptides from Different Leishmania infantum Proteins Induces Dendritic Cells Maturation and Promotes Peptide-Specific IFNγ-Producing CD8+ T Cells Essential for the Protection against Experimental Visceral Leishmaniasis

PubMed Central

Athanasiou, Evita; Agallou, Maria; Tastsoglou, Spyros; Kammona, Olga; Hatzigeorgiou, Artemis; Kiparissides, Costas; Karagouni, Evdokia

2017-01-01

Visceral leishmaniasis, caused by Leishmania (L.) donovani and L. infantum protozoan parasites, can provoke overwhelming and protracted epidemics, with high case-fatality rates. An effective vaccine against the disease must rely on the generation of a strong and long-lasting T cell immunity, mediated by CD4+ TH1 and CD8+ T cells. Multi-epitope peptide-based vaccine development is manifesting as the new era of vaccination strategies against Leishmania infection. In this study, we designed chimeric peptides containing HLA-restricted epitopes from three immunogenic L. infantum proteins (cysteine peptidase A, histone H1, and kinetoplastid membrane protein 11), in order to be encapsulated in poly(lactic-co-glycolic) acid nanoparticles with or without the adjuvant monophosphoryl lipid A (MPLA) or surface modification with an octapeptide targeting the tumor necrosis factor receptor II. We aimed to construct differentially functionalized peptide-based nanovaccine candidates and investigate their capacity to stimulate the immunomodulatory properties of dendritic cells (DCs), which are critical regulators of adaptive immunity generated upon vaccination. According to our results, DCs stimulation with the peptide-based nanovaccine candidates with MPLA incorporation or surface modification induced an enhanced maturation profile with prominent IL-12 production, promoting allogeneic T cell proliferation and intracellular production of IFNγ by CD4+ and CD8+ T cell subsets. In addition, DCs stimulated with the peptide-based nanovaccine candidate with MPLA incorporation exhibited a robust transcriptional activation, characterized by upregulated genes indicative of vaccine-driven DCs differentiation toward type 1 phenotype. Immunization of HLA A2.1 transgenic mice with this peptide-based nanovaccine candidate induced peptide-specific IFNγ-producing CD8+ T cells and conferred significant protection against L. infantum infection. Concluding, our findings supported that encapsulation

Studies on cocktails of 31-kDa, 36-kDa and 51-kDa antigens of Leishmania donovani along with saponin against murine visceral leishmaniasis.

PubMed

Kaur, H; Thakur, A; Kaur, S

2015-04-01

A substantial number of antigens of Leishmania donovani have been described in the past. However, identifying candidate antigens is not enough. Appropriate antigen delivery to induce the right type of immune response against leishmaniasis (i.e. induction of a strong antigen-specific Th1 type of immune response) is another crucial component of an effective vaccine. Therefore, 'cocktail' vaccines are proposed based on the assumption that such cocktails will show enhanced efficacy. Studies have been carried out on LD31 and LD51 polypeptides from L. donovani promastigotes, which have proven to be potential vaccine candidates. This study was designed to check the protective efficacy of various cocktails of low molecular weight antigens alone and along with saponin as adjuvant. Mice were sacrificed on different post-challenge days for evaluation of parasite load and other immunological parameters. Protective efficacy of different vaccine formulations was revealed by significant decline in parasite burden and increased DTH Delayed Type Hypersenstivity responses. The antibody response was of IgG type with elevated IgG2a and decreased production of IgG1, whereas cytokine levels pointed towards the generation of protective Th1 type of immune response. Among all vaccine formulations, cocktail of 31+51+saponin was found to be highly immunogenic and imparted maximum protection. © 2015 John Wiley & Sons Ltd.

Multifaceted Population Structure and Reproductive Strategy in Leishmania donovani Complex in One Sudanese Village

PubMed Central

Hide, Mallorie; Le Falher, Georges; Bucheton, Bruno; Dereure, Jacques; El-Safi, Sayda H.; Dessein, Alain; Bañuls, Anne-Laure

2011-01-01

Leishmania species of the subgenus Leishmania and especially L. donovani are responsible for a large proportion of visceral leishmaniasis cases. The debate on the mode of reproduction and population structure of Leishmania parasites remains opened. It has been suggested that Leishmania parasites could alternate different modes of reproduction, more particularly clonality and frequent recombinations either between related individuals (endogamy) or between unrelated individuals (outcrossing) within strongly isolated subpopulations. To determine whether this assumption is generalized to other species, a population genetics analysis within Leishmania donovani complex strains was conducted within a single village. The results suggest that a mixed-mating reproduction system exists, an important heterogeneity of subsamples and the coexistence of several genetic entities in Sudanese L. donovani. Indeed, results showed significant genetic differentiation between the three taxa (L. donovani, L. infantum and L. archibaldi) and between the human or canine strains of such taxa, suggesting that there may be different imbricated transmission cycles involving either dogs or humans. Results also are in agreement with an almost strict specificity of L. donovani stricto sensu to human hosts. This empirical study demonstrates the complexity of population structure in the genus Leishmania and the need to pursue such kind of analyses at the smallest possible spatio-temporal and ecological scales. PMID:22206035

Leishmania major large RAB GTPase is highly immunogenic in individuals immune to cutaneous and visceral leishmaniasis.

PubMed

Chamakh-Ayari, Rym; Chenik, Mehdi; Chakroun, Ahmed Sahbi; Bahi-Jaber, Narges; Aoun, Karim; Meddeb-Garnaoui, Amel

2017-04-17

We previously identified a Leishmania (L.) major large RAB GTPase (LmlRAB), a new atypical RAB GTPase protein. It is highly conserved in Leishmania species while displaying low level of homology with mammalian homologues. Leishmania small RAB GTPases proteins have been involved in regulation of exocytic and endocytic pathways whereas the role of large RAB GTPases proteins has not been characterized yet. We report here the immunogenicity of both recombinant rLmlRAB and rLmlRABC, in individuals with immunity against L. major or L. infantum. PBMC were isolated from individuals cured of L. major (CCLm) or from healthy individuals. The latter were subdivided into high or low IFN-γ responders. Healthy high IFN-γ responders, considered as asymptomatics, were living in an endemic area for L. major (HHRLm) or L. infantum (HHRLi). Healthy low IFN-γ responders (HLR) were considered as naïve controls. Cells from all volunteers were stimulated with rLmlRAB or rLmlRABC. Cytokines were analysed by CBA and ELISA and phenotypes of IFN-γ-producing cells were analysed by flow cytometry. Both rLmlRAB and rLmlRABC induced high significant levels of IFN-γ in CCLm, HHRLm and HHRLi groups. Phenotype analysis of rLmlRAB and rLmlRABC-stimulated T cells in CCLm individuals showed a significant increase in the percentage of specific IFN-γ-producing CD4+ and CD8+ T cells. rLmlRAB induced significant granzyme B levels in CCLm and HHRLm. Low but significant granzyme B levels were detected in naïve group. IL-10 was detected in immune and naïve individuals. We showed that rLmlRAB protein and its divergent carboxy-terminal part induced a predominant Th1 response in individuals immune to L. major or L. infantum. Our results suggest that rLmlRAB and rLmlRABC proteins are potential cross-species vaccine candidates against cutaneous and visceral leishmaniasis.

Natural infection of Algerian hedgehog, Atelerix algirus (Lereboullet 1842) with Leishmania parasites in Tunisia.

PubMed

Chemkhi, Jomaa; Souguir, Hejer; Ali, Insaf Bel Hadj; Driss, Mehdi; Guizani, Ikram; Guerbouj, Souheila

2015-10-01

In Tunisia, Leishmania parasites are responsible of visceral leishmaniasis, caused by Leishmania infantum species while three cutaneous disease forms are documented: chronic cutaneous leishmaniasis due to Leishmania killicki, sporadic cutaneous form (SCL) caused by L. infantum and the predominant zoonotic cutaneous leishmanaisis (ZCL) due to Leishmania major. ZCL reservoirs are rodents of the Psammomys and Meriones genera, while for SCL the dog is supposed to be a reservoir. Ctenodactylus gundii is involved in the transmission of L. killicki. However, other mammals could constitute potential reservoir hosts in Tunisia and other North African countries. In order to explore the role of hedgehogs as potential reservoirs of leishmaniasis, specimens (N=6) were captured during July-November period in 2011-2013 in an SCL endemic area in El Kef region, North-Western Tunisia. Using morphological characteristics, all specimens were described and measured. Biopsies from liver, heart, kidney and spleen of each animal were used to extract genomic DNA, which was further used in PCR assays to assess the presence of Leishmania parasites. Different PCRs targeting kinetoplast minicircles, ITS1, mini-exon genes and a repetitive Leishmania- specific sequence, were applied. To further identify Leishmania species involved, RFLP analysis of amplified fragments was performed with appropriate restriction enzymes. Using morphological characters, animals were identified as North African hedgehogs, also called Algerian hedgehogs, that belong to the Erinaceidae family, genus Atelerix Pomel 1848, and species algirus (Lereboullet, 1842). PCR results showed in total that all specimens were Leishmania infected, with different organs incriminated, mainly liver and spleen. Results were confirmed by direct sequencing of amplified fragments. Species identification showed that all specimens were infected with L. major, three of which were additionally co-infected with L. infantum. The present study

Evolutionary comparison of prenylation pathway in kinetoplastid Leishmania and its sister Leptomonas.

PubMed

Chauhan, Indira Singh; Kaur, Jaspreet; Krishna, Shagun; Ghosh, Arpita; Singh, Prashant; Siddiqi, Mohammad Imran; Singh, Neeloo

2015-11-21

Leptomonas is monogenetic kinetoplastid parasite of insects and is primitive in comparison to Leishmania. Comparative studies of these two kinetoplastid may share light on the evolutionary transition to dixenous parasitism in Leishmania. In order to adapt and survive within two hosts, Leishmania species must have acquired virulence factors in addition to mechanisms that mediate susceptibility/resistance to infection in the pathology associated with disease. Rab proteins are key mediators of vesicle transport and contribute greatly to the evolution of complexity of membrane transport system. In this study we used our whole genome sequence data of these two divergent kinetoplastids to analyze the orthologues/paralogues of Rab proteins. During change of lifestyle from monogenetic (Leptomonas) to digenetic (Leishmania), we found that the prenyl machinery remained unchanged. Geranylgeranyl transferase-I (GGTase-I) was absent in both Leishmania and its sister Leptomonas. Farnesyltransferase (FTase) and geranylgeranyl transferase-II (GGTase-II) were identified for protein prenylation. We predict that activity of the missing alpha-subunit (α-subunit) of GGTase-II in Leptomonas was probably contributed by the α-subunit of FTase, while beta-subunit (β-subunit) of GGTase-II was conserved and indicated functional conservation in the evolution of these two kinetoplastids. Therefore the β-subunit emerges as an excellent target for compounds inhibiting parasite activity in clinical cases of co-infections. We also confirmed that during the evolution to digenetic life style in Leishmania, the parasite acquired capabilities to evade drug action and maintain parasite virulence in the host with the incorporation of short-chain dehydrogenase/reductase (SDR/MDR) superfamily in Rab genes. Our study based on whole genome sequences is the first to build comparative evolutionary analysis and identification of prenylation proteins in Leishmania and its sister Leptomonas. The information

Affinity labeling of the folate-methotrexate transporter from Leishmania donovani

DOE Office of Scientific and Technical Information (OSTI.GOV)

Beck, J.T.; Ullman, B.

1989-08-22

An affinity labeling technique has been developed to identify the folate-methotrexate transporter of Leishmania donovani promastigotes using activated derivatives of the ligands. These activated derivatives were synthesized by incubating folate and methotrexate with a 10-fold excess of 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide (EDC) for 10 min at ambient temperature in dimethyl sulfoxide. When intact wild-type (DI700) Leishmania donovani or preparations of their membranes were incubated with a 0.4 {mu}M concentration of either activated ({sup 3}H)folate or activated ({sup 3}H)methotrexate, the radiolabeled ligands were covalently incorporated into a polypeptide with a molecular weight of approximately 46,000, as demonstrated by SDS-polyacrylamide gel electrophoresis. No affinity labelingmore » of a 46,000-dalton protein was observed when equimolar concentrations of activated radiolabeled ligands were incubated with intact cells or membranes prepared from a methotrexate-resistant mutant clone of Leishmania donovani, MTXA5, that is genetically defective in folate-methotrexate transport capability. Time course studies indicated that maximal labeling of the 46,000-dalton protein occurred within 5-10 min of incubation of intact cells with activated ligand. These studies provide biochemical evidence that the folate-methotrexate transporter of Leishmania donovani can be identified in crude extracts by an affinity labeling technique and serve as a prerequisite to further analysis of the transport protein by providing a vehicle for subsequent purification of this membrane carrier. Moreover, these investigations suggest that the affinity labeling technique using EDC-activated ligands may be exploitable to analyze other cell surface binding proteins in Leishmania donovani, as well as in other organisms.« less

Leishmania amazonensis Engages CD36 to Drive Parasitophorous Vacuole Maturation

PubMed Central

Okuda, Kendi; Tong, Mei; Dempsey, Brian; Moore, Kathryn J.; Gazzinelli, Ricardo T.; Silverman, Neal

2016-01-01

Leishmania amastigotes manipulate the activity of macrophages to favor their own success. However, very little is known about the role of innate recognition and signaling triggered by amastigotes in this host-parasite interaction. In this work we developed a new infection model in adult Drosophila to take advantage of its superior genetic resources to identify novel host factors limiting Leishmania amazonensis infection. The model is based on the capacity of macrophage-like cells, plasmatocytes, to phagocytose and control the proliferation of parasites injected into adult flies. Using this model, we screened a collection of RNAi-expressing flies for anti-Leishmania defense factors. Notably, we found three CD36-like scavenger receptors that were important for defending against Leishmania infection. Mechanistic studies in mouse macrophages showed that CD36 accumulates specifically at sites where the parasite contacts the parasitophorous vacuole membrane. Furthermore, CD36-deficient macrophages were defective in the formation of the large parasitophorous vacuole typical of L. amazonensis infection, a phenotype caused by inefficient fusion with late endosomes and/or lysosomes. These data identify an unprecedented role for CD36 in the biogenesis of the parasitophorous vacuole and further highlight the utility of Drosophila as a model system for dissecting innate immune responses to infection. PMID:27280707

Genomic Confirmation of Hybridisation and Recent Inbreeding in a Vector-Isolated Leishmania Population

PubMed Central

Smith, Barbara A.; Imamura, Hideo; Sanders, Mandy; Svobodova, Milena; Volf, Petr; Berriman, Matthew; Cotton, James A.; Smith, Deborah F.

2014-01-01

Although asexual reproduction via clonal propagation has been proposed as the principal reproductive mechanism across parasitic protozoa of the Leishmania genus, sexual recombination has long been suspected, based on hybrid marker profiles detected in field isolates from different geographical locations. The recent experimental demonstration of a sexual cycle in Leishmania within sand flies has confirmed the occurrence of hybridisation, but knowledge of the parasite life cycle in the wild still remains limited. Here, we use whole genome sequencing to investigate the frequency of sexual reproduction in Leishmania, by sequencing the genomes of 11 Leishmania infantum isolates from sand flies and 1 patient isolate in a focus of cutaneous leishmaniasis in the Çukurova province of southeast Turkey. This is the first genome-wide examination of a vector-isolated population of Leishmania parasites. A genome-wide pattern of patchy heterozygosity and SNP density was observed both within individual strains and across the whole group. Comparisons with other Leishmania donovani complex genome sequences suggest that these isolates are derived from a single cross of two diverse strains with subsequent recombination within the population. This interpretation is supported by a statistical model of the genomic variability for each strain compared to the L. infantum reference genome strain as well as genome-wide scans for recombination within the population. Further analysis of these heterozygous blocks indicates that the two parents were phylogenetically distinct. Patterns of linkage disequilibrium indicate that this population reproduced primarily clonally following the original hybridisation event, but that some recombination also occurred. This observation allowed us to estimate the relative rates of sexual and asexual reproduction within this population, to our knowledge the first quantitative estimate of these events during the Leishmania life cycle. PMID:24453988

Regional, age and respiratory-secretion-specific prevalence of respiratory viruses associated with asthma exacerbation: a literature review.

PubMed

Zheng, Xue-Yan; Xu, Yan-Jun; Guan, Wei-Jie; Lin, Li-Feng

2018-04-01

Despite increased understanding of how viral infection is involved in asthma exacerbations, it is less clear which viruses are involved and to what extent they contribute to asthma exacerbations. Here, we sought to determine the prevalence of different respiratory viruses during asthma exacerbations. Systematic computerized searches of the literature up to June 2017 without language limitation were performed. The primary focus was on the prevalence of respiratory viruses, including AdV (adenovirus), BoV (bocavirus), CoV (coronavirus), CMV (cytomegalovirus), EnV (enterovirus), HSV (herpes simplex virus), IfV (influenza virus), MpV (metapneumovirus), PiV (parainfluenzavirus), RV (rhinovirus) and RSV (respiratory syncytial virus) during asthma exacerbations. We also examined the prevalence of viral infection stratified by age, geographic region, type of respiratory secretion, and detection method. Sixty articles were included in the final analysis. During asthma exacerbations, the mean prevalence of AdV, BoV, CoV, CMV, EnV, HSV, IfV, MpV, PiV, RV and RSV was 3.8%, 6.9%, 8.4%, 7.2%, 10.1%, 12.3%, 10.0%, 5.3%, 5.6%, 42.1% and 13.6%, respectively. EnV, MPV, RV and RSV were more prevalent in children, whereas AdV, BoV, CoV, IfV and PiV were more frequently present in adults. RV was the major virus detected globally, except in Africa. RV could be detected in both the upper and lower airway. Polymerase chain reaction was the most sensitive method for detecting viral infection. Our findings indicate the need to develop prophylactic polyvalent or polyvirus (including RV, EnV, IfV and RSV) vaccines that produce herd immunity and reduce the healthcare burden associated with virus-induced asthma exacerbations.

Leishmania carbon metabolism in the macrophage phagolysosome- feast or famine?

PubMed

McConville, Malcolm J; Saunders, Eleanor C; Kloehn, Joachim; Dagley, Michael J

2015-01-01

A number of medically important microbial pathogens target and proliferate within macrophages and other phagocytic cells in their mammalian hosts. While the majority of these pathogens replicate within the host cell cytosol or non-hydrolytic vacuolar compartments, a few, including protists belonging to the genus Leishmania, proliferate long-term within mature lysosome compartments.  How these parasites achieve this feat remains poorly defined. In this review, we highlight recent studies that suggest that Leishmania virulence is intimately linked to programmed changes in the growth rate and carbon metabolism of the obligate intra-macrophage stages. We propose that activation of a slow growth and a stringent metabolic response confers resistance to multiple stresses (oxidative, temperature, pH), as well as both nutrient limitation and nutrient excess within this niche. These studies highlight the importance of metabolic processes as key virulence determinants in Leishmania.

[Leishmania infantum MON-1: the only zymodeme isolated in canine leishmaniasis in Tunisia].

PubMed

Aoun, K; Diouani, M F; Benikhlef, R; Bouratbine, A; Ben Haj Ali, S; Harrat, Z; Belkaïd, M; Kilani, M; Ben Ismaïl, R

2003-05-01

Iso-enzymatic characterization of 19 Leishmania strains isolated from Tunisian dogs revealed that all correspond to Leishmania infantum MON-1. This confirms the role of dog as a reservoir of the commonest zymodeme responsible for human visceral leishmaniasis in the Mediterranean basin. However, and in spite of the high number of identifications, many zymodemes, mainly the dermotropic ones as Leishmania infantum MON-24, frequently reported in humans, had never been isolated from dogs. The study of their cultural characteristics may contribute to explain the particular iso-enzymatic profile of dogs strains.

Detection of Leishmania spp. in Bats from an Area of Brazil Endemic for Visceral Leishmaniasis.

PubMed

de Rezende, M B; Herrera, H M; Carvalho, C M E; Carvalho Anjos, E A; Ramos, C A N; de Araújo, F R; Torres, J M; de Oliveira, C E

2017-12-01

The multihost parasites Leishmania spp. infect a broad range of wild mammalian species including bats. Several species of bats have adapted to a variety of food resources and shelters in urban areas. This study aimed to detect Leishmania spp. DNA in bats present in forest fragments located in metropolitan areas endemic for leishmaniasis in Campo Grande, Mato Grosso do Sul (MS), Brazil. Blood samples were obtained from 80 individuals, including eight species of Phyllostomidae and one species of Vespertilionidae. Thirty of the 80 bats were positive for Leishmania spp. using conventional PCR, all belonging to the family Phyllostomidae. Eighteen samples tested by real-time PCR (qPCR) using specific primers for the kDNA of Leishmania infantum were positive. To the best of our knowledge, this is the first report detecting Leishmania spp. in Platyrrhinus incarum in addition to being the first reported detection of L. infantum in the bat species Phyllostomus discolor, Platyrrhinus lineatus, Artibeus planirostris and Artibeus lituratus. Our results show that bats can host Leishmania spp. in areas endemic for leishmaniasis, which must be taken into account in disease control operations by public health authorities. © 2017 Blackwell Verlag GmbH.

The fusion of Toxoplasma gondii SAG1 vaccine candidate to Leishmania infantum heat shock protein 83-kDa improves expression levels in tobacco chloroplasts.

PubMed

Albarracín, Romina M; Becher, Melina Laguía; Farran, Inmaculada; Sander, Valeria A; Corigliano, Mariana G; Yácono, María L; Pariani, Sebastián; López, Edwin Sánchez; Veramendi, Jon; Clemente, Marina

2015-05-01

Chloroplast transformation technology has emerged as an alternative platform offering many advantages over nuclear transformation. SAG1 is the main surface antigen of the intracellular parasite Toxoplasma gondii and a promising candidate to produce an anti-T. gondii vaccine. The aim of this study was to investigate the expression of SAG1 using chloroplast transformation technology in tobacco plants. In order to improve expression in transplastomic plants, we also expressed the 90-kDa heat shock protein of Leishmania infantum (LiHsp83) as a carrier for the SAG1 antigen. SAG1 protein accumulation in transplastomic plants was approximately 0.1-0.2 μg per gram of fresh weight (FW). Fusion of SAG1 to LiHsp83 significantly increased the level of SAG1 accumulation in tobacco chloroplasts (by up to 500-fold). We also evaluated the functionality of the chLiHsp83-SAG1. Three human seropositive samples reacted with SAG1 expressed in transplastomic chLiHsp83-SAG1 plants. Oral immunization with chLiHsp83-SAG1 elicited a significant reduction of the cyst burden that correlated with an increase of SAG1-specific antibodies. We propose the fusion of foreign proteins to LiHsp83 as a novel strategy to increase the expression level of the recombinant proteins using chloroplast transformation technology, thus addressing one of the current challenges for this approach in antigen protein production. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Leishmania sand fly interaction: progress and challenges.

PubMed

Bates, Paul A

2008-08-01

Complex interactions occurs between Leishmania parasites and their sand fly vectors. Promastigotes of Leishmania live exclusively within the gut, possess flagella and are motile, and kinesins, kinases and G proteins have been described that play a role in regulating flagellar assembly. Movement within the gut is not random: promastigotes can detect gradients of solutes via chemotaxis and osmotaxis. Further they use their flagella to attach to the fly midgut using surface glyconconjugates, a key step in establishment of the infection. Differentiation of mammal-infective stages is characterised by significant biochemical and cellular remodelling. Further, the parasites can manipulate the behaviour of the vector to maximise their transmission, and flies may even deliver altruistic apoptotic forms to aid transmission of infective stages.

PKC/ROS-Mediated NLRP3 Inflammasome Activation Is Attenuated by Leishmania Zinc-Metalloprotease during Infection

PubMed Central

Jung, Jee Yong; Chang, Kwang-Poo; Olivier, Martin

2015-01-01

Parasites of the Leishmania genus infect and survive within macrophages by inhibiting several microbicidal molecules, such as nitric oxide and pro-inflammatory cytokines. In this context, various species of Leishmania have been reported to inhibit or reduce the production of IL-1β both in vitro and in vivo. However, the mechanism whereby Leishmania parasites are able to affect IL-1β production and secretion by macrophages is still not fully understood. Dependent on the stimulus at hand, the maturation of IL-1β is facilitated by different inflammasome complexes. The NLRP3 inflammasome has been shown to be of pivotal importance in the detection of danger molecules such as inorganic crystals like asbestos, silica and malarial hemozoin, (HZ) as well as infectious agents. In the present work, we investigated whether Leishmania parasites modulate NLRP3 inflammasome activation. Using PMA-differentiated THP-1 cells, we demonstrate that Leishmania infection effectively inhibits macrophage IL-1β production upon stimulation. In this context, the expression and activity of the metalloprotease GP63 - a critical virulence factor expressed by all infectious Leishmania species - is a prerequisite for a Leishmania-mediated reduction of IL-1β secretion. Accordingly, L. mexicana, purified GP63 and GP63-containing exosomes, caused the inhibition of macrophage IL-1β production. Leishmania-dependent suppression of IL-1β secretion is accompanied by an inhibition of reactive oxygen species (ROS) production that has previously been shown to be associated with NLRP3 inflammasome activation. The observed loss of ROS production was due to an impaired PKC-mediated protein phosphorylation. Furthermore, ROS-independent inflammasome activation was inhibited, possibly due to an observed GP63-dependent cleavage of inflammasome and inflammasome-related proteins. Collectively for the first time, we herein provide evidence that the protozoan parasite Leishmania, through its surface

In vitro antiprotozoal activity of (S)-cis-Verbenol against Leishmania spp. and Trypanosoma cruzi.

PubMed

Yaluff, Gloria; Vega, Celeste; Alvarenga, Nelson

2017-04-01

(S)-cis-Verbenol, a monoterpene frequently found as a component of essential oils, was assayed against Leishmania amazonensis, Leishmania infantum, Leishmania brasiliensis and against two strains of Trypanosoma cruzi. The cytotoxicity of the compound was also assayed against human fibroblast cells using a colorimetric method. Benznidazole was used as reference drug against T. cruzi and amphotericin B was used against Leishmania spp. The compound showed good activity against the trypanosomes, being more active against the CL Brenner strain, with an IC 50 value of 8.3μg/mL. Against Leishmania, the IC 50 values were between 2.1 and 3.8μg/mL. The compound showed no cytotoxicity against human fibroblasts at the concentrations assayed and was 100-500 times more toxic for the parasites than for the human cells, as indicated by the selectivity indexes. The results open interesting perspectives about the potential of (S)-cis-Verbenol and other individual components of essential oils for the treatment of these diseases. Copyright © 2016 Elsevier B.V. All rights reserved.

New primers for the detection Leishmania species by multiplex polymerase chain reaction.

PubMed

Conter, Carolina Cella; Lonardoni, Maria Valdrinez Campana; Aristides, Sandra Mara Alessi; Cardoso, Rosilene Fressatti; Silveira, Thaís Gomes Verzignassi

2018-02-01

Leishmaniasis is caused by protozoa of the Leishmania genus, which is divided into subgenus Viannia and Leishmania. In humans, the course of infection largely depends on the host-parasite relationship and primarily of the infective species. The objective of the present study was to design specific primers to the identification of Leishmania species using multiplex PCR. Four primers were designed, based on the GenBank sequences of the kDNA minicircle, amplifying 127 bp for subgenus Viannia, 100 bp for L. amazonensis, and 60 bp for Leishmania donovani complex and L. major. None of the primers amplified Trypanosoma cruzi or L. mexicana. The limit of detection of multiplex PCR was 2 × 10 -5 parasites for L. braziliensis, 2 x 10 -3 parasites for L. amazonensis, and 1.4 × 10 -3 parasites for L. infantum. The high sensitivity of multiplex PCR was confirmed by the detection of parasites in different biological samples, including lesion scrapings, spleen imprinting of a hamster, sandflies, and blood. The multiplex PCR that was developed herein presented good performance with regard to detecting and identifying the parasite in different biological samples and may thus be useful for diagnosis, decision making with regard to the proper therapeutic approach, and determining the geographic distribution of Leishmania species.

Increased Glycolytic ATP Synthesis Is Associated with Tafenoquine Resistance in Leishmania major▿

PubMed Central

Manzano, José Ignacio; Carvalho, Luis; Pérez-Victoria, José M.; Castanys, Santiago; Gamarro, Francisco

2011-01-01

Tafenoquine (TFQ), an 8-aminoquinoline used to treat and prevent Plasmodium infections, could represent an alternative therapy for leishmaniasis. Indeed, TFQ has shown significant leishmanicidal activity both in vitro and in vivo, where it targets Leishmania mitochondria and activates a final apoptosis-like process. In order not to jeopardize the life span of this potential antileishmania drug, it is important to determine the likelihood that Leishmania will develop resistance to TFQ and the mechanisms of resistance induced. To address this issue, a TFQ-resistant Leishmania major promastigote line (R4) was selected. This resistance, which is unstable in a drug-free medium (revertant line), was maintained in intramacrophage amastigote forms, and R4 promastigotes were found to be cross-resistant to other 8-aminoquinolines. A decreased TFQ uptake, which is probably associated with an alkalinization of the intracellular pH rather than drug efflux, was observed for both the R4 and revertant lines. TFQ induces a decrease in ATP synthesis in all Leishmania lines, although total ATP levels were maintained at higher values in R4 parasites. In contrast, ATP synthesis by glycolysis was significantly increased in R4 parasites, whereas mitochondrial ATP synthesis was similar to that in wild-type parasites. We therefore conclude that increased glycolytic ATP synthesis is the main mechanism underlying TFQ resistance in Leishmania. PMID:21199921

Increased glycolytic ATP synthesis is associated with tafenoquine resistance in Leishmania major.

PubMed

Manzano, José Ignacio; Carvalho, Luis; Pérez-Victoria, José M; Castanys, Santiago; Gamarro, Francisco

2011-03-01

Tafenoquine (TFQ), an 8-aminoquinoline used to treat and prevent Plasmodium infections, could represent an alternative therapy for leishmaniasis. Indeed, TFQ has shown significant leishmanicidal activity both in vitro and in vivo, where it targets Leishmania mitochondria and activates a final apoptosis-like process. In order not to jeopardize the life span of this potential antileishmania drug, it is important to determine the likelihood that Leishmania will develop resistance to TFQ and the mechanisms of resistance induced. To address this issue, a TFQ-resistant Leishmania major promastigote line (R4) was selected. This resistance, which is unstable in a drug-free medium (revertant line), was maintained in intramacrophage amastigote forms, and R4 promastigotes were found to be cross-resistant to other 8-aminoquinolines. A decreased TFQ uptake, which is probably associated with an alkalinization of the intracellular pH rather than drug efflux, was observed for both the R4 and revertant lines. TFQ induces a decrease in ATP synthesis in all Leishmania lines, although total ATP levels were maintained at higher values in R4 parasites. In contrast, ATP synthesis by glycolysis was significantly increased in R4 parasites, whereas mitochondrial ATP synthesis was similar to that in wild-type parasites. We therefore conclude that increased glycolytic ATP synthesis is the main mechanism underlying TFQ resistance in Leishmania.

Identification and assessment of COPD exacerbations.

PubMed

Oliveira, A S; Munhá, J; Bugalho, A; Guimarães, M; Reis, G; Marques, A

2017-12-24

Chronic Obstructive Pulmonary Disease (COPD) exacerbations play a central role in the disease natural history of the disease, affecting its overall severity, decreasing pulmonary function, worsening underlying co-morbidities, impairing quality of life (QoL) and leading to severe morbidity and mortality. Therefore, identification and correct assessment of COPD exacerbations is paramount, given it will strongly influence therapy success. For the identification of exacerbations, several questionnaires exist, with varying degrees of complexity. However, most questionnaires remain of limited clinical utility, and symptom scales seem to be more useful in clinical practice. In the assessment of exacerbations, the type and degree of severity should be ascertained in order to define the management setting and optimize treatment options. Still, a consensual and universal classification system to assess the severity and type of an exacerbation is lacking, and there are no established criteria for less severely ill patients not requiring hospital assessment. This might lead to under-reporting of minor to moderate exacerbations, which has an impact on patients' health status. There is a clear unmet need to develop clinically useful questionnaires and a comprehensive system to evaluate the severity of exacerbations that can be used in all settings, from primary health care to general hospitals. Copyright © 2017. Published by Elsevier España, S.L.U.

Virulence determinants of Moraxella catarrhalis: distribution and considerations for vaccine development.

PubMed

Blakeway, Luke V; Tan, Aimee; Peak, Ian R A; Seib, Kate L

2017-10-01

Moraxella catarrhalis is a human-restricted opportunistic bacterial pathogen of the respiratory mucosa. It frequently colonizes the nasopharynx asymptomatically, but is also an important causative agent of otitis media (OM) in children, and plays a significant role in acute exacerbations of chronic obstructive pulmonary disease (COPD) in adults. As the current treatment options for M. catarrhalis infection in OM and exacerbations of COPD are often ineffective, the development of an efficacious vaccine is warranted. However, no vaccine candidates for M. catarrhalis have progressed to clinical trials, and information regarding the distribution of M. catarrhalis virulence factors and vaccine candidates is inconsistent in the literature. It is largely unknown if virulence is associated with particular strains or subpopulations of M. catarrhalis, or if differences in clinical manifestation can be attributed to the heterogeneous expression of specific M. catarrhalis virulence factors in the circulating population. Further investigation of the distribution of M. catarrhalis virulence factors in the context of carriage and disease is required so that vaccine development may be targeted at relevant antigens that are conserved among disease-causing strains. The challenge of determining which of the proposed M. catarrhalis virulence factors are relevant to human disease is amplified by the lack of a standardized M. catarrhalis typing system to facilitate direct comparisons of worldwide isolates. Here we summarize and evaluate proposed relationships between M. catarrhalis subpopulations and specific virulence factors in the context of colonization and disease, as well as the current methods used to infer these associations.

An epidemic outbreak of canine cutaneous leishmaniasis in Colombia caused by Leishmania braziliensis and Leishmania panamensis.

PubMed

Vélez, Iván D; Carrillo, Lina M; López, Liliana; Rodríguez, Erwin; Robledo, Sara M

2012-05-01

The largest recorded outbreak of cutaneous leishmaniasis in Colombia's history occurred during 2005-2009 in soldiers of the Colombian Army, with ~40,000 cases. This outbreak was caused by the influx of military personnel into the jungle with the mission of combat illicit crops and the guerrilla. The soldiers remain for long periods within the rainforest and are exposed to the bite of infected sand flies. During the military activities, soldiers work with dogs specially trained to detect landmines, and therefore, dogs are also exposed to the infected sand flies and show high incidence of cutaneous leishmaniasis (CL). This work describes an epidemic outbreak of canine CL caused by Leishmania braziliensis and Leishmania panamensis in Colombia, South America. The clinical features of the disease and the response to treatment with pentavalent antimonials observed in 72 guard dogs from the Colombian Army are described. A program for prevention and control of canine CL is also discussed.

Antiproliferative and ultrastructural effects of phenethylamine derivatives on promastigotes and amastigotes of Leishmania (Leishmania) infantum chagasi.

PubMed

Brasil, Paula Ferreira; de Freitas, Júlia Araújo; Barreto, Anna Léa Silva; Adade, Camila Marques; Reis de Sá, Leandro Figueira; Constantino-Teles, Pamella; Toledo, Fabiano Travanca; de Sousa, Bruno A; Gonçalves, Augusto Cesar; Romanos, Maria Teresa Villela; Comasseto, João V; Dos Santos, Alcindo A; Tessis, Ana Claudia; Souto-Padrón, Thais; Soares, Rosangela Maria A; Ferreira-Pereira, Antonio

2017-04-01

Leishmania (Leishmania) infantum chagasi is one of the agents that cause visceral leishmaniasis. This disease occurs more frequently in third world countries, such as Brazil. The treatment is arduous, and is dependent on just a few drugs like the antimonial derivatives and amphotericin B. Moreover, these drugs are not only expensive, but they can also cause severe side effects and require long-term treatment. Therefore, it is very important to find new compounds that are effective against leishmaniasis. In the present work we evaluated a new group of synthetic amides against the promastigote and amastigote forms of L. infantum chagasi. The results showed that one of these amides in particular, presented very effective activity against the promastigotes and amastigotes of L. infantum chagasi at low concentrations and it also presented low toxicity for mammal cells, which makes this synthetic amide a promising drug for combating leishmaniasis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

An Epidemic Outbreak of Canine Cutaneous Leishmaniasis in Colombia Caused by Leishmania braziliensis and Leishmania panamensis

PubMed Central

Vélez, Iván D.; Carrillo, Lina M.; López, Liliana; Rodríguez, Erwin; Robledo, Sara M.

2012-01-01

The largest recorded outbreak of cutaneous leishmaniasis in Colombia's history occurred during 2005–2009 in soldiers of the Colombian Army, with ∼40,000 cases. This outbreak was caused by the influx of military personnel into the jungle with the mission of combat illicit crops and the guerrilla. The soldiers remain for long periods within the rainforest and are exposed to the bite of infected sand flies. During the military activities, soldiers work with dogs specially trained to detect landmines, and therefore, dogs are also exposed to the infected sand flies and show high incidence of cutaneous leishmaniasis (CL). This work describes an epidemic outbreak of canine CL caused by Leishmania braziliensis and Leishmania panamensis in Colombia, South America. The clinical features of the disease and the response to treatment with pentavalent antimonials observed in 72 guard dogs from the Colombian Army are described. A program for prevention and control of canine CL is also discussed. PMID:22556078

Leishmania infection: painful or painless?

PubMed

Borghi, Sergio M; Fattori, Victor; Conchon-Costa, Ivete; Pinge-Filho, Phileno; Pavanelli, Wander R; Verri, Waldiceu A

2017-02-01

The complex life cycle and immunopathological features underpinning the interaction of Leishmania parasites and their mammalian hosts poses frequent poorly explored and inconclusively resolved questions. The altered nociceptive signals over the course of leishmaniasis remain an intriguing issue for nociceptive and parasitology researchers. Experimental investigations have utilized behavioral, morphological, and neuro-immune approaches in the study of experimental cutaneous leishmaniasis (CL). The data generated indicates new venues for the study of the pathological characteristics of nociceptive processing in this parasitic disease. Leishmania-induced pain may be easily observed in mice and rats. However, nociceptive data is more complex in human investigations, including the occurrence of painless lesions in mucocutaneous and cutaneous leishmaniasis. Data from recent decades indicate that humans can also be affected by pain-related symptoms, often distinct from the region of body infection. The molecular and cellular mechanisms underlying such variable nociceptive states in humans during the course of leishmaniasis are an active area of research. The present article reviews nociception in leishmaniasis, including in experimental models of CL and clinical reports.

Sulfonamides as Inhibitors of Leishmania – Potential New Treatments for Leishmaniasis

PubMed Central

Katinas, Jade; Epplin, Rachel; Hamaker, Christopher; Jones, Marjorie A.

2017-01-01

Introduction: Leishmaniasis is an endemic disease caused by the protozoan parasite Leishmania. Current treatments for the parasite are limited by cost, availability and drug resistance as the occurrence of leishmaniasis continues to be more prevalent. Sulfonamides are a class of compounds with medicinal properties which have been used to treat bacterial and parasitic disease via various pathways especially as antimetabolites for folic acid. Methods: New derivatives of sulfonamide compounds were assessed for their impact on Leishmania cell viability and potential pathways for inhibition were evaluated. Leishmania tarentolae (ATCC Strain 30143) axenic promastigote cells were grown in brain heart infusion (BHI) medium and treated with varying concentrations of the new sulfonamide compounds. Light microscopy and viability tests were used to assess the cells with and without treatment. Discussion: A non-water soluble sulfonamide was determined to have 90-96% viability inhibition 24 hours after treatment with 100 µM final concentration. Because Leishmania are also autotrophs for folate precursors, the folic acid pathway was identified as a target for sulfonamide inhibition. When folic acid was added to untreated Leishmania, cell proliferation increased. A water soluble derivative of the inhibitory sulfonamide was synthesized and evaluated, resulting in less viability inhibition with a single dose (approximately 70% viability inhibition after 24 hours with 100 µM final concentration), but additive inhibition with multiple doses of the compound. Results: However, the potential mechanism of inhibition was different between the water-soluble and non-water soluble sulfonamides. The inhibitory effects and potential pathways of inhibition indicate that these compounds may be new treatments for this disease. PMID:29399442

Impaired Hemorheology in Exacerbations of COPD

PubMed Central

Can, Ilknur; Kilic-Erkek, Ozgen; Altinisik, Goksel; Bor-Kucukatay, Melek

2017-01-01

Background Chronic obstructive pulmonary disease (COPD) is characterized by progressive airflow limitation. Cardiovascular-related comorbidities are established to contribute to morbidity and mortality especially during exacerbations. The aim of the current study was to determine alterations in hemorheology (erythrocyte aggregation, deformability) in newly diagnosed COPD patients and their response to medical treatment and to compare with values of COPD patients with exacerbations. Materials and Methods The study comprised 13 COPD patients, 12 controls, and 16 COPD patients with exacerbations. The severity of COPD was determined according to Global Initiative for Chronic Obstructive Lung Disease guidelines. Red blood cell (RBC) deformability and aggregation were measured by an ektacytometer. Results RBC deformability of COPD patients with exacerbations was decreased compared to the other groups. Erythrocyte aggregation and plasma fibrinogen of COPD patients determined during exacerbations were higher than control. Conclusion Decreased RBC deformability and increased aggregation associated with exacerbations of COPD may serve as unfavorable mechanisms to worsen oxygenation and thus clinical symptoms of the patient. Treatment modalities that modify rheological parameters might be beneficial. PMID:29089816

Curcumin overcomes the inhibitory effect of nitric oxide on Leishmania.

PubMed

Chan, Marion Man-Ying; Adapala, Naga Suresh; Fong, Dunne

2005-04-01

Upon Leishmania infection, macrophages are activated to produce nitrogen and oxygen radicals simultaneously. It is well established that the infected host cells rely on nitric oxide (NO) as the major weapon against the intracellular parasite. In India where leishmaniasis is endemic, the spice turmeric is used prolifically in food and for insect bites. Curcumin, the active principle of turmeric, is a scavenger of NO. This report shows that curcumin protects promastigotes and amastigotes of the visceral species, Leishmania donovani, and promastigotes of the cutaneous species, L. major, against the actions of S-nitroso-N-acetyl-D,L-penicillamine (SNAP) and DETANONOate, which release NO, 3-morpholino-sydnonimine hydrochloride (SIN-1), which releases NO and superoxide, and peroxynitrite, which is formed from the reaction of NO with superoxide. Thus, curcumin, as an antioxidant, is capable of blocking the action of both NO and NO congeners on the Leishmania parasite.

Oligomerization of coronin: Implication on actin filament length in Leishmania.

PubMed

Srivastava, Rashmi; Prasadareddy Kajuluri, Lova; Pathak, Neelam; Gupta, Chhitar M; Sahasrabuddhe, Amogh A

2015-12-01

Coronin proteins bind with actin filaments and participate in regulation of actin-dependent processes. These proteins contain a coiled-coil domain at their C-terminus, which is responsible for their dimeric or trimeric forms. However, the functional significance of these oligomeric configurations in organizing the actin cytoskeleton is obscure. Here, we report that the Leishmania coronin exists in a higher oligomeric form through its coiled-coil domain, the truncation of which ablates the ability of Leishmania coronin to assist actin-filament formation. F-actin co-sedimentation assay using purified proteins shows that the coiled-coil domain does not interact with actin-filaments and its absence does not abrogate actin-coronin interaction. Furthermore, it was shown that unlike other coronins, Leishmania coronin interacts with actin-filaments through its unique region. These results provided important insights into the role of coronin oligomerization in modulating actin-network. © 2015 Wiley Periodicals, Inc.

Leishmania and human immunodeficiency virus coinfection: the first 10 years.

PubMed Central

Alvar, J; Cañavate, C; Gutiérrez-Solar, B; Jiménez, M; Laguna, F; López-Vélez, R; Molina, R; Moreno, J

1997-01-01

Over 850 Leishmania-human immunodeficiency virus (HIV) coinfection cases have been recorded, the majority in Europe, where 7 to 17% of HIV-positive individuals with fever have amastigotes, suggesting that Leishmania-infected individuals without symptoms will express symptoms of leishmaniasis if they become immunosuppressed. However, there are indirect reasons and statistical data demonstrating that intravenous drug addiction plays a specific role in Leishmania infantum transmission: an anthroponotic cycle complementary to the zoonotic one has been suggested. Due to anergy in patients with coinfection, L. infantum dermotropic zymodemes are isolated from patient viscera and a higher L. infantum phenotypic variability is seen. Moreover, insect trypanosomatids that are currently considered nonpathogenic have been isolated from coinfected patients. HIV infection and Leishmania infection each induce important analogous immunological changes whose effects are multiplied if they occur concomitantly, such as a Th1-to-Th2 response switch; however, the consequences of the viral infection predominate. In fact, a large proportion of coinfected patients have no detectable anti-Leishmania antibodies. The microorganisms share target cells, and it has been demonstrated in vitro how L. infantum induces the expression of latent HIV-1. Bone marrow culture is the most useful diagnostic technique, but it is invasive. Blood smears and culture are good alternatives. PCR, xenodiagnosis, and circulating-antigen detection are available only in specialized laboratories. The relationship with low levels of CD4+ cells conditions the clinical presentation and evolution of disease. Most patients have visceral leishmaniasis, but asymptomatic, cutaneous, mucocutaneous, diffuse cutaneous, and post-kala-azar dermal leishmaniasis can be produced by L. infantum. The digestive and respiratory tracts are frequently parasitized. The course of coinfection is marked by a high relapse rate. There is a lack

Arginase activity in pathogenic and non-pathogenic species of Leishmania parasites.

PubMed

Badirzadeh, Alireza; Taheri, Tahereh; Taslimi, Yasaman; Abdossamadi, Zahra; Heidari-Kharaji, Maryam; Gholami, Elham; Sedaghat, Baharehsadat; Niyyati, Maryam; Rafati, Sima

2017-07-01

Proliferation of Leishmania (L.) parasites depends on polyamine availability, which can be generated by the L-arginine catabolism and the enzymatic activity of arginase (ARG) of the parasites and of the mammalian hosts. In the present study, we characterized and compared the arginase (arg) genes from pathogenic L. major and L. tropica and from non-pathogenic L. tarentolae. We quantified the level of the ARG activity in promastigotes and macrophages infected with pathogenic L. major and L. tropica and non-pathogenic L. tarentolae amastigotes. The ARG's amino acid sequences of the pathogenic and non-pathogenic Leishmania demonstrated virtually 98.6% and 88% identities with the reference L. major Friedlin ARG. Higher ARG activity was observed in all pathogenic promastigotes as compared to non-pathogenic L. tarentolae. In vitro infection of human macrophage cell line (THP1) with pathogenic and non-pathogenic Leishmania spp. resulted in increased ARG activities in the infected macrophages. The ARG activities present in vivo were assessed in susceptible BALB/c and resistant C57BL/6 mice infected with L. major, L. tropica and L. tarentolae. We demonstrated that during the development of the infection, ARG is induced in both strains of mice infected with pathogenic Leishmania. However, in L. major infected BALB/c mice, the induction of ARG and parasite load increased simultaneously according to the time course of infection, whereas in C57BL/6 mice, the enzyme is upregulated solely during the period of footpad swelling. In L. tropica infected mice, the footpads' swellings were slow to develop and demonstrated minimal cutaneous pathology and ARG activity. In contrast, ARG activity was undetectable in mice inoculated with the non-pathogenic L. tarentolae. Our data suggest that infection by Leishmania parasites can increase ARG activity of the host and provides essential polyamines for parasite salvage and its replication. Moreover, the ARG of Leishmania is vital for parasite

Arginase activity in pathogenic and non-pathogenic species of Leishmania parasites

PubMed Central

Badirzadeh, Alireza; Taheri, Tahereh; Taslimi, Yasaman; Abdossamadi, Zahra; Heidari-Kharaji, Maryam; Gholami, Elham; Sedaghat, Baharehsadat; Niyyati, Maryam

2017-01-01

Proliferation of Leishmania (L.) parasites depends on polyamine availability, which can be generated by the L-arginine catabolism and the enzymatic activity of arginase (ARG) of the parasites and of the mammalian hosts. In the present study, we characterized and compared the arginase (arg) genes from pathogenic L. major and L. tropica and from non-pathogenic L. tarentolae. We quantified the level of the ARG activity in promastigotes and macrophages infected with pathogenic L. major and L. tropica and non-pathogenic L. tarentolae amastigotes. The ARG's amino acid sequences of the pathogenic and non-pathogenic Leishmania demonstrated virtually 98.6% and 88% identities with the reference L. major Friedlin ARG. Higher ARG activity was observed in all pathogenic promastigotes as compared to non-pathogenic L. tarentolae. In vitro infection of human macrophage cell line (THP1) with pathogenic and non-pathogenic Leishmania spp. resulted in increased ARG activities in the infected macrophages. The ARG activities present in vivo were assessed in susceptible BALB/c and resistant C57BL/6 mice infected with L. major, L. tropica and L. tarentolae. We demonstrated that during the development of the infection, ARG is induced in both strains of mice infected with pathogenic Leishmania. However, in L. major infected BALB/c mice, the induction of ARG and parasite load increased simultaneously according to the time course of infection, whereas in C57BL/6 mice, the enzyme is upregulated solely during the period of footpad swelling. In L. tropica infected mice, the footpads' swellings were slow to develop and demonstrated minimal cutaneous pathology and ARG activity. In contrast, ARG activity was undetectable in mice inoculated with the non-pathogenic L. tarentolae. Our data suggest that infection by Leishmania parasites can increase ARG activity of the host and provides essential polyamines for parasite salvage and its replication. Moreover, the ARG of Leishmania is vital for parasite

Comparative genomics of canine-isolated Leishmania (Leishmania) amazonensis from an endemic focus of visceral leishmaniasis in Governador Valadares, southeastern Brazil

PubMed Central

Valdivia, Hugo O.; Almeida, Laila V.; Roatt, Bruno M.; Reis-Cunha, João Luís; Pereira, Agnes Antônia Sampaio; Gontijo, Celia; Fujiwara, Ricardo Toshio; Reis, Alexandre B.; Sanders, Mandy J.; Cotton, James A.; Bartholomeu, Daniella C.

2017-01-01

Leishmaniasis is a highly diverse group of diseases caused by kinetoplastid of the genus Leishmania. These parasites are taxonomically diverse, with human pathogenic species separated into two subgenera according to their development site inside the alimentary tract of the sand fly insect vector. The disease encompasses a variable spectrum of clinical manifestations with tegumentary or visceral symptoms. Among the causative species in Brazil, Leishmania (Leishmania) amazonensis is an important etiological agent of human cutaneous leishmaniasis that accounts for more than 8% of all cases in endemic regions. L. (L.) amazonensis is generally found in the north and northeast regions of Brazil. Here, we report the first isolation of L. (L.) amazonensis from dogs with clinical manifestations of visceral leishmaniasis in Governador Valadares, an endemic focus in the southeastern Brazilian State of Minas Gerais where L. (L.) infantum is also endemic. These isolates were characterized in terms of SNPs, chromosome and gene copy number variations, confirming that they are closely related to a previously sequenced isolate obtained in 1973 from the typical Northern range of this species. The results presented in this article will increase our knowledge of L. (L.) amazonensis-specific adaptations to infection, parasite survival and the transmission of this Amazonian species in a new endemic area of Brazil. PMID:28091623

Comparative genomics of canine-isolated Leishmania (Leishmania) amazonensis from an endemic focus of visceral leishmaniasis in Governador Valadares, southeastern Brazil.

PubMed

Valdivia, Hugo O; Almeida, Laila V; Roatt, Bruno M; Reis-Cunha, João Luís; Pereira, Agnes Antônia Sampaio; Gontijo, Celia; Fujiwara, Ricardo Toshio; Reis, Alexandre B; Sanders, Mandy J; Cotton, James A; Bartholomeu, Daniella C

2017-01-16

Leishmaniasis is a highly diverse group of diseases caused by kinetoplastid of the genus Leishmania. These parasites are taxonomically diverse, with human pathogenic species separated into two subgenera according to their development site inside the alimentary tract of the sand fly insect vector. The disease encompasses a variable spectrum of clinical manifestations with tegumentary or visceral symptoms. Among the causative species in Brazil, Leishmania (Leishmania) amazonensis is an important etiological agent of human cutaneous leishmaniasis that accounts for more than 8% of all cases in endemic regions. L. (L.) amazonensis is generally found in the north and northeast regions of Brazil. Here, we report the first isolation of L. (L.) amazonensis from dogs with clinical manifestations of visceral leishmaniasis in Governador Valadares, an endemic focus in the southeastern Brazilian State of Minas Gerais where L. (L.) infantum is also endemic. These isolates were characterized in terms of SNPs, chromosome and gene copy number variations, confirming that they are closely related to a previously sequenced isolate obtained in 1973 from the typical Northern range of this species. The results presented in this article will increase our knowledge of L. (L.) amazonensis-specific adaptations to infection, parasite survival and the transmission of this Amazonian species in a new endemic area of Brazil.

Surveillance for antibodies to Leishmania spp. in dogs from Sri Lanka and India

USDA-ARS?s Scientific Manuscript database

The global distribution of leishmaniasis is rapidly expanding into new geographic regions. Dogs are the primary reservoir hosts for human visceral leishmaniasis (VL) caused by infection with Leishmania infantum. Natural infections with other Leishmania species can occur in dogs, but their role as re...

A novel recombinant Leishmania donovani p45, a partial coding region of methionine aminopeptidase, generates protective immunity by inducing a Th1 stimulatory response against experimental visceral leishmaniasis.

PubMed

Gupta, Reema; Kushawaha, Pramod K; Tripathi, Chandra Dev Pati; Sundar, Shyam; Dube, Anuradha

2012-05-01

The development of a vaccine against visceral leishmaniasis (VL) conferring long-lasting immunity remains a challenge. Identification and proteomic characterization of parasite proteins led to the detection of p45, a member of the methionine aminopeptidase family. To our knowledge the present study is the first known report that describes the molecular and immunological characterization of p45. Recombinant Leishmania donovani p45 (rLdp45) induced cellular responses in cured hamsters and generated Th1-type cytokines from peripheral blood mononuclear cells of cured/endemic VL patients. Immunization with rLdp45 exerted considerable prophylactic efficacy (∼85%) supported by an increase in mRNA expression of iNOS, IFN-γ, TNF-α and IL-12 and decrease in TGF-β and IL-4, indicating its potential as a vaccine candidate against VL. Copyright © 2012. Published by Elsevier Ltd.

Leishmania replication protein A-1 binds in vivo single-stranded telomeric DNA

DOE Office of Scientific and Technical Information (OSTI.GOV)

Neto, J.L. Siqueira; Instituto de Biologia, UNICAMP, Campinas, SP; Lira, C.B.B.

Replication protein A (RPA) is a highly conserved heterotrimeric single-stranded DNA-binding protein involved in different events of DNA metabolism. In yeast, subunits 1 (RPA-1) and 2 (RPA-2) work also as telomerase recruiters and, in humans, the complex unfolds G-quartet structures formed by the 3' G-rich telomeric strand. In most eukaryotes, RPA-1 and RPA-2 bind DNA using multiple OB fold domains. In trypanosomatids, including Leishmania, RPA-1 has a canonical OB fold and a truncated RFA-1 structural domain. In Leishmania amazonensis, RPA-1 alone can form a complex in vitro with the telomeric G-rich strand. In this work, we show that LaRPA-1 ismore » a nuclear protein that associates in vivo with Leishmania telomeres. We mapped the boundaries of the OB fold DNA-binding domain using deletion mutants. Since Leishmania and other trypanosomatids lack homologues of known telomere end binding proteins, our results raise questions about the function of RPA-1 in parasite telomeres.« less

Generation of Leishmania Hybrids by Whole Genomic DNA Transformation

PubMed Central

Coelho, Adriano C.; Leprohon, Philippe; Ouellette, Marc

2012-01-01

Genetic exchange is a powerful tool to study gene function in microorganisms. Here, we tested the feasibility of generating Leishmania hybrids by electroporating genomic DNA of donor cells into recipient Leishmania parasites. The donor DNA was marked with a drug resistance marker facilitating the selection of DNA transfer into the recipient cells. The transferred DNA was integrated exclusively at homologous locus and was as large as 45 kb. The independent generation of L. infantum hybrids with L. major sequences was possible for several chromosomal regions. Interfering with the mismatch repair machinery by inactivating the MSH2 gene enabled an increased efficiency of recombination between divergent sequences, hence favouring the selection of hybrids between species. Hybrids were shown to acquire the phenotype derived from the donor cells, as demonstrated for the transfer of drug resistance genes from L. major into L. infantum. The described method is a first step allowing the generation of in vitro hybrids for testing gene functions in a natural genomic context in the parasite Leishmania. PMID:23029579

Phototoxic effects of silicon bis (dimetilaminoetanoxi)-phthalocyanine (SiPc) on the viability of Leishmania major and Leishmania braziliensis promastigotes

NASA Astrophysics Data System (ADS)

Guerra Pinto, Juliana; Ferreira-Strixino, Juliana; Mittmann, Josane

2016-06-01

American cutaneous leishmaniasis (ACL) is an infectious disease caused by protozoans of the genus Leishmania. The treatment may consist of pentavalent antimonials or pentamidine and amphotericin. However, these treatments are extremely aggressive. Photodynamic antimicrobial chemotherapy (PACT) involves the same mechanism of photodynamic therapy which associates a photosensitizer with oxygen and a light source generating a photochemical reaction leading to cell death. The aim of this study was to verify the potential use of silicon bis (dimetilaminoetanoxi)-phthalocyanine (SiPc) compound in photodynamic treatment through evaluation of its phototoxic effect in promastigotes of the genus Leishmania braziliensis and Leishmania major. Treatment with SiPc was able to drastically affect the viability of the parasites as well as affect their growth and morphology, after PACT treatment. The data shown in this study allows us to conclude that SiPc is a promising photosensitizer (PS) since it does not affect parasite growth and viability in the dark. After PACT with this phthalocyanine, over 99% of parasites were killed with the higher concentration and a light dose used. These results suggest that SiPc can be used in future to treat CL, however, further studies are necessary to determine whether the PS are toxic to mononuclear phagocytic cells and epithelial cells which will also be affected by therapy when applied topically.

Natural infection of phlebotomines (Diptera: Psychodidae) by Leishmania (Leishmania) amazonensis in an area of ecotourism in Central-Western Brazil.

PubMed

Brilhante, Andreia Fernandes; Nunes, Vânia Lúcia Brandão; Kohatsu, Kleber Augusto; Galati, Eunice Aparecida Bianchi; Rocca, Maria Elizabeth Ghizzi; Ishikawa, Edna Aoba Yassui

2015-01-01

Bonito municipality, known as an area of ecoturism, in Mato Grosso do Sul state, Brazil, is also a focus of visceral and cutaneous leishmaniases, with cases registered in both human and canine populations. This study sought to investigate natural infection by flagellate forms of Leishmania in phlebotomines of the urban area of Bonito. Sand flies were collected fortnightly from October 2005 to July 2006 with modified automatic light traps installed in peridomiciles and animal shelters in the center and on the outskirts of the city. The females were dissected and their guts observed under an optical microscope. A total of 1977 specimens were captured, Lutzomyia longipalpis (88.4 %) and Bichromomyia flaviscutelata (3.0 %) being the most frequent species. Bi. flaviscutellata was found infected by flagellates that were identified as Leishmania (Leishmania) amazonensis by indirect immunofluorescence reaction, employing monoclonal antibodies and the biotin-avidin system. This is the first report of natural infection by L. amazonensis in Bi. flaviscutellata in a Brazilian urban area. As Bi. flaviscutellata is only slightly attracted by humans, the transmission of L. amazonensis in the study area may have a zoonotic character; however, the sympatric occurrence of this parasite and Lu. longipalpis should be taken into consideration by the local health authorities since this sand fly has already been found with L. amazonensis DNA in a focus of canine visceral leishmaniasis in Bonito municipality.

[Predictive factors associated with severity of asthma exacerbations].

PubMed

Atiş, Sibel; Kaplan, Eylem Sercan; Ozge, Cengiz; Bayindir, Suzan

2008-01-01

Several factors have been accused for asthma exacerbations, however, very few studies have evaluated whether different factors predict severity of asthma exacerbation. We aimed to determine the predictive factors for severity of asthma exacerbation. Retrospective analysis of data on 93 patients visited our emergency-department because of asthma exacerbation was reviewed. Hospitalization in intensive care unit and/or intubation because of asthma was accepted as the criteria for severe exacerbation. Logistic regression analysis estimated the strength of association of each variable, potentially related to severe asthmatic exacerbation, with severe/very severe as compared to mild/moderate asthmatic exacerbation. Independent variables included in the analysis were age, sex, smoking history, inhaler steroid using, compliance with medication, chronic asthma severity, presence of additional atopic diseases, prick test positivity, provocative factors, number of short-acting beta(2)-agonist using, number of visits to emergency department for asthma over one year period, previous severe exacerbation, pulmonary functions, and blood eosinophil count. 20 were severe/very severe and 73 mild/moderate asthmatic exacerbation. Frequent using of short-acting beta(2)-agonist (OR= 1.5, 95% CI= 1.08-5.3, p= 0.003), noncompliance with medication (OR= 3.6, 95% CI= 1.3-9.9, p= 0.013), previous severe asthmatic exacerbation (OR= 3.8, 95% CI= 1.48-10.01, p= 0.005) and recent admission to hospital (OR= 2.9, 95% CI= 1.07-8.09, p= 0.037) were found to be predictive factors for severe asthmatic exacerbation. Different predictive factors, in particular frequent using of short-acting beta(2)-agonist and noncompliance with medication may be associated with severe asthma exacerbations compared to milder exacerbations. This suggests different mechanisms are responsible for severity of asthma exacerbation.

Impact of Leishmania Infection on Host Macrophage Nuclear Physiology and Nucleopore Complex Integrity

PubMed Central

Isnard, Amandine; Christian, Jan G.; Kodiha, Mohamed; Stochaj, Ursula; McMaster, W. Robert; Olivier, Martin

2015-01-01

The protease GP63 is an important virulence factor of Leishmania parasites. We previously showed that GP63 reaches the perinuclear area of host macrophages and that it directly modifies nuclear translocation of the transcription factors NF-κB and AP-1. Here we describe for the first time, using molecular biology and in-depth proteomic analyses, that GP63 alters the host macrophage nuclear envelope, and impacts on nuclear processes. Our results suggest that GP63 does not appear to use a classical nuclear localization signal common between Leishmania species for import, but degrades nucleoporins, and is responsible for nuclear transport alterations. In the nucleoplasm, GP63 activity accounts for the degradation and mislocalization of proteins involved amongst others in gene expression and in translation. Collectively, our data indicates that Leishmania infection strongly affects nuclear physiology, suggesting that targeting of nuclear physiology may be a strategy beneficial for virulent Leishmania parasites. PMID:25826301

Dynamics of sterol synthesis during development of Leishmania spp. parasites to their virulent form.

PubMed

Yao, Chaoqun; Wilson, Mary E

2016-04-12

The Leishmania spp. protozoa, the causative agents of the "neglected" tropical disease leishmaniasis, are transmitted to mammals by sand fly vectors. Within the sand fly, parasites transform from amastigotes to procyclic promastigotes, followed by development of virulent (metacyclic) promastigote forms. The latter are infectious to mammalian hosts. Biochemical components localized in the parasite plasma membrane such as proteins and sterols play a pivotal role in Leishmania pathogenesis. Leishmania spp. lack the enzymes for cholesterol synthesis, and the dynamics of sterol acquisition and biosynthesis in parasite developmental stages are not understood. We hypothesized that dynamic changes in sterol composition during metacyclogenesis contribute to the virulence of metacyclic promastigotes. Sterols were extracted from logarithmic phase or metacyclic promastigotes grown in liquid culture with or without cholesterol, and analyzed qualitatively and quantitatively by gas chromatograph-mass spectrometry (GC-MS). TriTrypDB was searched for identification of genes involved in Leishmania sterol biosynthetic pathways. In total nine sterols were identified. There were dynamic changes in sterols during promastigote metacyclogenesis. Cholesterol in the culture medium affected sterol composition in different parasite stages. There were qualitative and relative quantitative differences between the sterol content of virulent versus avirulent parasite strains. A tentative sterol biosynthetic pathway in Leishmania spp. promastigotes was identified. Significant differences in sterol composition were observed between promastigote stages, and between parasites exposed to different extracellular cholesterol in the environment. These data lay the foundation for further investigating the role of sterols in the pathogenesis of Leishmania spp. infections.

Lulo cell line derived from Lutzomyia longipalpis (Diptera: Psychodidae): a novel model to assay Leishmania spp. and vector interaction.

PubMed

Côrtes, Luzia Mc; Silva, Roger Mm; Pereira, Bernardo As; Guerra, Camila; Zapata, Angela C; Bello, Felio J; Finkelstein, Léa C; Madeira, Maria F; Brazil, Reginaldo P; Côrte-Real, Suzana; Alves, Carlos R

2011-11-14

Leishmania (Vianna) braziliensis, Leishmania (Leishmania) amazonensis and Leishmania (Leishmania) chagasi are important parasites in the scenario of leishmaniasis in Brazil. During the life cycle of these parasites, the promastigote forms adhere to the midgut epithelial microvillii of phlebotomine insects to avoid being secreted along with digestive products. Lulo cells are a potential model that will help to understand the features of this adhesion phenomenon. Here, we analyze the interaction between Leishmania spp. promastigotes and Lulo cells in vitro, specifically focusing on adhesion events occurring between three Leishmania species and this cell line. Confluent monolayers of Lulo cells were incubated with promastigotes and adhesion was assessed using both light microscopy and scanning electron microscopy. The results indicate that species from the subgenera Leishmania and Viannia have great potential to adhere to Lulo cells. The highest adherence rate was observed for L. (L.) chagasi after 24 h of incubation with Lulo cells (27.3 ± 1.8% of cells with adhered promastigotes), followed by L. (L.) amazonensis (16.0 ± 0.7%) and L. (V.) braziliensis (3.0 ± 0.7%), both after 48 h. In the ultrastructural analysis, promastigote adherence was also assessed by scanning electron microscopy, showing that, for parasites from both subgenera, adhesion occurs by both the body and the flagellum. The interaction of Lulo cells with Leishmania (L.) chagasi showed the participation of cytoplasmic projections from the former closely associating the parasites with the cells. We present evidence that Lulo cells can be useful in studies of insect-parasite interactions for Leishmania species.

Lulo cell line derived from Lutzomyia longipalpis (Diptera: Psychodidae): a novel model to assay Leishmania spp. and vector interaction

PubMed Central

2011-01-01

Background Leishmania (Vianna) braziliensis, Leishmania (Leishmania) amazonensis and Leishmania (Leishmania) chagasi are important parasites in the scenario of leishmaniasis in Brazil. During the life cycle of these parasites, the promastigote forms adhere to the midgut epithelial microvillii of phlebotomine insects to avoid being secreted along with digestive products. Lulo cells are a potential model that will help to understand the features of this adhesion phenomenon. Here, we analyze the interaction between Leishmania spp. promastigotes and Lulo cells in vitro, specifically focusing on adhesion events occurring between three Leishmania species and this cell line. Methods Confluent monolayers of Lulo cells were incubated with promastigotes and adhesion was assessed using both light microscopy and scanning electron microscopy. Findings The results indicate that species from the subgenera Leishmania and Viannia have great potential to adhere to Lulo cells. The highest adherence rate was observed for L. (L.) chagasi after 24 h of incubation with Lulo cells (27.3 ± 1.8% of cells with adhered promastigotes), followed by L. (L.) amazonensis (16.0 ± 0.7%) and L. (V.) braziliensis (3.0 ± 0.7%), both after 48 h. In the ultrastructural analysis, promastigote adherence was also assessed by scanning electron microscopy, showing that, for parasites from both subgenera, adhesion occurs by both the body and the flagellum. The interaction of Lulo cells with Leishmania (L.) chagasi showed the participation of cytoplasmic projections from the former closely associating the parasites with the cells. Conclusions We present evidence that Lulo cells can be useful in studies of insect-parasite interactions for Leishmania species. PMID:22082050

Natural Infection of Phlebotomus sergenti by Leishmania tropica in Libya.

PubMed

Dokhan, Mostafa Ramadhan; Jaouadi, Kaouther; Salem, Sadok; Zenbil, Osama; Gonzalez, Jean Paul; Ben Salah, Afif; Annajar, Badreddin Bashir

2018-03-12

Cutaneous leishmaniasis (CL) is a public health concern caused by Leishmania ( L. ) major and ( L. ) tropica in Libya. Information on sandfly vectors, as well as their associated Leishmania species, is of paramount importance because vector dispersion is one of the major factors responsible for pathogen dissemination. A number of 515 sandflies (275 males and 240 females) were collected during June-November 2012 using the CDC miniature light traps from Al Rabta, northwest of Libya. Two hundred and forty unfed females were identified; Phlebotomus ( Ph. ) papatasi ( N = 97), Ph. sergenti ( N = 27), Ph. longicuspis ( N = 32), Sergentomyia ( Se. ) minuta ( N = 38), and Se. fallax ( N = 46). These flies were screened for Leishmania DNA using the polymerase chain reaction-restriction fragment length polymorphism analysis of the internal transcribed spacer 1 and sequencing. Two Ph. sergenti were found positive to L. tropica DNA. This finding should be considered for any further vector surveillance and epidemiological studies of CL in endemic areas across Libya.

On the relationship between human papilloma virus vaccine and autoimmune diseases.

PubMed

Pellegrino, Paolo; Carnovale, Carla; Pozzi, Marco; Antoniazzi, Stefania; Perrone, Valentina; Salvati, Dionigi; Gentili, Marta; Brusadelli, Tatiana; Clementi, Emilio; Radice, Sonia

2014-07-01

The human papilloma virus (HPV) vaccines were introduced to reduce the incidence of cervical cancer. The bivalent vaccine is effective against HPV-16, -18, -31, -33 and -45 while the quadrivalent vaccine is effective against HPV-16, 18, 31, 6 and 11 types. The immunisation, recommended for adolescent females, has led to high vaccine coverage in many countries. Along with the introduction of the HPV vaccines, several cases of onset or exacerbations of autoimmune diseases following the vaccine shot have been reported in the literature and pharmacovigilance databases, triggering concerns about its safety. This vaccination programme, however, has been introduced in a population that is at high risk for the onset of autoimmune diseases, making it difficult to assess the role of HPV vaccine in these cases and no conclusive studies have been reported thus far. We have thus analysed and reviewed comprehensively all case reports and studies dealing with either the onset of an autoimmune disease in vaccinated subject or the safety in patients with autoimmune diseases to define the role of the HPV vaccines in these diseases and hence its safety. A solid evidence of causal relationship was provided in few cases in the examined studies, and the risk vs. benefit of vaccination is still to be solved. The on-going vigilance for the safety of this vaccine remains thus of paramount importance. Copyright © 2014 Elsevier B.V. All rights reserved.

Imaging host-Leishmania interactions: significance in visceral leishmaniasis.

PubMed

Forestier, C-L

2013-01-01

Leishmaniasis is a neglected disease that is associated with a spectrum of clinical manifestations ranging from self-healing cutaneous lesions to fatal visceral infections, which primarily depends on the parasite species. In visceral leishmaniasis (VL), as opposed to cutaneous leishmaniasis (CL), parasites that infect host cells at the sand fly bite site have the striking ability to disseminate to visceral organs where they proliferate and persist for long periods of time. Imaging the dynamics of the host-Leishmania interaction in VL provides a powerful approach to understanding the mechanisms underlying host cell invasion, Leishmania dissemination and persistence within visceral organs and, to dissecting the immune responses to infection. Therefore, by allowing the visualization of the critical steps involved in the pathogenesis of VL, state-of-the-art microscopy technologies have the great potential to aid in the identification of better intervention strategies for this devastating disease. In this review, we emphasize the current knowledge and the potential significance of imaging technologies in understanding the infection process of visceralizing Leishmania species. Then, we discuss how application of innovative microscopy technologies to the study of VL will provide rich opportunities for investigating host-parasite interactions at a previously unexplored level and elucidating visceral disease-promoting mechanisms. © 2013 John Wiley & Sons Ltd.

Leishmania infection modulates beta-1 integrin activation and alters the kinetics of monocyte spreading over fibronectin.

PubMed

Figueira, Cláudio Pereira; Carvalhal, Djalma Gomes Ferrão; Almeida, Rafaela Andrade; Hermida, Micely d' El-Rei; Touchard, Dominique; Robert, Phillipe; Pierres, Anne; Bongrand, Pierre; dos-Santos, Washington L C

2015-08-07

Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue. In this work, we studied the early stages of bond formation between VLA4 and fibronectin, measured the kinetics of membrane alignment and the monocyte cytoplasm spreading area over a fibronectin-coated surface, and studied the expression of high affinity integrin epitope in uninfected and Leishmania-infected human monocytes. Our results show that the initial VLA4-mediated interaction of Leishmania-infected monocyte with a fibronectin-coated surface is preserved, however, the later stage, leukocyte spreading over the substrate is abrogated in Leishmania-infected cells. The median of spreading area was 72 [55-89] μm(2) for uninfected and 41 [34-51] μm(2) for Leishmania-infected monocyte. This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody. After the initial contact with the fibronectrin-coated surface, uninfected monocyte quickly spread the cytoplasm at a 15 μm(2) s(-1) ratio whilst Leishmania-infected monocytes only made small contacts at a 5.5 μm(2) s(-1) ratio. The expression of high affinity epitope by VLA4 (from 39 ± 21% to 14 ± 3%); and LFA1 (from 37 ± 32% to 18 ± 16%) molecules was reduced in Leishmania-infected monocytes. These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis.

Improving Leishmania Species Identification in Different Types of Samples from Cutaneous Lesions

PubMed Central

Cruz-Barrera, Mónica L.; Ovalle-Bracho, Clemencia; Ortegon-Vergara, Viviana; Pérez-Franco, Jairo E.

2015-01-01

The discrimination of Leishmania species from patient samples has epidemiological and clinical relevance. In this study, different gene target PCR-restriction fragment length polymorphism (RFLP) protocols were evaluated for their robustness as Leishmania species discriminators in 61 patients with cutaneous leishmaniasis. We modified the hsp70-PCR-RFLP protocol and found it to be the most reliable protocol for species identification. PMID:25609727

Development of Novel Prime-Boost Strategies Based on a Tri-Gene Fusion Recombinant L. tarentolae Vaccine against Experimental Murine Visceral Leishmaniasis

PubMed Central

Saljoughian, Noushin; Taheri, Tahereh; Zahedifard, Farnaz; Taslimi, Yasaman; Doustdari, Fatemeh; Bolhassani, Azam; Doroud, Delaram; Azizi, Hiva; Heidari, Kazem; Vasei, Mohammad; Namvar Asl, Nabiollah; Papadopoulou, Barbara; Rafati, Sima

2013-01-01

Visceral leishmaniasis (VL) is a vector-borne disease affecting humans and domestic animals that constitutes a serious public health problem in many countries. Although many antigens have been examined so far as protein- or DNA-based vaccines, none of them conferred complete long-term protection. The use of the lizard non-pathogenic to humans Leishmania (L.) tarentolae species as a live vaccine vector to deliver specific Leishmania antigens is a recent approach that needs to be explored further. In this study, we evaluated the effectiveness of live vaccination in protecting BALB/c mice against L. infantum infection using prime-boost regimens, namely Live/Live and DNA/Live. As a live vaccine, we used recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinases (CPA and CPB without its unusual C-terminal extension (CPB-CTE)) as a tri-fusion gene. For DNA priming, the tri-fusion gene was encoded in pcDNA formulated with cationic solid lipid nanoparticles (cSLN) acting as an adjuvant. At different time points post-challenge, parasite burden and histopathological changes as well as humoral and cellular immune responses were assessed. Our results showed that immunization with both prime-boost A2-CPA-CPB-CTE-recombinant L. tarentolae protects BALB/c mice against L. infantum challenge. This protective immunity is associated with a Th1-type immune response due to high levels of IFN-γ production prior and after challenge and with lower levels of IL-10 production after challenge, leading to a significantly higher IFN-γ/IL-10 ratio compared to the control groups. Moreover, this immunization elicited high IgG1 and IgG2a humoral immune responses. Protection in mice was also correlated with a high nitric oxide production and low parasite burden. Altogether, these results indicate the promise of the A2-CPA-CPB-CTE-recombinant L. tarentolae as a safe live vaccine candidate against VL. PMID:23638195

Exposure to mixed asymptomatic infections with Trypanosoma cruzi, Leishmania braziliensis and Leishmania chagasi in the human population of the greater Amazon.

PubMed

Mendes, Daniella G; Lauria-Pires, Liana; Nitz, Nadjar; Lozzi, Silene P; Nascimento, Rubens J; Monteiro, Pedro S; Rebelo, Manuel M; Rosa, Ana de Cássia; Santana, Jaime M; Teixeira, Antonio R L

2007-05-01

Lack of conservation of the Amazon tropical rainforest has imposed severe threats to its human population living in newly settled villages, resulting in outbreaks of some infectious diseases. We conducted a seroepidemiological survey of 1100 inhabitants of 15 villages of Paço do Lumiar County, Brazil. Thirty-five (3%) individuals had been exposed to Trypanosoma cruzi (Tc), 41 (4%) to Leishmania braziliensis (Lb) and 50 (4.5%) to Leishmania chagasi (Lc) infections. Also, 35 cases had antibodies that were cross-reactive against the heterologous kinetoplastid antigens. Amongst these, the Western blot assays revealed that 11 (1%) had Tc and Lb, that seven (0.6%) had Lc and Tc, and that 17 (1.6%) had Lb and Lc infections. All of these cases of exposures to mixed infections with Leishmania sp, and eight of 11 cases of Tc and Lb were confirmed by specific PCR assays and Southern hybridizations. Two cases had triple infections. We consider these asymptomatic cases showing phenotype and genotype markers consistent with mixed infections by two or more kinetoplastid flagellates a high risk factor for association with Psychodidae and Triatominae vectors blood feeding and transmitting these protozoa infections. This is the first publication showing human exposure to mixed asymptomatic kinetoplastid infections in the Amazon.

A respiratory syncytial virus (RSV) vaccine based on parainfluenza virus 5 (PIV5)

PubMed Central

Phan, Shannon I.; Chen, Zhenhai; Xu, Pei; Li, Zhuo; Gao, Xiudan; Foster, Stephanie L.; Teng, Michael N.; Tripp, Ralph A.; Sakamoto, Kaori; He, Biao

2014-01-01

Human respiratory syncytial virus (RSV) is a leading cause of severe respiratory disease and hospitalizations in infants and young children. It also causes significant morbidity and mortality in elderly and immune compromised individuals. No licensed vaccine currently exists. Parainfluenza virus 5 (PIV5) is a paramyxovirus that causes no known human illness and has been used as a platform for vector-based vaccine development. To evaluate the efficacy of PIV5 as a RSV vaccine vector, we generated two recombinant PIV5 viruses - one expressing the fusion (F) protein and the other expressing the attachment glycoprotein (G) of RSV strain A2 (RSV A2). The vaccine strains were used separately for single-dose vaccinations in BALB/c mice. The results showed that both vaccines induced RSV antigen-specific antibody responses, with IgG2a/IgG1 ratios similar to those seen in wild-type RSV A2 infection. After challenging the vaccinated mice with RSV A2, histopathology of lung sections showed that the vaccines did not exacerbate lung lesions relative to RSV A2-immunized mice. Importantly, both F and G vaccines induced protective immunity. Therefore, PIV5 presents an attractive platform for vector-based vaccines against RSV infection. PMID:24717150

Comparison of two commercial vaccines against visceral leishmaniasis in dogs from endemic areas: IgG, and subclasses, parasitism, and parasite transmission by xenodiagnosis.

PubMed

Fernandes, Consuelo Barreto; Junior, Jairo Torres Magalhães; de Jesus, Clauceane; Souza, Bárbara Maria Paraná da Silva; Larangeira, Daniela Farias; Fraga, Deborah Bittencourt Mothé; Tavares Veras, Patricia Sampaio; Barrouin-Melo, Stella Maria

2014-03-05

The incidence of zoonotic canine visceral leishmaniasis (CVL) would decrease if dogs were effectively vaccinated; however, additional data on the efficacy of canine vaccines are required for their approved preventative use. To prospectively evaluate vaccination outcomes using two products commercially available in Brazil, with respect to adverse reactions (reactogenicity), humoral response, disease signs, parasitism, and parasite infectiousness in naturally exposed pet dogs in an endemic area of visceral leishmaniasis (VL). From 2010 to 2012, healthy dogs were vaccinated with Leishmune(®) (50 animals) or Leish-Tec(®) (50 animals). Each dog was examined to identify clinical signs during peri- and post-vaccination procedures every 2 months for 11 months to identify the presence of parasites or parasite DNA in splenic samples using culturing or PCR, respectively. Levels of anti-Leishmania IgG, IgG1, and IgG2 were quantified in sera by ELISA and infectiousness was assessed by xenodiagnosis. Adverse effects occurred in 2.2% (1/45) and 13.0% (6/46) of the animals in the Leishmune(®) and Leish-Tec(®) groups, respectively. IgG levels peaked on the 21st day following the first dose of Leishmune(®) and on the 21st day after the second dose of Leish-Tec(®). The final seropositivity rate for IgG was 32.5% (13/40) and 30.9% (13/42) in the Leishmune(®) and Leish-Tec(®) groups, respectively. The Leishmune(®) group presented higher levels of IgG1 and IgG2 compared to the Leish-Tec(®) group (p<0.001), and ELISA reactivity in both vaccinated groups was significantly lower (p<0.001) than in infected positive control dogs. Parasitism was observed in 12.2% (5/41) of the Leishmune(®) group, and 7.9% (3/38) of the Leish-Tec(®) group, with xenodiagnostic transmission rates of Leishmania to Lutzomyia longipalpis of 5.1% (2/39), and 5.4% (2/37), respectively. No significant differences were observed in dogs vaccinated with Leishmune(®) or Leish-Tec(®), with respect to LVC

Wild and synanthropic reservoirs of Leishmania species in the Americas

PubMed Central

Roque, André Luiz R.; Jansen, Ana Maria

2014-01-01

The definition of a reservoir has changed significantly in the last century, making it necessary to study zoonosis from a broader perspective. One important example is that of Leishmania, zoonotic multi-host parasites maintained by several mammal species in nature. The magnitude of the health problem represented by leishmaniasis combined with the complexity of its epidemiology make it necessary to clarify all of the links in transmission net, including non-human mammalian hosts, to develop effective control strategies. Although some studies have described dozens of species infected with these parasites, only a minority have related their findings to the ecological scenario to indicate a possible role of that host in parasite maintenance and transmission. Currently, it is accepted that a reservoir may be one or a complex of species responsible for maintaining the parasite in nature. A reservoir system should be considered unique on a given spatiotemporal scale. In fact, the transmission of Leishmania species in the wild still represents an complex enzootic “puzzle”, as several links have not been identified. This review presents the mammalian species known to be infected with Leishmania spp. in the Americas, highlighting those that are able to maintain and act as a source of the parasite in nature (and are thus considered potential reservoirs). These host/reservoirs are presented separately in each of seven mammal orders – Marsupialia, Cingulata, Pilosa, Rodentia, Primata, Carnivora, and Chiroptera – responsible for maintaining Leishmania species in the wild. PMID:25426421

Attenuation of Leishmania infantum chagasi Metacyclic Promastigotes by Sterol Depletion

PubMed Central

Gaur Dixit, Upasna; Barker, Jason H.; Teesch, Lynn M.; Love-Homan, Laurie; Donelson, John E.; Wilson, Mary E.

2013-01-01

The infectious metacyclic promastigotes of Leishmania protozoa establish infection in a mammalian host after they are deposited into the dermis by a sand fly vector. Several Leishmania virulence factors promote infection, including the glycosylphosphatidylinositol membrane-anchored major surface protease (MSP). Metacyclic Leishmania infantum chagasi promastigotes were treated with methyl-beta-cyclodextrin (MβCD), a sterol-chelating reagent, causing a 3-fold reduction in total cellular sterols as well as enhancing MSP release without affecting parasite viability in vitro. MβCD-treated promastigotes were more susceptible to complement-mediated lysis than untreated controls and reduced the parasite load 3-fold when inoculated into BALB/c mice. Paradoxically, MβCD-treated promastigotes caused a higher initial in vitro infection rate in human or murine macrophages than untreated controls, although their intracellular multiplication was hindered upon infection establishment. There was a corresponding larger amount of covalently bound C3b than iC3b on the parasite surfaces of MβCD-treated promastigotes exposed to healthy human serum in vitro, as well as loss of MSP, a protease that enhances C3b cleavage to iC3b. Mass spectrometry showed that MβCD promotes the release of proteins into the extracellular medium, including both MSP and MSP-like protein (MLP), from virulent metacyclic promastigotes. These data support the hypothesis that plasma membrane sterols are important for the virulence of Leishmania protozoa at least in part through retention of membrane virulence proteins. PMID:23630964

C-terminal domain deletion enhances the protective activity of cpa/cpb loaded solid lipid nanoparticles against Leishmania major in BALB/c mice.

PubMed

Doroud, Delaram; Zahedifard, Farnaz; Vatanara, Alireza; Taslimi, Yasaman; Vahabpour, Rouholah; Torkashvand, Fatemeh; Vaziri, Behrooz; Rouholamini Najafabadi, Abdolhossein; Rafati, Sima

2011-07-01

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. 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. 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

Serological and molecular survey of Leishmania infection in dogs from Luanda, Angola

PubMed Central

2014-01-01

Background Canine leishmaniosis (CanL) due to Leishmania infantum is a global zoonosis endemic in more than 70 countries in Europe, North Africa, Asia and America; however, data on this infection is scarce from southern Africa. The aim of this study was to survey dogs in Luanda, Angola, for Leishmania infection. Findings One hundred-and-three dogs presented to a veterinary medical centre in Luanda were serologically and molecularly assessed for Leishmania with the direct agglutination test (DAT) and polymerase chain reaction (PCR). Two dogs were seropositive, with DAT titres of 800 and ≥6400; the latter was also found to be PCR-positive and confirmed to be infected with L. infantum by DNA sequence analysis. No other dog was found to be PCR-positive. The first dog had been imported from Portugal, but the latter had never left Angola (neither had its parents), strongly suggesting an autochthonous infection. Conclusions Although other cases of CanL have previously been described in the country, this is the first reported study of canine Leishmania infection at the population level, as well as the first report on the molecular characterization of L. infantum in dogs from Angola. PMID:24655415

Characterization of monomeric DNA-binding protein Histone H1 in Leishmania braziliensis.

PubMed

Carmelo, Emma; González, Gloria; Cruz, Teresa; Osuna, Antonio; Hernández, Mariano; Valladares, Basilio

2011-08-01

Histone H1 in Leishmania presents relevant differences compared to higher eukaryote counterparts, such as the lack of a DNA-binding central globular domain. Despite that, it is apparently fully functional since its differential expression levels have been related to changes in chromatin condensation and infectivity, among other features. The localization and the aggregation state of L. braziliensis H1 has been determined by immunolocalization, mass spectrometry, cross-linking and electrophoretic mobility shift assays. Analysis of H1 sequences from the Leishmania Genome Database revealed that our protein is included in a very divergent group of histones H1 that is present only in L. braziliensis. An antibody raised against recombinant L. braziliensis H1 recognized specifically that protein by immunoblot in L. braziliensis extracts, but not in other Leishmania species, a consequence of the sequence divergences observed among Leishmania species. Mass spectrometry analysis and in vitro DNA-binding experiments have also proven that L. braziliensis H1 is monomeric in solution, but oligomerizes upon binding to DNA. Finally, despite the lack of a globular domain, L. braziliensis H1 is able to form complexes with DNA in vitro, with higher affinity for supercoiled compared to linear DNA.

Photo-induced Leishmania DNA degradation by silver-doped zinc oxide nanoparticle: an in-vitro approach.

PubMed

Nadhman, Akhtar; Sirajuddin, Muhammad; Nazir, Samina; Yasinzai, Masoom

2016-06-01

Recently, the authors reported newly synthesised polyethylene glycol (PEG)ylated silver (9%)-doped zinc oxide nanoparticle (doped semiconductor nanoparticle (DSN)) which has high potency for killing Leishmania tropica by producing reactive oxygen species on exposure to sunlight. The current report is focused on Leishmania DNA interaction and damage caused by the DSN. Here, we showed that the damage to Leishmania DNA was indirect, as the DSN was unable to interact with the DNA in intact Leishmania cell, indicating the incapability of PEGylated DSN to cross the nucleus barrier. The DNA damage was the result of high production of singlet oxygen on exposure to sunlight. The DNA damage was successfully prevented by singlet oxygen scavenger (sodium azide) confirming involvement of the highly energetic singlet oxygen in the DNA degradation process.

Molecular and Serological Evidence of Leishmania Infection in Stray Dogs from Visceral Leishmaniasis-Endemic Areas of Bangladesh.

PubMed

Akter, Shirin; Alam, Mohammad Zahangir; Nakao, Ryo; Yasin, Golam; Kato, Hirotomo; Katakura, Ken

2016-10-05

Visceral leishmaniasis (VL), or kala-azar, is mainly caused by two closely related Leishmania species, Leishmania infantum and Leishmania donovani Leishmania infantum is responsible for zoonotic VL, with dogs as the main reservoir host in the Mediterranean, the Middle East, Asia, and South America. In the Indian subcontinent, VL is caused by L. donovani and is considered anthroponotic, although the only known vector, the sand fly, is zoophilic in nature. The role of domestic and stray dogs in VL transmission is still unclear in this area. We screened 50 stray dogs from VL-endemic areas of Bangladesh for serological and molecular evidence of Leishmania infection. We detected anti-Leishmania antibodies in six (12%) dog serum samples using rK39 immunochromatographic tests. We observed Leishmania kinetoplast DNA in 10 (20%) buffy coat DNA samples by real-time polymerase chain reaction (PCR), five of which were positive based on internal transcribed spacer 1-PCR. A sequencing analysis of the amplified products confirmed that the parasitic DNA was derived from L. donovani Our findings support the hypothesis that stray dogs are an animal reservoir for L. donovani in this endemic region. Further studies are required to determine the precise role of dogs in the epidemiology of VL in Bangladesh. © The American Society of Tropical Medicine and Hygiene.

An agent-based model for Leishmania major infection

NASA Astrophysics Data System (ADS)

Dancik, Garrett M.; Jones, Douglas E.; Dorman, Karin S.

Leishmania are protozoan parasites transmitted by bites of infected sandflies. Over 20 species of Leishmania, endemic in 88 countries, are capable of causing human disease. Disease is either cutaneous, where skin ulcers occur on exposed surfaces of the body, or visceral, with near certain mortality if untreated. C3HeB/FeJ mice are resistant to L. major, but develop chronic cutaneous lesions when infected with another species L. amazonensis. The well-characterized mechanism of resistance to L. major depends on a CD4+ Thl immune response, macrophage activation, and elimination of the parasite [Sacks 2002]. The factors that account for host susceptibility to L. Amazonensis, however, are not completely understood, despite being generally attributed to a weakened Th1 response [Vanloubbeck 2004].

An agent-based model for Leishmania major infection

NASA Astrophysics Data System (ADS)

Dancik, Garrett M.; Jones, Douglas E.; Dorman, Karin S.

Leishmania are protozoan parasites transmitted by bites of infected sandflies. Over 20 species of Leishmania, endemic in 88 countries, are capable of causing human disease. Disease is either cutaneous, where skin ulcers occur on exposed surfaces of the body, or visceral, with near certain mortality if left untreated. C3HeB/FeJ mice are resistant to L. major, but develop chronic cutaneous lesions when infected with another species L. amazonensis. The well-characterized mechanism of resistance to L. major depends on a CD4+ Thl immune response, macrophage activation, and elimination of the parasite [Sacks 2002]. The factors that account for host susceptibility to L. Amazonensis, however, are not completely understood, despite being generally attributed to a weakened Th1 response [Vanloubbeck 2004].

Detection of Leishmania parasites in the blood of patients with isolated cutaneous leishmaniasis.

PubMed

Nakkash-Chmaisse, Hania; Makki, Raja; Nahhas, Georges; Knio, Khouzama; Nuwayri-Salti, Nuha

2011-07-01

The consequences of the spread of Leishmania parasites to the blood from lesions in patients with cutaneous leishmaniasis are numerous. To assess the magnitude of this invasion we conducted the present study on patients referred to the American University of Beirut Medical Center for cutaneous leishmaniasis. Patients referred for the management of cutaneous leishmaniasis were included in the study. Skin and blood cultures for Leishmania were taken from these patients. One hundred sixty-two patients were proven to have cutaneous leishmaniasis by pathology; 52% were males and 44% females (gender information was missing for 4%). Patient age ranged from 5 months to 70 years. None of the patients had received treatment for Leishmania. We obtained parasite isolates from 85 patients (52.5%), proven by cultures from skin and blood/blood components. Interestingly, the parasite was isolated in the blood and blood components of 50 patients (30.9%). Isoenzyme analysis confirmed the fact that the organisms in blood and skin were the same; from the 28 isolates that were positive in both skin and blood, eight isolates were Leishmania major and two were Leishmania tropica. The remaining isolates, whether positive in both blood and skin or in either of these tissues, skin or blood and its products, were Leishmania infantum sensu lato. In the current study, the detection rate of parasites in the blood of patients with cutaneous leishmaniasis was high. This illustrates the invasive characteristic of the parasite that has escaped the skin. Testing should be considered in areas other than Lebanon, especially around the Mediterranean basin. Whether these findings support the administration of systemic treatment for cutaneous leishmaniasis or not needs to be confirmed in larger prospective studies. Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.

Differential Subcellular Localization of Leishmania Alba-Domain Proteins throughout the Parasite Development.

PubMed

Dupé, Aurélien; Dumas, Carole; Papadopoulou, Barbara

2015-01-01

Alba-domain proteins are RNA-binding proteins found in archaea and eukaryotes and recently studied in protozoan parasites where they play a role in the regulation of virulence factors and stage-specific proteins. This work describes in silico structural characterization, cellular localization and biochemical analyses of Alba-domain proteins in Leishmania infantum. We show that in contrast to other protozoa, Leishmania have two Alba-domain proteins, LiAlba1 and LiAlba3, representative of the Rpp20- and the Rpp25-like eukaryotic subfamilies, respectively, which share several sequence and structural similarities but also important differences with orthologs in other protozoa, especially in sequences targeted for post-translational modifications. LiAlba1 and LiAlba3 proteins form a complex interacting with other RNA-binding proteins, ribosomal subunits, and translation factors as supported by co-immunoprecipitation and sucrose gradient sedimentation analysis. A higher co-sedimentation of Alba proteins with ribosomal subunits was seen upon conditions of decreased translation, suggesting a role of these proteins in translational repression. The Leishmania Alba-domain proteins display differential cellular localization throughout the parasite development. In the insect promastigote stage, Alba proteins co-localize predominantly to the cytoplasm but they translocate to the nucleolus and the flagellum upon amastigote differentiation in the mammalian host and are found back to the cytoplasm once amastigote differentiation is completed. Heat-shock, a major signal of amastigote differentiation, triggers Alba translocation to the nucleolus and the flagellum. Purification of the Leishmania flagellum confirmed LiAlba3 enrichment in this organelle during amastigote differentiation. Moreover, partial characterization of the Leishmania flagellum proteome of promastigotes and differentiating amastigotes revealed the presence of other RNA-binding proteins, as well as differences in

Differential Subcellular Localization of Leishmania Alba-Domain Proteins throughout the Parasite Development

PubMed Central

Dupé, Aurélien; Dumas, Carole; Papadopoulou, Barbara

2015-01-01

Alba-domain proteins are RNA-binding proteins found in archaea and eukaryotes and recently studied in protozoan parasites where they play a role in the regulation of virulence factors and stage-specific proteins. This work describes in silico structural characterization, cellular localization and biochemical analyses of Alba-domain proteins in Leishmania infantum. We show that in contrast to other protozoa, Leishmania have two Alba-domain proteins, LiAlba1 and LiAlba3, representative of the Rpp20- and the Rpp25-like eukaryotic subfamilies, respectively, which share several sequence and structural similarities but also important differences with orthologs in other protozoa, especially in sequences targeted for post-translational modifications. LiAlba1 and LiAlba3 proteins form a complex interacting with other RNA-binding proteins, ribosomal subunits, and translation factors as supported by co-immunoprecipitation and sucrose gradient sedimentation analysis. A higher co-sedimentation of Alba proteins with ribosomal subunits was seen upon conditions of decreased translation, suggesting a role of these proteins in translational repression. The Leishmania Alba-domain proteins display differential cellular localization throughout the parasite development. In the insect promastigote stage, Alba proteins co-localize predominantly to the cytoplasm but they translocate to the nucleolus and the flagellum upon amastigote differentiation in the mammalian host and are found back to the cytoplasm once amastigote differentiation is completed. Heat-shock, a major signal of amastigote differentiation, triggers Alba translocation to the nucleolus and the flagellum. Purification of the Leishmania flagellum confirmed LiAlba3 enrichment in this organelle during amastigote differentiation. Moreover, partial characterization of the Leishmania flagellum proteome of promastigotes and differentiating amastigotes revealed the presence of other RNA-binding proteins, as well as differences in

Dual Transcriptome Profiling of Leishmania-Infected Human Macrophages Reveals Distinct Reprogramming Signatures.

PubMed

Fernandes, Maria Cecilia; Dillon, Laura A L; Belew, Ashton Trey; Bravo, Hector Corrada; Mosser, David M; El-Sayed, Najib M

2016-05-10

Macrophages are mononuclear phagocytes that constitute a first line of defense against pathogens. While lethal to many microbes, they are the primary host cells of Leishmania spp. parasites, the obligate intracellular pathogens that cause leishmaniasis. We conducted transcriptomic profiling of two Leishmania species and the human macrophage over the course of intracellular infection by using high-throughput RNA sequencing to characterize the global gene expression changes and reprogramming events that underlie the interactions between the pathogen and its host. A systematic exclusion of the generic effects of large-particle phagocytosis revealed a vigorous, parasite-specific response of the human macrophage early in the infection that was greatly tempered at later time points. An analogous temporal expression pattern was observed with the parasite, suggesting that much of the reprogramming that occurs as parasites transform into intracellular forms generally stabilizes shortly after entry. Following that, the parasite establishes an intracellular niche within macrophages, with minimal communication between the parasite and the host cell later during the infection. No significant difference was observed between parasite species transcriptomes or in the transcriptional response of macrophages infected with each species. Our comparative analysis of gene expression changes that occur as mouse and human macrophages are infected by Leishmania spp. points toward a general signature of the Leishmania-macrophage infectome. Little is known about the transcriptional changes that occur within mammalian cells harboring intracellular pathogens. This study characterizes the gene expression signatures of Leishmania spp. parasites and the coordinated response of infected human macrophages as the pathogen enters and persists within them. After accounting for the generic effects of large-particle phagocytosis, we observed a parasite-specific response of the human macrophages early in

Murine Visceral Leishmaniasis: IgM and Polyclonal B-Cell Activation Lead to Disease Exacerbation

PubMed Central

Deak, Eszter; Jayakumar, Asha; Wing Cho, Ka; Goldsmith-Pestana, Karen; Dondji, Blaise; Lambris, John D.; McMahon-Pratt, Diane

2010-01-01

In visceral leishmaniasis, the draining lymph node (DLN) is the initial site for colonization and establishment of infection after intradermal transmission by the sand fly vector; however, little is known about the developing immune response within this site. Using an intradermal infection model, which allows for parasite visceralization, we have examined the ongoing immune responses in the DLN of BALB/c mice infected with L. infantum. Although not unexpected, at early times post-infection there is a marked B cell expansion in the DLN, which persists throughout infection. However, the characteristics of this response were of interest; as early as day 7 post-infection, polyclonal antibodies (TNP, OVA, chromatin) were observed and the levels appeared comparable to the specific anti-leishmania response. Although B-cell-deficient JHD BALB/c mice are relatively resistant to infection, neither B-cell-derived IL-10 nor B-cell antigen presentation appear to be primarily responsible for the elevated parasitemia. However, passive transfer and reconstitution of JHD BALB/c with secretory immunoglobulins, (IgM or IgG; specific or non-specific immune complexes) results in increased susceptibility to L. infantum infection. Further, JHD BALB/c mice transgenetically reconstituted to secrete IgM demonstrated exacerbated disease in comparison to wild type BALB/c mice as early as 2 days post-infection. Evidence suggests that complement activation (generation of C5a) and signaling via the C5aR (CD88) is related to the disease exacerbation caused by IgM rather than cytokine levels (IL-10 or IFN-γ). Overall these studies indicate that polyclonal B cell activation, which is known to be associated with human visceral leishmaniasis, is an early and intrinsic characteristic of disease and may represent a target for therapeutic intervention. PMID:20213734

Leishmania infection modulates beta-1 integrin activation and alters the kinetics of monocyte spreading over fibronectin

PubMed Central

Figueira, Cláudio Pereira; Carvalhal, Djalma Gomes Ferrão; Almeida, Rafaela Andrade; Hermida, Micely d’ El-Rei; Touchard, Dominique; Robert, Phillipe; Pierres, Anne; Bongrand, Pierre; dos-Santos, Washington LC

2015-01-01

Contact with Leishmania leads to a decreases in mononuclear phagocyte adherence to connective tissue. In this work, we studied the early stages of bond formation between VLA4 and fibronectin, measured the kinetics of membrane alignment and the monocyte cytoplasm spreading area over a fibronectin-coated surface, and studied the expression of high affinity integrin epitope in uninfected and Leishmania-infected human monocytes. Our results show that the initial VLA4-mediated interaction of Leishmania-infected monocyte with a fibronectin-coated surface is preserved, however, the later stage, leukocyte spreading over the substrate is abrogated in Leishmania-infected cells. The median of spreading area was 72 [55–89] μm2 for uninfected and 41 [34–51] μm2 for Leishmania-infected monocyte. This cytoplasm spread was inhibited using an anti-VLA4 blocking antibody. After the initial contact with the fibronectrin-coated surface, uninfected monocyte quickly spread the cytoplasm at a 15 μm2 s−1 ratio whilst Leishmania-infected monocytes only made small contacts at a 5.5 μm2 s−1 ratio. The expression of high affinity epitope by VLA4 (from 39 ± 21% to 14 ± 3%); and LFA1 (from 37 ± 32% to 18 ± 16%) molecules was reduced in Leishmania-infected monocytes. These changes in phagocyte function may be important for parasite dissemination and distribution of lesions in leishmaniasis. PMID:26249106

An overview on Leishmania (Mundinia) enriettii: biology, immunopathology, LRV and extracellular vesicles during the host-parasite interaction.

PubMed

Paranaiba, Larissa F; Pinheiro, Lucélia J; Macedo, Diego H; Menezes-Neto, Armando; Torrecilhas, Ana C; Tafuri, Wagner L; Soares, Rodrigo P

2017-12-10

One of the Leishmania species known to be non-infective to humans is Leishmania (Mundinia) enriettii whose vertebrate host is the guinea pig Cavia porcellus. It is a good model for cutaneous leishmaniasis, chemotherapeutic and molecular studies. In the last years, an increased interest has emerged concerning the L. (Mundinia) subgenus after the finding of Leishmania (M.) macropodum in Australia and with the description of other new/putative species such as L. (M.) martiniquensis and 'L. (M.) siamensis'. This review focused on histopathology, glycoconjugates and innate immunity. The presence of Leishmania RNA virus and shedding of extracellular vesicles by the parasite were also evaluated.

Canine cutaneous leishmaniasis caused by neotropical Leishmania infantum despite of systemic disease: A case report.

PubMed

Cavalcanti, Amanda; Lobo, Rogério; Cupolillo, Elisa; Bustamante, Fábio; Porrozzi, Renato

2012-12-01

Visceral leishmaniasis is an anthropozoonosis caused by a protozoan Leishmania infantum (syn. Leishmania chagasi). Here, we report a typical case of canine cutaneous leishmaniasis due to L. infantum infection without any other systemic symptom in one dog in the city of Rio de Janeiro, Brazil. A mongrel female dog was admitted in a veterinary clinic with reports of chronic wounds in the body. Physical examination revealed erosive lesions in the limbs, nasal ulcers, presence of ectoparasites and seborrheic dermatitis. Blood samples and fragments of healthy and injured skin were collected. The complete hemogram revealed aregenerative normocytic normochromic anemia and erythrocyte rouleaux, and biochemical analysis revealed normal renal and hepatic functions. Cytology of the muzzle and skin lesions suggested pyogranulomatous inflammatory process. The histopathology of a skin fragment was performed and revealed suspicion of protozoa accompanied by necrotizing dermatitis. The diagnosis of leishmaniasis was accomplished by positive serology, isolation of Leishmania from the skin lesion, and also by molecular test (PCR targeting the conserved region of Leishmania kDNA). Culture was positive for damaged skin samples. PCR targeting a fragment of Leishmania hsp70 gene was performed employing DNA extracted from damaged skin. RFLP of the amplified hsp70 fragment identified the parasite as L. infantum, instead of Leishmania braziliensis, the main agent of cutaneous leishmaniasis in Rio de Janeiro. Characterization of isolated promastigotes by five different enzymatic systems confirmed the species identification of the etiological agent. Serology was positive by ELISA and rapid test. This case warns to the suspicion of viscerotropic Leishmania in cases of chronic skin lesions and brings the discussion of the mechanisms involved in the parasite tissue tropism. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

A Vaccine Therapy for Canine Visceral Leishmaniasis Promoted Significant Improvement of Clinical and Immune Status with Reduction in Parasite Burden

PubMed Central

Roatt, Bruno Mendes; Aguiar-Soares, Rodrigo Dian de Oliveira; Reis, Levi Eduardo Soares; Cardoso, Jamille Mirelle de Oliveira; Mathias, Fernando Augusto Siqueira; de Brito, Rory Cristiane Fortes; da Silva, Sydnei Magno; Gontijo, Nelder De Figueiredo; Ferreira, Sidney de Almeida; Valenzuela, Jesus G.; Corrêa-Oliveira, Rodrigo; Giunchetti, Rodolfo Cordeiro; Reis, Alexandre Barbosa

2017-01-01

Herein, we evaluated the treatment strategy employing a therapeutic heterologous vaccine composed of antigens of Leishmania braziliensis associated with MPL adjuvant (LBMPL vaccine) for visceral leishmaniasis (VL) in symptomatic dogs naturally infected by Leishmania infantum. Sixteen dogs received immunotherapy with MPL adjuvant (n = 6) or with a vaccine composed of antigens of L. braziliensis associated with MPL (LBMPL vaccine therapy, n = 10). Dogs were submitted to an immunotherapeutic scheme consisting of 3 series composed of 10 subcutaneous doses with 10-day interval between each series. The animals were evaluated before (T0) and 90 days after treatment (T90) for their biochemical/hematological, immunological, clinical, and parasitological variables. Our major results showed that the vaccine therapy with LBMPL was able to restore and normalize main biochemical (urea, AST, ALP, and bilirubin) and hematological (erythrocytes, hemoglobin, hematocrit, and platelets) parameters. In addition, in an ex vivo analysis using flow cytometry, dogs treated with LBMPL vaccine showed increased CD3+ T lymphocytes and their subpopulations (TCD4+ and TCD8+), reduction of CD21+ B lymphocytes, increased NK cells (CD5−CD16+) and CD14+ monocytes. Under in vitro conditions, the animals developed a strong antigen-specific lymphoproliferation mainly by TCD4+ and TCD8+ cells; increasing in both TCD4+IFN-γ+ and TCD8+IFN-γ+ as well as reduction of TCD4+IL-4+ and TCD8+IL-4+ lymphocytes with an increased production of TNF-α and reduced levels of IL-10. Concerning the clinical signs of canine visceral leishmaniasis, the animals showed an important reduction in the number and intensity of the disease signs; increase body weight as well as reduction of splenomegaly. In addition, the LBMPL immunotherapy also promoted a reduction in parasite burden assessed by real-time PCR. In the bone marrow, we observed seven times less parasites in LBMPL animals compared with MPL group. The

Leishmania and its quest for iron: An update and overview.

PubMed

Zaidi, Amir; Singh, Krishn Pratap; Ali, Vahab

2017-01-01

Parasites of genus Leishmania are the causative agents of complex neglected diseases called leishmaniasis and continue to be a significant health concern globally. Iron is a vital nutritional requirement for virtually all organisms, including pathogenic trypanosomatid parasites, and plays a crucial role in many facets of cellular metabolism as a cofactor of several enzymes. Iron acquisition is essential for the survival of parasites. Yet parasites are also vulnerable to the toxicity of iron and reactive oxygen species. The aim of this review is to provide an update on the current knowledge about iron acquisition and usage by Leishmania species. We have also discussed about host strategy to modulate iron availability and the strategies deployed by Leishmania parasites to overcome iron withholding defences and thus favour parasite growth within host macrophages. Since iron plays central roles in the host's response and parasite metabolism, a comprehensive understanding of the iron metabolism is beneficial to identify potential viable therapeutic opportunities against leishmaniasis. Copyright © 2016 Elsevier B.V. All rights reserved.

Leishmania (Viannia) Infection in the Domestic Dog in Chaparral, Colombia

PubMed Central

Santaella, Julián; Ocampo, Clara B.; Saravia, Nancy G.; Méndez, Fabián; Góngora, Rafael; Gomez, Maria Adelaida; Munstermann, Leonard E.; Quinnell, Rupert J.

2011-01-01

Peridomestic transmission of American cutaneous leishmaniasis is increasingly reported and dogs may be a reservoir of Leishmania (Viannia) in this setting. We investigated the prevalence of infection in dogs in Chaparral County, Colombia, the focus of an epidemic of human cutaneous leishmaniasis caused by Leishmania (Viannia) guyanensis. Two (0.72%) of 279 dogs had lesions typical of cutaneous leishmaniasis that were biopsy positive by kinetoplast DNA polymerase chain reaction–Southern blotting. Seroprevalence was 2.2% (6 of 279) by enzyme-linked immunosorbent assay. Buffy coat and ear skin biopsy specimens were positive by polymerase chain reaction–Southern blotting in 7.3% (10 of 137) and 11.4% (12 of 105) of dogs, respectively. Overall 20% of dogs (21 of 105) showed positive results for one or more tests. Amplification and sequencing of the Leishmania 7SL RNA gene identified L. guyanensis in one dog and L. braziliensis in two dogs. No association was identified between the risk factors evaluated and canine infection. Dogs may contribute to transmission but their role in this focus appears to be limited. PMID:21540374

The case test-negative design for studies of the effectiveness of influenza vaccine in inpatient settings.

PubMed

Foppa, Ivo M; Ferdinands, Jill M; Chaves, Sandra S; Haber, Michael J; Reynolds, Sue B; Flannery, Brendan; Fry, Alicia M

2016-12-01

The test-negative design (TND) to evaluate influenza vaccine effectiveness is based on patients seeking care for acute respiratory infection, with those who test positive for influenza as cases and the test-negatives serving as controls. This design has not been validated for the inpatient setting where selection bias might be different from an outpatient setting. We derived mathematical expressions for vaccine effectiveness (VE) against laboratory-confirmed influenza hospitalizations and used numerical simulations to verify theoretical results exploring expected biases under various scenarios. We explored meaningful interpretations of VE estimates from inpatient TND studies. VE estimates from inpatient TND studies capture the vaccine-mediated protection of the source population against laboratory-confirmed influenza hospitalizations. If vaccination does not modify disease severity, these estimates are equivalent to VE against influenza virus infection. If chronic cardiopulmonary individuals are enrolled because of non-infectious exacerbation, biased VE estimates (too high) will result. If chronic cardiopulmonary disease status is adjusted for accurately, the VE estimates will be unbiased. If chronic cardiopulmonary illness cannot be adequately be characterized, excluding these individuals may provide unbiased VE estimates. The inpatient TND offers logistic advantages and can provide valid estimates of influenza VE. If highly vaccinated patients with respiratory exacerbation of chronic cardiopulmonary conditions are eligible for study inclusion, biased VE estimates will result unless this group is well characterized and the analysis can adequately adjust for it. Otherwise, such groups of subjects should be excluded from the analysis. © The Author 2016. Published by Oxford University Press on behalf of the International Epidemiological Association.

Molecular Identification of Leishmania spp. in Sand Flies (Diptera: Psychodidae: Phlebotominae) in the Lençóis Maranhenses National Park, Brazil.

PubMed

Pereira-Filho, Adalberto Alves; Fonteles, Raquel Silva; Bandeira, Maria da Conceição Abreu; Moraes, Jorge Luiz Pinto; Rebêlo, José Manuel Macário; Melo, Maria Norma

2018-02-20

Sand flies are very common in the region of Lençóis Maranhenses National Park, an important tourist attraction in Brazil. However, the role of some species and their relative importance locally in Leishmania Ross 1903 transmission is unclear. The objective of this study was to identify Leishmania infection in phlebotomine sand flies collected around the Lençóis Maranhenses National Park, an important conservation area and popular international/national tourist destination with a high incidence of leishmaniasis. Sand flies were collected in peridomiciliary areas on the tourist route from September 2012 to August 2013. The captured females were subjected to molecular analyses for the detection of Leishmania DNA. Sand flies were infected with four Leishmania species: Leishmania (Viannia) braziliensis (Vianna, 1911) was found in Lutzomyia whitmani (Antunes and Coutinho, 1939) (2.1%) and Lutzomyia longipalpis (Lutz and Neiva, 1912) (1.7%); Leishmania (Leishmania) infantum (Nicole, 1908) infected Lutzomyia wellcomei (Fraiha, Shaw, and Lainson, 1971) (20%), Lutzomyia sordellii (Shannon and Del Ponte, 1927) (4.3%), Lu. longipalpis (3.7%), and Lu. whitmani (0.8%); Leishmania (Leishmania) amazonensis (Lainson & Shaw, 1972) was found in Lu. whitmani (0.58%), while Leishmania (Viannia) lainsoni infected Lutzomyia evandroi (Costa Lima and Antunes, 1936) (3.4%), Lu. longipalpis (1.06%), and Lu. whitmani (0.29%). The occurrence of these parasites requires control measures to reduce the incidence of cutaneous leishmaniasis and to contain a possible epidemic of visceral leishmaniasis, the most severe form of the disease.

Detection of Leishmania amazonensis and Leishmania braziliensis in Culicoides (Diptera, Ceratopogonidae) in an endemic area of cutaneous leishmaniasis in the Brazilian Amazonia.

PubMed

Rebêlo, José Manuel Macário; Rodrigues, Bruno Leite; Bandeira, Maria da Conceição Abreu; Moraes, Jorge Luiz Pinto; Fonteles, Raquel Silva; Pereira, Silma Regina Ferreira

2016-12-01

Biting midges in the genus Culicoides act as vectors of arboviruses throughout the world and as vectors of filariasis in Latin America, the Caribbean, and parts of Africa. Although Culicoides spp. are currently not considered to be vectors of Leishmania protozoa, the high abundance of biting midges in areas with active cutaneous leishmaniasis transmission points to the possibility of Culicoides infection by these pathogens. We used PCR to test captured Culicoides species for natural infection with Leishmania spp. We tested 450 Culicoides females, divided into 30 pools of 15 individuals each, as follows: nine pools of C. foxi (135 specimens), seven pools of C. filariferus (105), seven pools of C. insignis (105), five pools of C. ignacioi (75), and two pools of C. flavivenula (30). PCR confirmed the presence of Leishmania braziliensis DNA in C. ignacioi (0.14%), C. insignis (0.14%), and C. foxi (0.11); and Le. amazonensis DNA in C. filariferus (0.14%) and C. flavivenula (0.50%). We conclude that these Culicoides species can be naturally infected, but vector competence and transmission capability must be confirmed in future studies. Our results warrant further investigation into the role of these biting midge species in the leishmaniasis epidemiological cycle. © 2016 The Society for Vector Ecology.

Leishmania (Viannia) braziliensis infection in wild small mammals in ecotourism area of Brazil

PubMed Central

Tonelli, Gabriel Barbosa; Tanure, Aline; Rego, Felipe Dutra; Carvalho, Gustavo Mayr de Lima; Stumpp, Rodolfo; Ássimos, Gabriela Ribeiro; Campos, Aldenise Martins; Lima, Ana Cristina Viana Mariano da Rocha; Gontijo, Célia Maria Ferreira

2017-01-01

Leishmaniases are parasitic diseases transmitted to mammalian hosts by sand fly vectors (Diptera: Psychodidae). Despite the increasing occurrence of visceral and cutaneous leishmaniasis cases in urban centers, their transmission still occur primarily in wild environments and may be associated with professional activities and recreation, such as ecotourism. The Reserva Particular do Patrimônio Natural Santuário do Caraça (RPPNSC) is one of the largest ecotourism attractions in the State of Minas Gerais, Brazil, and comprises an area of environmental preservation with 11,233 hectares presenting a transitional vegetation between Cerrado and Atlantic Forest. The present study describes the abundance of small mammals in RPPNSC, the isolation and identification of Leishmania in five wild animals. Small mammals were bimonthly trapped along 6 trails within the RPPNSC with 10 Tomahawk traps each. Two trails were located in peridomiciliary areas near tourist lodging facilities, and four trails were located at sites visited by tourists in forest areas. The most prevalent species were Akodon cursor, Cerradomys subflavus and Oligoryzomys nigripes. Six isolates of Leishmania were obtained from these animals and identified as Leishmania braziliensis through HSP70-PCR RFLP method. Leishmania spp. DNA was detected by kDNA-PCR method and isolated by biphasic culture. Studies point to some of the captured species as potential wild reservoirs of Leishmania, suggesting they may be involved in the transmission cycle in these wild environments. PMID:29284049

Differential Activation of Human Keratinocytes by Leishmania Species Causing Localized or Disseminated Disease.

PubMed

Scorza, Breanna M; Wacker, Mark A; Messingham, Kelly; Kim, Peter; Klingelhutz, Aloysius; Fairley, Janet; Wilson, Mary E

2017-10-01

All Leishmania species parasites are introduced into mammalian skin through a sand fly bite, but different species cause distinct clinical outcomes. Mouse studies suggest that early responses are critical determinants of subsequent adaptive immunity in leishmaniasis, yet few studies address the role of keratinocytes, the most abundant cell in the epidermis. We hypothesized that Leishmania infection causes keratinocytes to produce immunomodulatory factors that influence the outcome of infection. Incubation of primary or immortalized human keratinocytes with Leishmania infantum or Leishmania major, which cause visceral or cutaneous leishmaniasis, respectively, elicited dramatically different responses. Keratinocytes incubated with L. infantum significantly increased expression of proinflammatory genes for IL-6, IL-8, tumor necrosis factor, and IL-1B, whereas keratinocytes exposed to several L. major isolates did not. Furthermore, keratinocyte-monocyte co-incubation studies across a 4 µM semipermeable membrane suggested that L. infantum-exposed keratinocytes release soluble factors that enhance monocyte control of intracellular L. infantum replication (P < 0.01). L. major-exposed keratinocytes had no comparable effect. These data suggest that L. infantum and L. major differentially activate keratinocytes to release factors that limit infection in monocytes. We propose that keratinocytes initiate or withhold a proinflammatory response at the site of infection, generating a microenvironment uniquely tailored to each Leishmania species that may affect the course of disease. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Leishmania (Viannia) braziliensis infection in wild small mammals in ecotourism area of Brazil.

PubMed

Tonelli, Gabriel Barbosa; Tanure, Aline; Rego, Felipe Dutra; Carvalho, Gustavo Mayr de Lima; Stumpp, Rodolfo; Ássimos, Gabriela Ribeiro; Campos, Aldenise Martins; Lima, Ana Cristina Viana Mariano da Rocha; Gontijo, Célia Maria Ferreira; Paz, Gustavo Fontes; Andrade Filho, José Dilermando

2017-01-01

Leishmaniases are parasitic diseases transmitted to mammalian hosts by sand fly vectors (Diptera: Psychodidae). Despite the increasing occurrence of visceral and cutaneous leishmaniasis cases in urban centers, their transmission still occur primarily in wild environments and may be associated with professional activities and recreation, such as ecotourism. The Reserva Particular do Patrimônio Natural Santuário do Caraça (RPPNSC) is one of the largest ecotourism attractions in the State of Minas Gerais, Brazil, and comprises an area of environmental preservation with 11,233 hectares presenting a transitional vegetation between Cerrado and Atlantic Forest. The present study describes the abundance of small mammals in RPPNSC, the isolation and identification of Leishmania in five wild animals. Small mammals were bimonthly trapped along 6 trails within the RPPNSC with 10 Tomahawk traps each. Two trails were located in peridomiciliary areas near tourist lodging facilities, and four trails were located at sites visited by tourists in forest areas. The most prevalent species were Akodon cursor, Cerradomys subflavus and Oligoryzomys nigripes. Six isolates of Leishmania were obtained from these animals and identified as Leishmania braziliensis through HSP70-PCR RFLP method. Leishmania spp. DNA was detected by kDNA-PCR method and isolated by biphasic culture. Studies point to some of the captured species as potential wild reservoirs of Leishmania, suggesting they may be involved in the transmission cycle in these wild environments.