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DNA-Protein Immunization Using Leishmania Peroxidoxin-1 Induces a Strong CD4+ T Cell Response and Partially Protects Mice from Cutaneous Leishmaniasis: Role of Fusion Murine Granulocyte-Macrophage Colony-Stimulating Factor DNA Adjuvant  [PDF]
Abebe Genetu Bayih ,Nada S. Daifalla,Lashitew Gedamu
PLOS Neglected Tropical Diseases , 2014, DOI: 10.1371/journal.pntd.0003391
Abstract: Background To date, no universally effective and safe vaccine has been developed for general human use. Leishmania donovani Peroxidoxin-1 (LdPxn-1) is a member of the antioxidant family of proteins and is predominantly expressed in the amastigote stage of the parasite. The aim of this study was to evaluate the immunogenicity and protective efficacy of LdPxn-1 in BALB/c mice in heterologous DNA-Protein immunization regimen in the presence of fusion murine granulocyte-macrophage colony-stimulating factor (mGMCSF) DNA adjuvant. Methodology and Principal Findings A fusion DNA of LdPxn1 and mGMCSF was cloned into a modified pcDNA vector. To confirm the expression in mammalian system, Chinese hamster ovary cells were transfected with the plasmid vector containing LdPxn1 gene. BALB/c mice were immunized twice with pcDNA-mGMCSF-LdPxn-1 or pcDNA-LdPxn1 DNA and boosted once with recombinant LdPxn-1 protein. Three weeks after the last immunization, mice were infected with Leishmania major promastigotes. The result showed that immunization with pcDNA-mGMCSF-LdPxn1 elicited a mixed Th-1/Th-2 immune response with significantly higher production of IFN-γ than controls. Intracellular cytokine staining of antigen-stimulated spleen cells showed that immunization with this antigen elicited significantly higher proportion of CD4+ T cells that express IFN-γ, TNF-α, or IL-2. The antigen also induced significantly higher proportion of multipotent CD4+ cells that simultaneously express the three Th-1 cytokines. Moreover, a significant reduction in the footpad swelling was seen in mice immunized with pcDNA-mGMCSF-LdPxn1 antigen. Expression study in CHO cells demonstrated that pcDNA-mGMCSF-LdPxn-1 was expressed in mammalian system. Conclusion The result demonstrates that immunization of BALB/c mice with a plasmid expressing LdPxn1 in the presence of mGMCSF adjuvant elicits a strong specific immune response with high level induction of multipotent CD4+ cells that mediate protection of the mice from Leishmania major infection. To our knowledge, this is the first study showing the vaccine potential of Leishmania peroxidoxin -1.
Development of Cutaneous Leishmaniasis after Leishmania Skin Test
Paulo R. Machado,Augusto M. Carvalho,Gustavo U. Machado,Marina L. Dantas,Sérgio Arruda
Case Reports in Medicine , 2011, DOI: 10.1155/2011/631079
Abstract: Thirty-year-old female with a previous history of a cutaneous ulcer suspicious of leishmaniasis 20 years ago presented with a new complaint of a depressed papular lesion 8×7 mm in the right lower extremity. The lesion was of 10-day duration. Because early cutaneous leishmaniasis (CL) lesions may have a non-ulcerated appearance, a Leishmania skin test (LST) was performed on the forearm with a strong positive result (38×32 mm). After 8 days, the lesion in the leg, which was diagnosed as folliculitis, completely healed. However, a typical CL ulcer (26×24 mm) developed at the LST site. Histopathology of the new lesion did not identifiy parasites, but the findings were consistent with a diagnosis of CL. Further analysis identified amastigotes by immunohistochemical stain. Mononuclear cells harvested from the patient were stimulated with Leishmania antigen and showed high levels of production of both tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ): 2,943 pg/mL and 2,313 pg/mL, respectively. After 40 days of treatment with antimony and pentoxifylline, the ulcer resolved. The development of CL at the LST site suggests a strong Th1 immune response, and it is an in vivo documentation of the role of the host immune response in the pathology of CL. It teaches us that LST should be cautiously, if at all, used in patients with self-healing CL ulcers.
Active cutaneous leishmaniasis in Brazil, induced by Leishmania donovani chagasi
Oliveira Neto, M. P.;Grimaldi Junior, Gabriel;Momen, Hooman;Pacheco, R. S.;Marzochi, Mauro Celio de A.;McMahon Pratt, D.;
Memórias do Instituto Oswaldo Cruz , 1986, DOI: 10.1590/S0074-02761986000300006
Abstract: l.d. chagasi was isolated from active cutaneous leishmaniasis in both human and canine infections in an endemic area in rio de janeiro, brazil. both isolates were identified by molecular and immunological characterization of the parasite using three different methods: electrophoretic mobility of isoenzymes; restriction endonuclease fragment analysis of kdna and serodeme analysis using monoclonal antibodies. this seems to be the first well documented case in the new world of a "viscerotropic" leishmania inducing a case of cutaneous leishmaniasis. this observation emphasizes that the diagnosis of the etiologic agent of human or canine visceral leishmaniasis based solely upon clinical and epidemiological critwria may lead to erroneous conclusions.
The histopathology of cutaneous leishmaniasis due to Leishmania (Leishmania) mexicana in the Yucatan peninsula, Mexico
Andrade-Narvaez, Fernando J.;Medina-Peralta, Salvador;Vargas-Gonzalez, Alberto;Canto-Lara, Silvia B.;Estrada-Parra, Sergio;
Revista do Instituto de Medicina Tropical de S?o Paulo , 2005, DOI: 10.1590/S0036-46652005000400003
Abstract: localized cutaneous leishmaniasis (lcl) known as "chiclero's ulcer" in southeast mexico, was described by seidelin in 1912. since then the sylvatic region of the yucatan peninsula has been documented as an endemic focus of lcl. this study of 73 biopsies from parasitological confirmed lesions of lcl cases of leishmania (leishmania) mexicana infection was undertaken: 1) to examine host response at tissue level; and 2) to relate manifestations of this response to some characteristics of clinical presentation. based on magalh?es' classification we found that the most common pattern in our lcl cases caused by l. (l.) mexicana was predominantly characterized by the presence of unorganized granuloma without necrosis, (43.8%). another important finding to be highlighted is the fact that in 50/73 (68.5%) parasite identification was positive. there was direct relation between the size of the lesion and time of evolution (rs = 0.3079, p = 0.03), and inverse correlation between size of the lesion and abundance of amastigotes (rs = -0.2467, p = 0.03). in view of the complexity of clinical and histopathological findings, cell-mediated immune response of the disease related to clinical and histopathological features, as so genetic background should be studied.
Molecular identification of Leishmania species causing cutaneous leishmaniasis in Mashhad, Iran
Mohammad Reza Mahmoodi,Masoud Mohajery,Jalil Tavakkol Afshari,Mohhamad Taghae Shakeri
Jundishapur Journal of Microbiology , 2010,
Abstract: Introduction and objective: Cutaneous leishmaniasis (CL) is considered as an important health problem in many parts of Iran especially in Mashhad, north-eastern part of Iran. Various species of Leishmania cause the disease. Identification of Leishmania parasites is useful for control and preventive plans. Although epidemiological and clinical findings are necessary but they are not sufficient for identification of causative agents of CL. In order to identify Leishmania spp. a definite molecular technique, Polymerase Chain Reaction (PCR) method was used over a 12 months period.Materials and methods: A total of twenty-one patients participated. Direct smear and culture in modified NNN medium followed by sub-culture in RPMI-1640 were performed for each case. Genomic DNA was extracted by using proteinase k and amplified by specific primers of kDNA. The PCR product was analysed by gel electrophoresis using 2% agarose. Gel staining was performed by ethidium bromide. The presence of 620bp fragment indicated Leishmania major and 800bp indicated L. tropica.Results: Of 21 positive cultures out of 53 positive samples, nineteen isolates were identified as L. tropica and two others were identified as L. major. However, by previous investigations, Mashhad was known, as an endemic focus for Anthroponotic Cutaneous Leishmaniasis (ACL), but it is now concluded that both ACL and Zoonotic Cutaneous Leishmaniasis (ZCL) are present in Mashhad and L. tropica is the dominant species.Conclusion: Both L. tropica and L. major are the causative agents of cutaneous leishmaniasis in Mashhad. L. tropica is the dominant Leishmania species in Mashhad. However PCR technique is a very reliable method to detect Leishmania DNA, but it is not easy to obtain Leishmania culture samples.
Experimental infection of Lutzomyia longipalpis fed on a patient with cutaneous leishmaniasis due to Leishmania mexicana amazonensis
Deane, Leonidas M.;Rangel, Elizabeth Ferreira;Paes-Oliveira, Manoel;Grimaldi Junior, Gabriel;Momen, Hooman;Souza, Nataly de;Wermelinger, Eduardo D.;Barbosa, André F.;
Memórias do Instituto Oswaldo Cruz , 1986, DOI: 10.1590/S0074-02761986000100020
Abstract: the authors were able to infect phlebotomine sandflies on a human case of american cutaneous leishmaniasis by feeding females of lutzomyia longipalpis on a patient with a lesion due to leishmania mexicana amazonensis.
Comparison of the specificity of PCR and the histopathological detection of leishmania for the diagnosis of American cutaneous leishmaniasis
Medeiros, A.C.R.;Rodrigues, S.S.;Roselino, A.M.F.;
Brazilian Journal of Medical and Biological Research , 2002, DOI: 10.1590/S0100-879X2002000400002
Abstract: more precise and rapid diagnostic methods for american cutaneous leishmaniasis (acl) are necessary because of the growing number of cases observed in brazil, including the northeastern region of the state of s?o paulo. we applied pcr to 54 skin or mucosal biopsies from patients with a clinical and/or laboratory diagnosis of acl using primers 13a and 13b, with positive results being obtained for 82% of the samples. when the pcr results were compared to those of histopathological leishmania detection, pcr showed superior results with 81.5% sensitivity and 95% ci of 68.0-95.1%. the montenegro skin test (mst) was positive in 88.7% of patients. since mst cannot be used as a diagnostic tool in endemic areas, the present results strongly suggest the use of pcr for the etiological confirmation of acl, with emphasis on the mucosal form.
Comparison of the specificity of PCR and the histopathological detection of leishmania for the diagnosis of American cutaneous leishmaniasis  [cached]
Medeiros A.C.R.,Rodrigues S.S.,Roselino A.M.F.
Brazilian Journal of Medical and Biological Research , 2002,
Abstract: More precise and rapid diagnostic methods for American cutaneous leishmaniasis (ACL) are necessary because of the growing number of cases observed in Brazil, including the northeastern region of the State of S o Paulo. We applied PCR to 54 skin or mucosal biopsies from patients with a clinical and/or laboratory diagnosis of ACL using primers 13A and 13B, with positive results being obtained for 82% of the samples. When the PCR results were compared to those of histopathological leishmania detection, PCR showed superior results with 81.5% sensitivity and 95% CI of 68.0-95.1%. The Montenegro skin test (MST) was positive in 88.7% of patients. Since MST cannot be used as a diagnostic tool in endemic areas, the present results strongly suggest the use of PCR for the etiological confirmation of ACL, with emphasis on the mucosal form.
Cutaneous leishmaniasis caused by members of Leishmania braziliensis complex in Nayarit, State of Mexico
Sanchez-Tejeda, Gustavo;Rodríguez, Noris;Parra, Carlos I;Hernandez-Montes, Omar;C Barker, Douglas;Monroy-Ostria, Amalia;
Memórias do Instituto Oswaldo Cruz , 2001, DOI: 10.1590/S0074-02762001000100002
Abstract: an epidemiological study was carried out in the northern mexican state, nayarit. fourteen patients with possible cutaneous leishmaniasis skin lesions gave positive montenegro skin tests. biopsies were taken from the skin ulcer and analyzed by polymerase chain reaction (pcr) with specific primers for the leishmania mexicana complex; however all biopsies were not amplified. pcr carried out with specific primers for the l. braziliensis complex resulted in the amplification of all patient dna. dna from 12 out of 14 biopsies gave positive amplification with primers species specific for l. (viannia) braziliensis and hybridized with a species specific l. (v.) braziliensis probe. these results demonstrate the presence in nayarit of at least two members of the l. braziliensis complex. most of the cutaneous lesions were caused by l. (v.) braziliensis and two by another species belonging to the l. braziliensis complex. as far as we are aware, this is the first report of l. (v.) braziliensis in nayarit. the main risk factor associated with the contraction of this disease in nayarit is attributed to working on coffee plantations.
Cutaneous leishmaniasis caused by members of Leishmania braziliensis complex in Nayarit, State of Mexico  [cached]
Sanchez-Tejeda Gustavo,Rodríguez Noris,Parra Carlos I,Hernandez-Montes Omar
Memórias do Instituto Oswaldo Cruz , 2001,
Abstract: An epidemiological study was carried out in the northern Mexican state, Nayarit. Fourteen patients with possible cutaneous leishmaniasis skin lesions gave positive Montenegro skin tests. Biopsies were taken from the skin ulcer and analyzed by polymerase chain reaction (PCR) with specific primers for the Leishmania mexicana complex; however all biopsies were not amplified. PCR carried out with specific primers for the L. braziliensis complex resulted in the amplification of all patient DNA. DNA from 12 out of 14 biopsies gave positive amplification with primers species specific for L. (Viannia) braziliensis and hybridized with a species specific L. (V.) braziliensis probe. These results demonstrate the presence in Nayarit of at least two members of the L. braziliensis complex. Most of the cutaneous lesions were caused by L. (V.) braziliensis and two by another species belonging to the L. braziliensis complex. As far as we are aware, this is the first report of L. (V.) braziliensis in Nayarit. The main risk factor associated with the contraction of this disease in Nayarit is attributed to working on coffee plantations.
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