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Searching for virulence factors in the non-pathogenic parasite to humans Leishmania tarentolae

  • H. AZIZI (a1), K. HASSANI (a1) (a2), Y. TASLIMI (a1), H. SHATERI NAJAFABADI (a3), B. PAPADOPOULOU (a4) and S. RAFATI (a1)...


Leishmania protozoa are obligate intracellular parasites that reside in the phagolysosome of host macrophages and cause a large spectrum of pathologies to humans known as leishmaniases. The outcome of the disease is highly dependent on the parasite species and on its ascribed virulence factors and the immune status of the host. Characterization of the genome composition of non-pathogenic species could ultimately open new horizons in Leishmania developmental biology and also the disease monitoring. Here, we provide evidence that the lizard non-pathogenic to humans Leishmania tarentolae species expresses an Amastin-like gene, cysteine protease B (CPB), lipophosphoglycan LPG3 and the leishmanolysin GP63, genes well-known for their potential role in the parasite virulence. These genes were expressed at levels comparable to those in L. major and L. infantum both at the level of mRNA and protein. Alignment of the L. tarentolae proteins with their counterparts in the pathogenic species demonstrated that the degree of similarity varied from 59% and 60% for Amastin, 89% for LPG3 and 71% and 68% for CPB, in L. major and L. infantum, respectively. Interestingly, the A2 gene, expressed specifically by the L. donovani complex which promotes visceralization, was absent in L. tarentolae. These findings suggest that the lack of pathogenicity in L. tarentolae is not associated with known virulence genes such as LPG3, CPB, GP63 and Amastin, and that other factors either unique to L. tarentolae or missing from this species may be responsible for the non-pathogenic potential of this lizard parasite.


Corresponding author

*Corresponding author: Molecular Immunology and Vaccine Research Laboratory, Pasteur Institute of Iran, Tehran, Iran. Tel: +98 21 66953311. Fax: +98 21 66465132. E-mail: or


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Akopyants, N. S., Matlib, R. S., Bukanova, E. N., Smeds, M. R., Brownstein, B. H., Stormo, G. D. and Beverley, S. M. (2004). Expression profiling using random genomic DNA microarrays identifies differentially expressed genes associated with three major developmental stages of the protozoan parasite Leishmania major. Molecular and Biochemical Parasitology 136, 7186.
Alexander, J., Coombs, G. H. and Mottram, J. C. (1998). Leishmania mexicana cysteine proteinase-deficient mutants have attenuated virulence for mice and potentiate a Th1 response. Journal of Immunology 161, 67946801.
Bordier, C. (1987). The promastigote surface protease of Leishmania. Parsitology Today 3, 151153.
Breton, M., Tremblay, M. J., Ouellette, M. and Papadopoulou, B. (2005). Live nonpathogenic parasitic vector as a candidate vaccine against visceral leishmaniasis. Infection and Immunity 73, 63726382.
Buxbaum, L. U., Denise, H., Coombs, G. H., Alexander, J., Mottram, J. C. and Scott, P. (2003). Cysteine protease B of Leishmania mexicana inhibits host Th1 responses and protective immunity. Journal of Immunology 171, 37113717.
Cameron, P., McGachy, A., Anderson, M., Paul, A., Coombs, G. H., Mottram, J. C., Alexander, J. and Plevin, R. (2004). Inhibition of lipopolysaccharide-induced macrophage IL-12 production by Leishmania mexicana amastigotes: the role of cysteine peptidases and the NF-kappaB signaling pathway. Journal of Immunology 173, 32973304.
Campbell, D., Kurath, U. and Fleischmann, J. (1992). Identification of a gp63 surface glycoprotein in Leishmania tarentolae. FEMS Microbiology Letters 75, 8992.
Chen, D. Q., Kolli, B. K., Yadava, N., Lu, H. G., Gilman-Sachs, A., Peterson, D. A. and Chang, K-P. (2000). Episomal expression of specific sense and antisense mRNAs in Leishmania amazonensis: modulation of gp63 level in promastigotes and their infection of macrophages in vitro. Infection and Immunity 68, 8086.
Coelho, E. A., Tavares, C. A., Carvalho, F. A., Chaves, K. F., Teixeira, K. N., Rodrigues, R. C., Charest, H., Matlashewski, G., Gazzinelli, R. T. and Fernandes, A. P. (2003). Immune responses induced by the Leishmania (Leishmania) donovani A2 antigen, but not by the LACK antigen, are protective against experimental Leishmania (Leishmania) amazonensis infection. Infection and Immunity 71, 39883994.
Croan, D. G., Morrison, D. A. and Ellis, J. T. (1997). Evolution of the genus Leishmania revealed by comparison of DNA and RNA polymerase gene sequences. Molecular and Biochemical Parasitology 89, 149159.
Descoteaux, A., Avila, H. A., Zhang, K., Turco, S. J. and Beverley, S. M. (2002). Leishmania LPG3 encodes a GRP94 homolog required for phosphoglycan synthesis implicated in parasite virulence but not viability. The EMBO Journal 21, 44584469.
Desjeux, P. (2004). Leishmaniasis: current situation and new perspectives. Comparative Immunology, Microbiology and Infectious Diseases 27, 305318.
Dietzsch, J., Gehlenborg, N. and Nieselt, K. (2006). Mayday – a Microarray Data Analysis Workbench. Bioinformatics 22, 10101012.
Dollahon, N. R. and Janovy, J. (1973). Leismania adleri: in vitro phagocytosis by lizard leukocytes and peritoneal cells. Experimental Parasitology 34, 5661.
Frame, M. J., Mottram, J. C. and Coombs, G. H. (2000). Analysis of the roles of cysteine proteinases of Leishmania mexicana in the host-parasite interaction. Parasitology 121, 367377.
Fukao, T., Matsuda, S. and Koyasu, S. (2000). Synergistic effects of IL-4 and IL-18 on IL-12-dependent IFN-gamma production by dendritic cells. Journal of Immunology 164, 6471.
Garin, Y. J., Meneceur, P., Pratlong, F., Dedet, J. P., Derouin, F. and Lorenzo, F. (2005). A2 gene of Old World cutaneous Leishmania is a single highly conserved functional gene. BMC Infectious Disease 28, 18.
Genest, P. A., Haimeur, A., Légaré, D., Sereno, D., Roy, G., Messier, N., Papadopoulou, B. and Ouellette, M. (2008). A protein of the leucine-rich repeats (LRRs) superfamily is implicated in antimony resistance in Leishmania infantum amastigotes. Molecular and Biochemical Parasitology 158, 9599.
Ghedin, E., Charest, H. and Matlashewski, G. (1998). A2rel: a constitutively expressed Leishmania gene linked to an amastigote-stage-specific gene. Molecular and Biochemical Parasitology 93, 2329.
Ghosh, D. K. and Honigberg, B. M. (1976). Activities of glucose-6-phosphate, 6-phosphogluconate, and isocitrate dehydrogenases from Leishmania donovani cultivated at 25 and 37 C. Journal of Parasitology 23, 450455.
Gregory, D. J., Godbout, M., Contreras, I., Forget, G. and Olivier, M. (2008). A novel form of NF-κB is induced by Leishmania infection: involvement in macrophage gene expression. European Journal of Immunology 38, 10711081.
Hide, M., Bras-Goncalves, R. and Bañuls, A.-L. (2007). Specific cpb copies within the Leishmania donovani complex: evolutionary interpretations and potential clinical implications in humans. Parasitology 134, 379389.
Ilg, T. (2000). Lipophosphoglycan is not required for infection of macrophages or mice by Leishmania mexicana. The EMBO Journal 19, 19531962.
Ivens, A. C., Peacock, C. S., Worthey, E. A., Murphy, L., Aggarwal, G., Berriman, M., Sisk, E., Rajandream, M. A., Adlem, E., Aert, R., Anupama, A., Apostolou, Z., Attipoe, P., Bason, N., Bauser, C., Beck, A., Beverley, S. M., Bianchettin, G., Borzym, K., Bothe, G., Bruschi, C. V., Collins, M., Cadag, E., Ciarloni, L., Clayton, C., Coulson, R. M., Cronin, A., Cruz, A. K., Davies, R. M., De Gaudenzi, J., Dobson, D. E., Duesterhoeft, A., Fazelina, G., Fosker, N., Frasch, A. C., Fraser, A., Fuchs, M., Gabel, C., Goble, A., Goffeau, A., Harris, D., Hertz-Fowler, C., Hilbert, H., Horn, D., Huang, Y., Klages, S., Knights, A., Kube, M., Larke, N., Litvin, L., Lord, A., Louie, T., Marra, M., Masuy, D., Matthews, K., Michaeli, S., Mottram, J. C., Muller-Auer, S., Munden, H., Nelson, S., Norbertczak, H., Oliver, K., O'Neil, S., Pentony, M., Pohl, T. M., Price, C., Purnelle, B., Quail, M. A., Rabbinowitsch, E., Reinhardt, R., Rieger, M., Rinta, J., Robben, J., Robertson, L., Ruiz, J. C., Rutter, S., Saunders, D., Schafer, M., Schein, J., Schwartz, D. C., Seeger, K., Seyler, A., Sharp, S., Shin, H., Sivam, D., Squares, R., Squares, S., Tosato, V., Vogt, C., Volckaert, G., Wambutt, R., Warren, T., Wedler, H., Woodward, J., Zhou, S., Zimmermann, W., Smith, D. F., Blackwell, J. M., Stuart, K. D., Barrell, B. and Myler, P. J. (2005). The genome of the kinetoplastid parasite, Leishmania major. Science 309, 436442.
Janovy, J. (1972). Temperature and metabolism in Leishmania. 3. Some dehydrogenases of L. donovani, L. mexicana, and L. tarentolae. Experimental Parasitology 32, 196205.
Joshi, B. P., Kelly, B. L., Kamhawi, H., Sacks, D. L. and McMaster, W. R. (2002). Targeted gene deletion in Leishmania major identifies leishmanolysin (GP63) as a virulence factor. Molecular and Biochemical Parasitology 120, 3340.
Krogh, A., Larsson, B., von Heijne, G. and Sonnhammer, E. L. (2001). Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. Journal of Molecular Biology 19, 567580.
Landweber, L. F. and Gilbert, W. (1994). Phylogenetic analysis of RNA editing: a primitive genetic phenomenon. Proceedings of the National Academy of Sciences, USA 91, 918921.
Larreta, R., Soto, M., Alonso, C. and Requena, J. M. (2000). Leishmania infantum: gene cloning of the GRP94 homologue, its expression as recombinant protein and analysis of antigenicity. Experimental Parasitology 96, 108115.
Leifso, K., Cohen-Freue, G., Dogra, N., Murray, A. and McMaster, W. R. (2007). Genomic and proteomic expression analysis of Leishmania promastigote and amastigote life stages. The Leishmania genome is constitutively expressed. Molecular and Biochemical Parasitology 152, 3546.
Maslov, D. A., Avila, H. A., Lake, J. A. and Simpson, L. (1994). Evolution of RNA editing in kinetoplastid protozoa. Nature, London 368, 345348.
McGwire, B. S., O'Connell, W. A., Chang, K. P. and Engman, D. M. (2002). Extracellular release of the glycosylphosphatidylinositol (GPI)-linked Leishmania surface metalloprotease, gp63, is independent of GPI phospholipolysis: implications for parasite virulence. Journal of Biological Chemistry 277, 88028809.
McMaster, W. R., Morrison, C. J., MacDonald, M. H. and Joshi, P. B. (1994). Mutational and functional analysis of the Leishmania surface metalloproteinase GP63: similarities to matrix metalloproteinases. Parasitology 108, S29S36.
McNicoll, F., Drummelsmith, J., Muller, M., Madore, E., Boilard, N., Ouellette, M. and Papadopoulou, B. (2006). A combined proteomic and transcriptomic approach to the study of stage differentiation in Leishmania infantum. Proteomics 6, 35673581.
McNicoll, F., Muller, M., Cloutier, S., Boilard, N., Rochette, A., Dube, M. and Papadopoulou, B. (2005). Distinct 3′-untranslated region elements regulate stage-specific mRNA accumulation and translation in Leishmania. Journal of Biological Chemistry 280, 3523835246.
Medina-Acosta, E., Beverley, S. M. and Russell, D. G. (1993). Evolution and expression of the Leishmania surface proteinase (gp63) gene locus. Infectious Agents and Diseases 2, 2534
Mottram, J. C., Brooks, D. R. and Coombs, G. H. (1998). Roles of cysteine proteinases of trypanosomes and Leishmania in host–parasite interactions. Current Opinion in Microbiology 1, 455460.
Mottram, J. C., Coombs, G. H. and Alexander, J. (2004). Cysteine peptidases as virulence factors of Leishmania. Current Opinion in Microbiology 7, 375381.
Mundodi, V., Somanna, A., Farrell, P. J. and Gedamu, L. (2002). Genomic organization and functional expression of differentially regulated cysteine protease genes of Leishmania donovani complex. Gene 282, 257265.
Murray, H. W., Berman, J. D., Davies, C. R. and Saravia, N. G. (2005). Advances in leishmaniasis. Lancet 366, 15611577.
Nakhaee, A., Taheri, T., Taghikhani, M., Mohebali, M., Salmanian, A. H., Fasel, N. and Rafati, S. (2004). Humoral and cellular immune responses against type I cysteine proteinase of Leishmania infantum are higher in asymptomatic than symptomatic dogs selected from a naturally infected population. Veterinary Parasitology 119, 107123.
Noyes, H. A., Chance, M. L., Croan, D. G. and Ellis, J. T. (1998). Leishmania (Sauroleishmania): a comment on classification. Parasitology Today 14, 167.
Orlando, T. C., Rubio, M. A., Sturm, N. R., Campbell, D. A. and Floeter-Winter, L. M. (2002). Intergenic and external transcribed spacers of ribosomal RNA genes in lizard-infecting Leishmania: molecular structure and phylogenetic relationship to mammal-infecting Leishmania in the subgenus Leishmania (Leishmania). Memórias do Instituto Oswaldo Cruz 97, 695701.
Ouakad, M., Bahi-Jaber, N., Chenik, M., Dellagi, K. and Louzir, H. (2007 a). Selection of endogenous reference genes for gene expression analysis in Leishmania major developmental stages. Parasitology Research 101, 473477.
Ouakad, M., Chenik, M., Achour-Chenik, Y. B., Louzir, H. and Dellagi, K. (2007 b). Gene expression analysis of wild Leishmania major isolates: identification of genes preferentially expressed in amastigotes. Parasitology Research 100, 255264.
Paape, D., Lippuner, C., Schmid, M., Ackermann, R., Barrios-Llerena, M. E., Zimny-Arndt, U., Brinkmann, V., Arndt, B., Pleissner, K. P., Jungblut, P. R. and Aebischer, T. (2008). Transgenic, fluorescent Leishmania mexicana allow direct analysis of the proteome of intracellular amastigotes. Molecular and Cellular Proteomics 7, 16881701.
Peacock, C. S., Seeger, K., Harris, D., Murphy, L., Ruiz, J. C., Quail, M. A., Peters, N., Adlem, E., Tivey, A., Aslett, M., Kerhornou, A., Ivens, A., Fraser, A., Rajandream, M. A., Carver, T., Norbertczak, H., Chillingworth, T., Hance, Z., Jagels, K., Moule, S., Ormond, D., Rutter, S., Squares, R., Whitehead, S., Rabbinowitsch, E., Arrowsmith, C., White, B., Thurston, S., Bringaud, F., Baldauf, S. L., Faulconbridge, A., Jeffares, D., Depledge, D. P., Oyola, S. O., Hilley, J. D., Brito, L. O., Tosi, L. R., Barrell, B., Cruz, A. K., Mottram, J. C., Smith, D. F. and Berriman, M. (2007). Comparative genomic analysis of three Leishmania species that cause diverse human disease. Nature Genetics 39, 839847.
Previato, J. O., Jones, C., Wait, R., Routier, F., Saraiva, E. and Mendonça-Previato, L. (1997). Leishmania adleri, a lizard parasite, expresses structurally similar glycoinositolphospholipids to mammalian Leishmania. Glycobiology 7, 687695.
Rafati, S., Hassani, N., Taslimi, Y., Movassagh, H., Rochette, A. and Papadopoulou, B. (2006). Amastin peptide-binding antibodies as biomarkers of active human visceral leishmaniasis. Clinical Vaccine Immunology 13, 11041110.
Rafati, S., Nakhaee, A., Taheri, T., Ghashghaii, A., Salmanian, A. H., Jimenez, M., Mohebali, M., Masina, S. and Fasel, N. (2003). Expression of cysteine proteinase type I and II of Leishmania infantum and their recognition by sera during canine and human visceral leishmaniasis. Experimental Parasitology 103, 143151.
Rafati, S., Nakhaee, A., Taheri, T., Taslimi, Y., Darabi, H., Eravani, D., Sanos, S., Kaye, P., Taghikhani, M., Jamshidi, S. and Rad, M. A. (2005). Protective vaccination against experimental canine visceral leishmaniasis using a combination of DNA and protein immunization with cysteine proteinases type I and II of L. infantum. Vaccine 23, 37163725.
Rafati, S., Salmanian, A. H., Hashemi, K., Schaff, C., Belli, S. and Fasel, N. (2001). Identification of Leishmania major cysteine proteinases as targets of the immune response in humans. Molecular and Biochemical Parasitology 113, 3543.
Requena, J. M., Alonso, C. and Soto, M. (2000). Evolutionarily conserved proteins as prominent immunogens during Leishmania infections. Parasitology Today 16, 246250.
Rochette, A., McNicoll, F., Girard, J., Breton, M., Leblanc, E., Bergeron, M. G. and Papadopoulou, B. (2005). Characterization and developmental gene regulation of a large gene family encoding amastin surface proteins in Leishmania spp. Molecular and Biochemical Parasitology 140, 205220.
Rochette, A., Raymond, F., Ubeda, J. M., Smith, M., Messier, N., Boisvert, S., Rigault, P., Corbeil, J., Ouellette, M. and Papadopoulou, B. (2008). Genome-wide gene expression profiling analysis of Leishmania major and Leishmania infantum developmental stages reveals substantial differences between the two species. BMC Genomics 9, 255281.
Rosenthal, P. J. (1999). Proteases of protozoan parasites. Advances in Parasitology 43, 105159.
Rosenzweig, D., Smith, D., Opperdoes, F., Stern, S., Olafson, R. W. and Zilberstein, D. (2008). Retooling Leishmania metabolism: from sand fly gut to human macrophage. The FASEB Journal 22, 590602.
Salotra, P., Duncan, R. C., Singh, R., Subba Raju, B. V., Sreenivas, G. and Nakhasi, H. L. (2006). Upregulation of surface proteins in Leishmania donovani isolated from patients of post kala-azar dermal leishmaniasis. Microbes and Infection 8, 637644.
Sambrook, J. and Russell, D. (2001). Molecular Cloning: a Laboratory Manual, 3rd Edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.
Spath, G. F., Epstein, L., Leader, B., Singer, S. M., Avila, H. A., Turco, S. J. and Beverley, S. M. (2000). Lipophosphoglycan is a virulence factor distinct from related glycoconjugates in the protozoan parasite Leishmania major. Proceedings of the National Academy of Sciences, USA 97, 92589263.
Spath, G. F., Garraway, L. A., Turco, S. J. and Beverley, S. M. (2003). The role(s) of lipophosphoglycan (LPG) in the establishment of Leishmania major infections in mammalian hosts. Proceedings of the National Academy of Sciences, USA 100, 95369541.
Stiles, J. K., Hicock, P. I., Shah, P. H. and Mead, J. C. (1999). Genomic organization, transcription, splicing and gene regulation in Leishmania. Annals of Tropical Medicine and Parasitology 93, 781807.
Williams, R. A., Tetley, L., Mottram, J. C. and Coombs, G. H. (2006).Cysteine peptidases CPA and CPB are vital for autophagy and differentiation in Leishmania mexicana. Molecular Microbiology 61, 655674.
Wu, Y., El-Fakhry, Y., Sereno, D., Tamar, S. and Papadopoulou, B. (2000). A new developmentally regulated gene family in Leishmania amastigotes encoding a homolog of amastin surface proteins. Molecular and Biochemical Parasitology 110, 345357.
Yao, C., Donelson, J. E. and Wilson, M. E. (2003). The major surface protease (MSP or GP63) of Leishmania sp. Biosynthesis, regulation of expression, and function. Molecular and Biochemical Parasitology 132, 116.
Zadeh-Vakili, A., Taheri, T., Taslimi, Y., Doustdari, F., Salmanian, A. H. and Rafati, S. (2004). Immunization with the hybrid protein vaccine, consisting of Leishmania major cysteine proteinases Type I (CPB) and Type II (CPA), partially protects against leishmaniasis. Vaccine 22, 19301940.
Zhang, W. W. and Matlashewski, G. (2001). Characterization of the A2–A2rel gene cluster in Leishmania donovani: involvement of A2 in visceralization during infection. Molecular Microbiology 39, 935948.
Zhang, W. W. and Matlashewski, G. (2004). In vivo selection for Leishmania donovani miniexon genes that increase virulence in Leishmania major. Molecular Microbiology 54, 10511062.
Zhang, W. W. and Matlashewski, G. (1997). Loss of virulence in Leishmania donovani deficient in an amastigote-specific protein, A2. Proceedings of the National Academy of Sciences, USA 94, 88078811.
Zhang, W. W., Mendez, S., Ghosh, A., Myler, P., Ivens, A., Clos, J., Sacks, D. L. and Matlashewski, G. (2003). Comparison of the A2 gene locus in Leishmania donovani and Leishmania major and its control over cutaneous infection. Journal of Biological Chemistry 278, 3550835515.


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