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A non-commercial approach for the generation of transgenic Leishmania tarentolae and its application in antileishmanial drug discovery

Published online by Cambridge University Press:  13 May 2016

TATIANA PINEDA ,
YESENIA VALENCIA ,
MARÍA F. FLÓREZ ,
SERGIO A. PULIDO ,
IVÁN D. VÉLEZ  and
SARA M. ROBLEDO
TATIANA PINEDA
Affiliation:
PECET, Medical Research Institute, School of Medicine, University of Antioquia, Medellín, Colombia
YESENIA VALENCIA
Affiliation:
PECET, Medical Research Institute, School of Medicine, University of Antioquia, Medellín, Colombia
MARÍA F. FLÓREZ
Affiliation:
PECET, Medical Research Institute, School of Medicine, University of Antioquia, Medellín, Colombia
SERGIO A. PULIDO
Affiliation:
PECET, Medical Research Institute, School of Medicine, University of Antioquia, Medellín, Colombia
IVÁN D. VÉLEZ
Affiliation:
PECET, Medical Research Institute, School of Medicine, University of Antioquia, Medellín, Colombia
SARA M. ROBLEDO*
Affiliation:
PECET, Medical Research Institute, School of Medicine, University of Antioquia, Medellín, Colombia
*
*Corresponding author: PECET, Medical Research Institute, School of Medicine, University of Antioquia, Medellín, Calle 70 # 52-21, Colombia. Phone: +574 2196503. Fax: +574 2196511. E-mail: [email protected]
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Summary

Leishmaniasis is a parasitic infection caused by several species of the genus Leishmania that is considered as a neglected disease. Drug development process requires a robust and updated high-throughput technology to the evaluation of candidate compounds that imply the manipulation of the pathogenic species of the parasite in the laboratory. Therefore, it is restricted to trained personal and level II biosafety environments. However, it has been established the utility of Leishmania tarentolae as a model for in vitro screening of antileishmanial agents without the necessity of level II biosafety setups. In parallel the transfection of Leishmania parasites with reporter genes as the eGFP using non-commercial integration vectors like the pIRmcs3(−) has proved to be a powerful tool for the implementation of semi automatized high-throughput platforms for the evaluation of antileishmanial compounds. Here we report the generation of a new L. tarentolae strain overexpressing the eGFP gene harboured by the non-commercial vector pIR3(−). We also demonstrate its utility for the semi-automatized screening of antileshmanial compounds in intracellular forms of the L. tarentolae parasite.

Keywords

Leishmania tarentolaegreen fluorescent proteintransgenic parasitespIRmsc3(−)
Type
Research Article
Information
Parasitology , Volume 143 , Issue 9 , August 2016 , pp. 1133 - 1142
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Copyright
Copyright © Cambridge University Press 2016 

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Footnotes

†

These authors contributed equally to this work.

References

REFERENCES

Alvar, J., Vélez, I. D., Bern, C., Herrero, M., Desjeux, P., Cano, J., Jannin, J., den Boer, M. and WHO Leishmaniasis Control Team (2012). Leishmaniasis worldwide and global estimates of its incidence. PLoS ONE 7, e35671.CrossRefGoogle ScholarPubMed
Ashutosh, S., Ramesh, S. and Goyal, N. (2005). Use of Leishmania donovani field isolates expressing the luciferase reporter gene in in vitro drug screening. Antimicrobial Agents and Chemotherapy 49, 3776–3783.CrossRefGoogle ScholarPubMed
Aslett, M., Aurrecoechea, C., Berriman, M., Brestelli, J., Brunk, B. P., Carrington, M., Depledge, D. P., Fischer, S., Gajria, B., Gao, X., Gardner, M. J., Gingle, A., Grant, G., Harb, O. S., Heiges, M., Hertz-Fowler, C., Houston, R., Innamorato, F., Iodice, J., Kissinger, J. C., Kraemer, E., Li, W., Logan, F. J., Miller, J. A., Mitra, S., Myler, P. J., Nayak, V., Pennington, C., Phan, I., Pinney, D. F., Ramasamy, G., Rogers, M. B., Roos, D. S., Ross, C., Sivam, D., Smith, D. F., Srinivasamoorthy, G., Stoeckert, C. J. Jr., Subramanian, S., Thibodeau, R., Tivey, A., Treatman, C., Velarde, G. and Wang, H. (2010). TriTrypDB: a functional genomic resource for the Trypanosomatidae. Nucleic Acids Research 38 (Database issue), D457–D462.CrossRefGoogle ScholarPubMed
Azizi, H., Hassani, K., Taslimi, Y., Najafabadi, H. S., Papadopoulou, B. and Rafati, S. (2009). Searching for virulence factors in the non-pathogenic parasite to humans Leishmania tarentolae . Parasitology 136, 723–735.CrossRefGoogle ScholarPubMed
Beverley, S. B. (2000). Protozoan expression system WO 2000058483 A2. http://www.google.com/patents/WO2000058483A2 Google Scholar
Bolhassani, A., Taheri, T., Taslimi, Y., Zamanilui, S., Zahedifard, F., Seyed, N., Torkashvand, F., Vaziri, B. and Rafati, S. (2011). Fluorescent Leishmania species: development of stable GFP expression and its application for in vitro and in vivo studies. Experimental Parasitology 127, 637–645.CrossRefGoogle ScholarPubMed
Breitling, R., Klingner, S., Callewaert, N., Pietrucha, R., Geyer, A., Ehrlich, G., Hartung, R., Müller, A., Contreras, R., Beverley, S. M. and Alexandrov, K. (2002). Non-pathogenic trypanosomatid protozoa as a platform for protein research and production. Protein Expression and Purification 25, 209–218.CrossRefGoogle ScholarPubMed
Buckner, F. S. and Wilson, A. J. (2005). Colorimetric assay for screening compounds against Leishmania amastigotes grown in macrophages. American Journal of Tropical Medicine and Hygiene 72, 600–605.CrossRefGoogle ScholarPubMed
Chan, M. M., Bulinski, J. C., Chang, K. P. and Fong, D. (2003). A microplate assay for Leishmania amazonensis promastigotes expressing multimeric green fluorescent protein. Parasitology Research 89, 266–271.CrossRefGoogle ScholarPubMed
Den Boer, M., Argaw, D., Jannin, J. and Alvar, J. (2011). Leishmaniasis impact and treatment access. Clinical Microbiology and Infection 17, 1471–1477.CrossRefGoogle ScholarPubMed
Finney, D. J. (1978). Statistical Method in Biological Assay, 3rd Edn. Charles Griffin & Co., London and High Wycombe, 508.Google Scholar
Henao, H. H., Osorio, Y., Saravia, N. G., Gómez, A. and Travi, B. (2004). [Efficacy and toxicity of pentavalent antimonials (Glucantime and Pentostam) in an American cutaneous leishmaniasis animal model: luminometry application]. Biomedica 24, 393–402.CrossRefGoogle Scholar
Hoyer, C., Zander, D., Fleischer, S., Schilhabel, M., Kroener, M., Platzer, M. and Clos, J. A. (2004). Leishmania donovani gene that confers accelerated recovery from stationary phase growth arrest. International Journal of Parasitology 34, 803–811.CrossRefGoogle ScholarPubMed
Kamau, S. W., Grimm, F. and Hehl, A. B. (2001). Expression of green fluorescent protein as a marker for effects of antileishmanial compounds in vitro . Antimicrobial Agents and Chemotherapy 45, 3654–3656.CrossRefGoogle ScholarPubMed
Lang, T., Goyard, S., Lebastard, M. and Milon, G. (2005). Bioluminescent Leishmania expressing luciferase for rapid and high throughput screening of drugs acting on amastigote-harbouring macrophages and for quantitative real-time monitoring of parasitism features in living mice. Cellular Microbiology 7, 383–392.CrossRefGoogle ScholarPubMed
Okuno, T., Goto, Y., Matsumoto, Y., Otsuka, H. and Matsumoto, Y. (2003). Applications of recombinant Leishmania amazonensis expressing egfp or the beta-galactosidase gene for drug screening and histopathological analysis. Experimental Animals 52, 109–118.CrossRefGoogle ScholarPubMed
Papadopoulou, B. and Dumas, C. (1997). Parameters controlling the rate of gene targeting frequency in the protozoan parasite Leishmania . Nucleic Acids Research 25, 4278–4286.CrossRefGoogle ScholarPubMed
Pulido, S. A., Muñoz, D. L., Restrepo, A. M., Mesa, C. V., Alzate, J. F., Vélez, I. D. and Robledo, S. M. (2012). Improvement of the green fluorescent protein reporter system in Leishmania spp. for the in vitro and in vivo screening of antileishmanial drugs. Acta Tropica 122, 36–45.CrossRefGoogle ScholarPubMed
Raymond, F., Boisvert, S., Roy, G., Ritt, J. F., Légaré, D., Isnard, A., Stanke, M., Olivier, M., Tremblay, M. J., Papadopoulou, B., Ouellette, M. and Corbeil, J. (2012). Genome sequencing of the lizard parasite Leishmania tarentolae reveals loss of genes associated to the intracellular stage of human pathogenic species. Nucleic Acids Research 40, 1131–1147.CrossRefGoogle Scholar
Rice, P., Longden, I. and Bleasby, A. (2000). EMBOSS: the European Molecular Biology Open Software Suite. Trends in Genetics 16, 276–277.CrossRefGoogle ScholarPubMed
Rios, L. A., Ocampo, R., Duque, S. M., Robledo, S. M., Velez, I. D., Cedeño, D. L. and Jones, M. A. (2015). Quaternary N-(halomethyl) ammonium salts as therapeutic agents US20140194640 A1. http://www.google.com/patents/US20140194640 Google Scholar
Robledo, S. M., Valencia, A. Z. and Saravia, N. G. (1999). Sensitivity to Glucantime of Leishmania Viannia isolated from patients prior to treatment. Journal of Parasitology 85, 360–366.CrossRefGoogle ScholarPubMed
Roy, G., Dumas, C., Sereno, D., Wu, Y., Singh, A. K., Tremblay, M. J., Ouellette, M., Olivier, M. and Papadopoulou, B. (2000). Episomal and stable expression of the luciferase reporter gene for quantifying Leishmania spp. infections in macrophages and in animal models. Molecular and Biochemical Parasitology 110, 195–206.CrossRefGoogle ScholarPubMed
Sereno, D., Roy, G., Lemesre, J. L., Papadopoulou, B. and Ouellette, M. (2001). DNA transformation of Leishmania infantum axenic amastigotes and their use in drug screening. Antimicrobial Agents and Chemotherapy 45, 1168–1173.CrossRefGoogle ScholarPubMed
Sereno, D., Cordeiro da Silva, A., Mathieu-Daude, F. and Ouaissi, A. (2007). Advances and perspectives in Leishmania cell based drug-screening procedures. Parasitology International 56, 3–7.CrossRefGoogle ScholarPubMed
Singh, N. and Dube, A. (2004). Short report: fluorescent Leishmania: application to anti-leishmanial drug testing. American Journal of Tropical Medicine and Hygiene 71, 400–402.CrossRefGoogle ScholarPubMed
Singh, N., Gupta, R., Jaiswa, L. A. K., Sundar, S. and Dube, A. (2009). Transgenic Leishmania donovani clinical isolates expressing green fluorescent protein constitutively for rapid and reliable ex vivo drug screening. Journal of Antimicrobial Chemotherapy 64, 370–374.CrossRefGoogle ScholarPubMed
Taylor, V. M., Muñoz, D. L., Cedeño, D. L., Vélez, I. D., Jones, M. A. and Robledo, S. M. (2010). Leishmania tarentolae: utility as an in vitro model for screening of antileishmanial agents. Experimental Parasitology 126, 471–475.CrossRefGoogle Scholar
U.S. Department of Health and Human Services (2009). Biosafety in Microbiological and Biomedical Laboratories, 5th edn. HHS Publication No. (CDC) 21-1112. December 2009. http://www.cdc.gov/biosafety/publications/bmbl5/bmbl.pdf Google Scholar
Varela, M. R. E., Muñoz, D. L., Robledo, S. M., Kolli, B. K., Dutta, S., Chang, K. P. and Muskus, C. (2009). Leishmania (Viannia) panamensis: an in vitro assay using the expression of GFP for screening of antileishmanial drug. Experimental Parasitology 122, 134–139.CrossRefGoogle Scholar
World Health Organization (2010). Control of the Leishmaniasis: Report of a Meeting of the WHO Expert Committee on the Control of Leishmaniases. WHO Technical Report. Series No. 949. World Health Organization, Geneva, Switzerland.Google Scholar