Comparison of Proteome Profiling of Two Sensitive and Resistant Field Iranian Isolates of Leishmania major to Glucantime® by 2- Dimensional Electrophoresis.
AbstractBackground: In this study, two-dimensional gel electrophoresis (2-DE) method was applied to determine and compare the protein spots expressed in the two field isolates of Leishmania major and recovered from the patients who were clinically sensitive and resistant to Glucantime® treatment. Methods:Leishmania parasites were isolated from the cutaneous lesions of two CL infected patients in Shiraz, south of Iran. The species of the two isolates were identified as L. major using Nested-PCR. Sensitivity (Sh-214S) and resistance (Sh-120R) of the two isolates to meglumine antimonite were checked by the standard in vitro assays. Both sensitive and resistant L. major isolates were harvested in RPMI 1640 medium. Protein extractions were performed using TCA/Acetone method and the protein spots were determined by a two-dimensional gel electrophoresis (2-DE). The gels were stained with silver nitrate and analyzed by Image Master 2D Melanie-6 software.Results:About 2967 protein spots were detected. Overall, 89 protein spots repre-sented considerable changes of expression in the resistant isolate of L. major compared to the sensitive isolate. Of these, 60 and 29 protein spots were up-and down regulated, respectively. In addition, 11 protein spots present in the resistant isolate were noticed to be absent in the sensitive isolate.Conclusion:A number of proteins showed significant changes of expression in the drug-resistant L. major; moreover, the roles of these proteins probably enhanced the parasite resistance to the drug and increased parasite survival in the cells.
Alvar J, Vélez ID, Bern C, Herrero M, Desjeux P, Cano J, et al. Leishmaniasis worldwide and global estimates of its incidence. PLoS One. 2012; 7 (5): e35671.
Nadim A, Javadian E, Mohebali M, Zamen Momeni A. Leishmania parasites and Leishman-iasis, in Epidemiology of Leishmaniasis in Iran. Tehran University Press. 2009 (Persian).
Hajjaran H, Mohebali M, Mamishi S, Vasigheh F, Oshaghi MA, Naddaf SR, et al. Molecular identification and polymorphism determination of cutaneous and visceral leishmaniasis agents isolated from human and animal hosts in Iran. Bio Med Res Int. 2013;http: //dx.doi.org /10.1155 /2013/ 789 326.
Mohebali M, Javadian E, Yaghoobi-Ershadi MR, Akhavan AA, Abaei MR, Hajjaran H. Study on Leishmania infection in caught rodents in some parts of I.R. of Iran. East Mediterr Health J. 2004; 10: 591- 599.
Hajjaran H, Mohebali M, Teimouri A, Oshaghi MA, Mirjalali H, Kazemi-Rad E, et al. Identifi-cation and Phylogenetic Relationship of Iranian Strains of Various Leishmania Species Isolated from Cutaneous and Visceral Cases of Leish-maniasis Based on N-Acetyl Glucosamine-1-phosphate Transferase Gene. Infect Genet Evol. 2014; 26: 203- 212.
Mohebali M, Fotouhi A, Hooshmand B, Zarei Z, Akhoundi B, Rahnemac A, et al. Compari-son of miltefosine and meglumine antimoniate for the treatment of zoonotic cutaneous leish-maniasis (ZCL) by a randomized clinical trial in Iran. Acta Tropica. 2007; 103 (1):33-40.
Croft SL, Sundar S, Fairlamb AH. Drug re-sistance in leishmaniasis. Clin Microbiol Rev. 2006; 19 (1):111- 126.
Hadighi R, Mohebali M, Boucher P, Hajjaran H, Khamesipour A, Ouellette M. Unrespon-siveness to Glucantime treatment in Iranian cu-taneous leishmaniasis due to drug resistant Leishmania tropica parasites. PLoS Med. 2006; 3 (5): e162.
Asilian A, Sadeghinia S, Faghihi G, Momeni AZ. Comparative study of the efficacy of com-bined cryotherapy and intralesional meglumine antimoniate vs. cryotherapy and interalesional meglumine antimoniate alone for the treatment of cutaneous leishmaniasis. Int J Dermatol. 2004; 43 (4): 281- 283.
Vergnes B, Gourbal B, Girard I, Sundar S, Drummelsmith J, Ouellette M. A proteomics screen implicates HSP 83 and a small kineto-plastid calpain related protein in drug resistance in Leishmania donovani clinical field isolates by Modulating Drug induced Programmed cell death. Mol Cell Proteomics. 2007; 6(1): 88-101.
Ashutosh A, SundarSh, Goyal N. Molecular mechanisms of antimony resistance in Leishma-nia. J Med Microbiol. 2007; 56:143-153.
Vincensini L, Richert S, Blisnick T, Van Dorsselaer A, Leize- Wagner E, Rabilloud T, et al. Proteomic analysis identifies novel proteins of the Maurer's clefts, a secretory compartment delivering Plasmodium falciparum proteins to the surface of its host cell. Mol Cell Proteomics. 2005; 4 (4): 582- 93.
Kucknoor AS, Mundodi V, Alderete JF. The proteins secreted by Trichomonas vaginalis and vaginal epithelial cell response to secreted and episomally expressed AP65. Cell Microbiol. 2007; 9 (11): 2586- 97.
Ringqvist E, Palm JE, Skarin H, Hehl AB, Weiland M, Davids BJ, et al. Release of meta-bolic enzymes by Giardia in response to interac-tion with intestinal epithelial cells. Mol Bio-chem Parasitol. 2008 (2); 159: 85- 91.
Soong L, Chang CH, Sun J, Longley BJ, Rud-dle NH, Flavell RA, et al. Role of CD4+ T-cells in pathogenesis associated with Leishmania amazonensis infection. J Immunol. 1997; 158 (11): 5374- 83.
Drummelsmith J, Brochu V, Girard I, Messier N, Ouellette M. Proteome mapping of the pro-tozoan parasite Leishmania and application to the study of drug targets and resistance mecha-nisms. Mol Cell Proteomics. 2003; 2 (3):146- 55.
Gongora R, Acestor N, Quadroni M, Fasel N, Saravia NG, Walker J. Mapping the proteome of Leishmania Viannia parasites using two-dimensional polyacrylamide gel electrophoresis and associated technologies. Biomedica. 2003; 23 (2):153- 60.
Sarkari B, Bavarsad Ahmadpour N, Motazedi-an MH, Mirjalali H, Akhoundi M, et al. Inter and Intra-Specific variations of Leishmania Strains Isolated from Cutaneous and Visceral Leishmaniasis Patients in Fars Province, South of Iran. Iran J Med Sci. 2014 (under publish).
Maraghi S, Mardanshah O, Rafiei A, Samar-bafzadeh A, Vazirianzadeh B. Identification of Cutaneous Leishmaniasis Agents in Four Geo-graphical Regions of Khuzestan Province Us-ing Nested PCR. Jundishapur J Microbiol. 2013; 6(4):e4866.
Sharifi F, Sharifi I, Zarean M, Hakimi- Parizi M, Aflatoonian MR, Fasihi Harandi M, et al. Spa-tial distribution and molecular identification of Leishmania species from endemic foci of South- Eastern Iran. Iran J Parasitol. 2012; 7(1): 45- 52.
Kazemi-Rad E, Mohebali M, Khadem- Erfan MB, Saffari M, Raoofian R, Hajjaran H, et al. Identification of antimony resistance markers in Leishmania tropica field isolates through a cDNA-AFLP approach. Exp Parasitol. 2013; 135: 344- 349.
Khadem- Erfan MB, Mohebali M, Kazemi-Rad E et al. Down regulation of calcineurin gene is associated with Glucantime® resistance in Leishmania infantum. Iran J Parasitol. 2013; 8 (3): 359- 366.
Damerval C, De Vienne D, Zivy M, Thiel-lement H. Technical improvement two dimen-sional electrophoresis increase the level of ge-netic variation detected in wheat-seedling pro-teins. Electrophoresis. 1986; 7 (1): 52- 54.
Hajjaran H, Mohebali M, Assareh A, Heidari M, Hadighi R. Protein profiling on Meglumine Antimoniate (Glucantime®) sensitive and re-sistant L. tropica isolates by 2-dimentional gel electrophoresis: A preliminary study. Iran J Parasitol. 2009; 4(1): 8- 14.
Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Anal Biochem. 1976; 72: 248- 254.
Shevchenko A, Wilm M, Vorm O, Mann M. Mass spectrometric sequencing of proteins sil-ver stained polyacrylamide gels. Anal Chem. 1996; 68 (5): 850- 858.
Bassam BJ, Caetano Anolles G, Gresshoff PM. Fast and sensitive silver staining of DNA in polyacrylamide gels. Anal Biochem. 1991; 196 (1): 80- 3.
Kedzierski L, Sakthianandeswaren A, Curtis JM, Andrews PC, Junk PC, Kedzierska K. Leishmaniasis: current treatment and prospects for new drugs and vaccines. Curr Med Chem. 2009; 16 (5): 599- 614.
Reynolds KB, Fang CM, Xiadong C, Lynn S. Comparative two-dimensional gel electropho-resis maps for promastigotes of Leishmania amazonensis and L. major. Brazilian J Infec Dis. 2006; 10 (1): 1- 6.
Acestor N, Masina S, Walker J, Saravia NG, Fasel N, Quadroni. Establishing two dimen-sional gels for the analysis of Leishmania proteo-mes. Proteomics. 2002; 2 (7): 877- 9.
Kazemi- Rad E, Mohebali M, Khadem- Erfan MB, Hajjaran H, Hadighi R, Khamesipour A, et al. Overexpression of ubiquitin and amino acid permease genes in association with anti-mony resistance in Leishmania tropica field iso-lates. Korean J Parasitol. 2013; 51(4): 413- 419.
Hajjaran H, Azarian B, Mohebali M, Hadighi R., Assareh A, Vaziri B. Comparative proteomics study on meglumine antimoniate sensitive and resistant Leishmania tropica isolated from Iranian anthroponotic cutaneous
leishmaniasis patients. East Mediterr Health J. 2012;18 (2):165-171.
Cordwell SJ, Nouwens AS, Walsh BJ. Compar-ative proteomics of bacterial pathogens. Prote-omics. 2001; 1 (4): 461- 72.
Das VN, Ranjan A, Bimal S, Siddique NA, Pandey K, Kumar N, et al. Magnitude of unre-sponsiveness to sodium stibogluconate in the treatment of visceral leishmaniasis in Bihar. Natl Med J India. 2005; 18 (3):131-133.
Ivens AC, Peacock CS, Worthey EA, Murphy L, Aggarwal G, Berriman M, et al. The genome of the kinetoplastid parasite, Leishmania major. Science. 2005; 309 (5733): 436- 42.
Kamoun-Essghaier S, Guizani I, Strub JM, Van Dorsselaer A, Mabrouk K, Ouelhazi L, Dellagi K. Proteomic approach for characteri-zation of immune dominant membrane associ-ated 30 to 36 kilodalton fraction antigens of Leishmania infantum promastigotes, reacting with sera from Mediterranean visceral leishmaniasis patients. Clin Diagn Lab Immunol. 2005; 12 (2): 310- 20.
Copyright (c) Iranian Journal of Parasitology
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.