اخبار و اعلانات

 آمار

تعداد نشریات284
تعداد نشریات سازمان82
تعداد شماره‌ها3,037
تعداد مقالات33,984
تعداد مشاهده مقاله45,779,998
تعداد دریافت فایل اصل مقاله77,192,479
bakhshandeh, nina, Tebianian, Majid, Khaki, Pejvak, Esmaelizad, Majid, Saadatmand, Maryam. (1403). Design, Development and Immunogenic Evaluation of a Novel Lipl32 Recombinant Protein of Local Pathogenic Leptospira Serovars. سامانه مدیریت نشریات علمی, 80(1), 103-115. doi: 10.32592/ARI.2025.80.1.103
nina bakhshandeh; Majid Tebianian; Pejvak Khaki; Majid Esmaelizad; Maryam Saadatmand. "Design, Development and Immunogenic Evaluation of a Novel Lipl32 Recombinant Protein of Local Pathogenic Leptospira Serovars". سامانه مدیریت نشریات علمی, 80, 1, 1403, 103-115. doi: 10.32592/ARI.2025.80.1.103
bakhshandeh, nina, Tebianian, Majid, Khaki, Pejvak, Esmaelizad, Majid, Saadatmand, Maryam. (1403). 'Design, Development and Immunogenic Evaluation of a Novel Lipl32 Recombinant Protein of Local Pathogenic Leptospira Serovars', سامانه مدیریت نشریات علمی, 80(1), pp. 103-115. doi: 10.32592/ARI.2025.80.1.103
bakhshandeh, nina, Tebianian, Majid, Khaki, Pejvak, Esmaelizad, Majid, Saadatmand, Maryam. Design, Development and Immunogenic Evaluation of a Novel Lipl32 Recombinant Protein of Local Pathogenic Leptospira Serovars. سامانه مدیریت نشریات علمی, 1403; 80(1): 103-115. doi: 10.32592/ARI.2025.80.1.103

Design, Development and Immunogenic Evaluation of a Novel Lipl32 Recombinant Protein of Local Pathogenic Leptospira Serovars

Archives of Razi Institute
مقاله 12، دوره 80، شماره 1، فروردین و اردیبهشت 2025، صفحه 103-115    اصل مقاله (929.65 K)
نوع مقاله: Original Articles
شناسه دیجیتال (DOI): 10.32592/ARI.2025.80.1.103
نویسندگان
nina bakhshandeh* 1؛ Majid Tebianian2؛ Pejvak Khaki3؛ Majid Esmaelizad4؛ Maryam Saadatmand5
1Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Ira
2Department of human vaccines and serum, Razi Institute, Karaj, Iran
3Department of Microbiology, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
4Department of Biotechnology, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran member
5Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran
چکیده
Leptospirosis is a significant yet often ignored zoonotic disease found globally, especially in tropical and subtropical regions. Its symptoms frequently lead to misdiagnosis as they resemble those of other infectious diseases. The design of rapid diagnostic tests or detection of possible vaccine candidates against leptospirosis can be challenging. Surface-exposed antigens are located on the outer layer of Leptospira, and these virulence factors likely contribute to the initial interactions between the host and the pathogen.Lipl32 is highly conserved and exclusively produced by pathogenic Leptospires and has a significant role as a prominent immunogen in leptospirosis. The goal of this study is to establish the ideal conditions for the expression and purification of the rLipl32 protein of Iranian pathogenic leptospira and to assess its ability to stimulate cellular and humoral immune responses.
All Lipl32 protein sequences were analyzed from the NCBI database. Codon sequences of serovars were designed and synthesized, and one local dominant Lipl32 pattern was selected after optimization. The construct was sub-cloned into a pET32a+ vector with His-tag and Trx, then transformed into E.coli (BL21) for expression using IPTG. Then purified, and confirmed by immune blotting. BALB/c Mice (4-6 weeks old) were vaccinated with 3 doses containing 50 mg of rLipl32, at a 14-day interval, and compared with controls. The humoral immune response and the cytokine profile were evaluated using an indirect sandwich ELISA test.
Results showed that rLipl32 protein was highly expressed in the presence of 0.5 mM IPTG after 16 hours of incubation at 22°C. The optimal conditions for the Ni-NTA pull-down involved a one-hour binding period at 37°C, followed by five washing steps and the use of an elution buffer with a pH of 7.4 and a 0.3 mM concentration of imidazole and successfully purified in soluble form. The administration of rLipl32 led to strong total antibody titers (p < 0.05) and dramatically increased cytokine levels (P < 0.05). In conclusion, rLipl32 could strongly stimulate specific humoral and cellular immune responses. It has been suggested that it could be further utilized for immune-dominant Lipl32-based diagnosis and has potential as a candidate to be a subunit vaccine.
کلیدواژه‌ها
Leptospira؛ Recombinant protein LipL32؛ Gene expression؛ Purification؛ Immune Response
مراجع
  1. Langston CE, Heuter KJ. Leptospirosis: A re-emerging zoonotic disease. Veterinary Clinics: Small Animal Practice. 2003;33(4):791-807.
  2. Adler B, de la Peña Moctezuma A. Leptospira and leptospirosis. Veterinary microbiology. 2010;140(3-4):287-96.
  3. Haake D, Levett P. Leptospirosis in humans. Curr To p Microbiol Immunol. 2015; 387: 65-97. doi: 10.1007. 978-3-662-45059.
  4. Torres FDA, Borges ALdSB, Kolesnikovas C, Domit C, Barbosa CB, Carvalho-Costa FA, et al. Pinnipeds carriers of pathogenic Leptospira: New data based on molecular characterization. Research in Veterinary Science. 2023;155:62-8.
  5. Costa F, Hagan JE, Calcagno J, Kane M, Torgerson P, Martinez-Silveira MS, et al. Global morbidity and mortality of leptospirosis: a systematic review. PLoS neglected tropical diseases. 2015;9(9):e0003898.
  6. Bandara M, Ananda M, Wickramage K, Berger E, Agampodi S. Globalization of leptospirosis through travel and migration. Globalization and health. 2014;10:1-9.
  7. Faisal SM, McDonough SP, Chang Y-F. Leptospira: Invasion, pathogenesis and persistence. The pathogenic spirochetes: Strategies for evasion of host immunity and persistence. 2012:143-72.
  8. Marotto PC, Nascimento CM, Eluf-Neto J, Marotto MS, Andrade L, Sztajnbok J, et al. Acute lung injury in leptospirosis: clinical and laboratory features, outcome, and factors associated with mortality. Clinical Infectious Diseases. 1999;29(6):1561-3.
  9. Schreier S, Doungchawee G, Chadsuthi S, Triampo D, Triampo W. Leptospirosis: current situation and trends of specific laboratory tests. Expert review of clinical immunology. 2013;9(3):263-80.
  10. Martínez R, Pérez A, Cruz R, Alvarez A, Armesto M, Fernández C, et al. Efficacy and safety of a vaccine against human leptospirosis in Cuba. Revista panamericana de salud publica= Pan American journal of public health. 2004;15(4):249-55.
  11. Fitzgerald J. Availability of leptospirosis vaccine. The Veterinary Record. 2009;164(5):157.
  12. Koizumi N, Watanabe H. Leptospirosis vaccines: past, present, and future. Journal of postgraduate medicine. 2005;51(3):210.
  13. Klaasen H, Molkenboer M, Vrijenhoek M, Kaashoek M. Duration of immunity in dogs vaccinated against leptospirosis with a bivalent inactivated vaccine. Veterinary microbiology. 2003;95(1-2):121-32.
  14. Barazzone GC, Teixeira AF, Azevedo BO, Damiano DK, Oliveira MP, Nascimento AL, et al. Revisiting the development of vaccines against pathogenic Leptospira: innovative approaches, present challenges, and future perspectives. Frontiers in immunology. 2022;12:760291.
  15. Haake DA, Zückert WR. The leptospiral outer membrane. Leptospira and Leptospirosis. 2014:187-221.
  16. Luo D, Xue F, Ojcius DM, Zhao J, Mao Y, Li L, et al. Protein typing of major outer membrane lipoproteins from Chinese pathogenic Leptospira spp. and characterization of their immunogenicity. Vaccine. 2009;28(1):243-55.
  17. Yan Y, Chen Y, Liou W, Ding J, Chen J, Zhang J, et al. An evaluation of the serological and epidemiological effects of the outer envelope vaccine to leptospira. Journal of the Chinese Medical Association: JCMA. 2003;66(4):224-30.
  18. Grassmann AA, Kremer FS, Dos Santos JC, Souza JD, Pinto LdS, McBride AJA. Discovery of novel leptospirosis vaccine candidates using reverse and structural vaccinology. Frontiers in immunology. 2017;8:463.
  19. Chalayon P, Chanket P, Boonchawalit T, Chattanadee S, Srimanote P, Kalambaheti T. Leptospirosis serodiagnosis by ELISA based on recombinant outer membrane protein. Transactions of the Royal Society of Tropical Medicine and Hygiene. 2011;105(5):289-97.
  20. Fernandes LG, Avelar KE, Romero EC, Heinemann MB, Kirchgatter K, Nascimento AL. A new recombinant multiepitope chimeric protein of Leptospira interrogans is a promising marker for the serodiagnosis of leptospirosis. Tropical Medicine and Infectious Disease. 2022;7(11):362.
  21. Cullen PA, Haake DA, Adler B. Outer membrane proteins of pathogenic spirochetes. FEMS microbiology reviews. 2004;28(3):291-318.
  22. Aviat F, Rochereau-Roulet S, Branger C, Estavoyer J, Chatrenet B, Orsonneau J, et al. Synthetic peptide issued from Hap1/LipL32 for new early serodiagnosis of human leptospirosis. Comparative immunology, microbiology and infectious diseases. 2010;33(5):375-87.
  23. Haake DA, Chao G, Zuerner RL, Barnett JK, Barnett D, Mazel M, et al. The leptospiral major outer membrane protein LipL32 is a lipoprotein expressed during mammalian infection. Infection and immunity. 2000;68(4):2276-85.
  24. Dey S, Mohan CM, Kumar TS, Ramadass P, Nainar AM, Nachimuthu K. Recombinant LipL32 antigen-based single serum dilution ELISA for detection of canine leptospirosis. Veterinary microbiology. 2004;103(1-2):99-106.
  25. Boonyod D, Poovorawan Y, Bhattarakosol P, Chirathaworn C. LipL32, an outer membrane protein of Leptospira, as an antigen in a dipstick assay for diagnosis of leptospirosis. Asian Pacific Journal of Allergy and Immunology. 2005;23(2-3):133.
  26. Seixas FK, Fernandes CH, Hartwig DD, Conceicao FR, Aleixo JAG, Dellagostin OA. Evaluation of different ways of presenting LipL32 to the immune system with the aim of developing a recombinant vaccine against leptospirosis. Canadian Journal of Microbiology. 2007;53(4):472-9.
  27. Yaakob Y, Rodrigues KF, Opook F, William T, John DV. Recombinant LipL32 protein developed using a synthetic gene detects Leptospira-specific antibodies in human serum samples. The Malaysian journal of medical sciences: MJMS. 2017;24(5):44.
  28. Yeoh TS, Tang TH, Citartan M. Isolation of a novel DNA aptamer against LipL32 as a potential diagnostic agent for the detection of pathogenic Leptospira. Biotechnology Journal. 2023;18(3):2200418.
  29. Govindan P, Manjusha P, Saravanan KM, Natesan V, Salmen SH, Alfarraj S, et al. Expression and preliminary characterization of the potential vaccine candidate LipL32 of leptospirosis. Applied Nanoscience. 2021:1-15.
  30. Fraga TR, Barbosa AS, Isaac L. Leptospirosis: aspects of innate immunity, immunopathogenesis and immune evasion from the complement system. Scandinavian journal of immunology. 2011;73(5):408-19.
  31. Dellagostin OA, Grassmann AA, Hartwig DD, Félix SR, da Silva EF, McBride AJ. Recombinant vaccines against leptospirosis. Human vaccines. 2011;7(11):1215-24.
  32. Reis EA, Hagan JE, Ribeiro GS, Teixeira-Carvalho A, Martins-Filho OA, Montgomery RR, et al. Cytokine response signatures in disease progression and development of severe clinical outcomes for leptospirosis. PLoS neglected tropical diseases. 2013;7(9):e2457.
  33. Qiagen A. handbook for high-level expression and purification of 6xHis-tagged proteins. Qiagen, Chatsworth, Calif, USA. 2003.
  34. Torgerson PR, Hagan JE, Costa F, Calcagno J, Kane M, Martinez-Silveira MS, et al. Global burden of leptospirosis: estimated in terms of disability adjusted life years. PLoS neglected tropical diseases. 2015;9(10):e0004122.
  35. Bharti AR, Nally JE, Ricaldi JN, Matthias MA, Diaz MM, Lovett MA, et al. Leptospirosis: a zoonotic disease of global importance. The Lancet infectious diseases. 2003;3(12):757-71.
  36. Putz EJ, Nally JE. Investigating the immunological and biological equilibrium of reservoir hosts and pathogenic leptospira: balancing the solution to an acute problem? Frontiers in microbiology. 2020;11:2005.
  37. Khaki P. Clinical laboratory diagnosis of human leptospirosis. Int J Enteric Pathog. 2016;4(1):1-7.
  38. Bakhshandeh N, Tebianian M, Khaki P, Esmaelizad M, Saadatmand S. Development of an In-house Indirect ELISA Test as a Diagnostic Tool for Bovine Leptospirosis. International Journal of Enteric Pathogens. 2021;9(2):47-51.
  39. Maia MA, Bettin EB, Barbosa LN, de Oliveira NR, Bunde TT, Pedra ACK, et al. Challenges for the development of a universal vaccine against leptospirosis revealed by the evaluation of 22 vaccine candidates. Frontiers in cellular and infection microbiology. 2022:1503.
  40. Feng C, Li Q, Zhang X, Dong K, Hu B, Guo X. Immune strategies using single-component LipL32 and multi-component recombinant LipL32-41-OmpL1 vaccines against leptospira. Brazilian Journal of Medical and Biological Research. 2009;42:796-803.
  41. Golab N, Khaki P, Harzandi N, Esmaelizad M, Tebianian M. Expression and purification of the Lipl41, a surface-exposed lipoprotein antigen of pathogenic Leptospira spp. Veterinarski arhiv. 2020;90(3):297-305.
  42. Teixeira AF, Cavenague MF, Kochi LT, Fernandes LG, Souza GO, de Souza Filho AF, et al. Immunoprotective activity induced by leptospiral outer membrane proteins in hamster model of acute leptospirosis. Frontiers in Immunology. 2020;11:568694.
  43. Lin Xa, Zhao J, Qian J, Mao Y, Pan J, Li L, et al. Identification of immunodominant B-and T-cell combined epitopes in outer membrane lipoproteins LipL32 and LipL21 of Leptospira interrogans. Clinical and Vaccine Immunology. 2010;17(5):778-83.
  44. Haake DA, Matsunaga J. Leptospiral immunoglobulin-like domain proteins: Roles in virulence and immunity. Frontiers in Immunology. 2021;11:579907.
  45. Hauk P, Macedo F, Romero EC, Vasconcellos SlA, de Morais ZM, Barbosa AS, et al. In LipL32, the major leptospiral lipoprotein, the C terminus is the primary immunogenic domain and mediates interaction with collagen IV and plasma fibronectin. Infection and immunity. 2008;76(6):2642-50.
  46. Vieira ML, Atzingen MV, Oliveira TR, Oliveira R, Andrade DM, Vasconcellos SA, et al. In vitro identification of novel plasminogen-binding receptors of the pathogen Leptospira interrogans. PloS one. 2010;5(6):e11259.
  47. Golab N, Khaki P, Tebianian M, Esmaelizad M, Harzandi N. Recombinant outer membrane protein Lipl41 from Leptospira interrogans robust immune responses in mice model. Iranian Journal of Microbiology. 2024;16(4):545.
  48. Hartwig DD, Bacelo KL, de Oliveira PD, Oliveira TL, Seixas FK, Amaral MG, et al. Mannosylated LigANI produced in Pichia pastoris protects hamsters against leptospirosis. Current microbiology. 2014;68:524-30.
  49. Gharakhani M, Ghasemi MF, Khaki P, Esmaelizad M, Tebianian M. Improvement the Expression and Purification of Loa22: A Lipoprotein with OmpA domain from Pathogenic Leptospira Serovars. 2022.
  50. Plotkin SA. Updates on immunologic correlates of vaccine-induced protection. Vaccine. 2020;38(9):2250-7.
  51. Senavirathna I, Rathish D, Agampodi S. Cytokine response in human leptospirosis with different clinical outcomes: a systematic review. BMC Infectious Diseases. 2020;20(1):1-8.
آمار
تعداد مشاهده مقاله: 143
تعداد دریافت فایل اصل مقاله: 274


counter
سامانه مدیریت نشریات علمی. طراحی و پیاده سازی از سیناوب