DOI: https://doi.org/10.14710/ijfst.15.2.105-111

UJI IN VITRO DAN KARAKTER PROTEIN EKSTRASELULER (ECP) Edwardsiella tarda DENGAN KONSENTRASI ETHANOL BERBEDA (In Vitro Test and Character of Extracellular Protein (ECP) Edwardsiella tarda with Different Ethanol Concentration)

*Cucun Herlina orcid  -  Brawijaya University, Indonesia
Uun Yanuhar  -  Brawijaya University, Indonesia
Maftuch Maftuch  -  Brawijaya University, Indonesia

Citation Format:
Abstract

Protein ekstraseluler (ECP) bakteri memiliki kemampuan imunogenik dan dapat meningkatkan sistem imun tubuh inang. Komponen ECP diantaranya flagellin, protease dan metalloprotease mampu berdifusi pada sel inang untuk mengaktifkan respon imun. Tujuan penelitian adalah untuk mengetahui produksi protein ekstraseluler E. tarda dengan presipitasi ethanol pada konsentrasi berbeda sebagai potensi protein imunogenik secara in vitro. Metode penelitian yaitu kultur bakteri E. tarda, preparasi protein ekstraseluler, presipitasi protein, konsentrasi protein ditentukan denganspektrofotometer dan SDS-PAGE, uji in vitro protein ECP E. tarda, dan uji nilai RPS. Hasil protein ekstraseluler yang diperoleh dengan presipitasi ethanol tertinggi pada konsentrasi 90% sebanyak 5,69 mg/ml dan terendah pada konsentrasi 80% sebanyak 4,81 mg/ml. Hasil karakterisasi protein dengan SDS-SDSPAGE antara 30-60 kDa. Hasil uji in vitro vaksin E. tarda (presipitasi ethanol 100%) baik dari uji viabilitas maupun sterilitas menunjukkan tidak terdapat koloni yang tumbuh. Hasil nilai RPS pada ikan nila (Oreochromis niloticus) mencapai 100% dengan masa pemelihaaraan 9 hari pasca imunisasi.

 

ECP bacteria have immunogenic abilities and can increase the host's immune system. ECP components which are flagellin, protease, metalloprotease can diffuse to host cell for the activation immune response. The study aimed to determine the production of extracellular protein E. tarda with ethanol precipitation at different concentrations as a potential immunogenic protein by in vitro test. The research methods were E. tarda bacterial culture, extracellular protein preparation, protein precipitation, protein concentration determined by spectrophotometer and SDS-PAGE, in vitro ECP E. tarda protein test, and RPS value test. The results of extracellular protein obtained by the highest ethanol precipitation at a concentration of 90% as much as 5.69 mg/ml and the lowest at 80% concentration as much as 4.81 mg/ml. The results of the characterization of proteins with SDS-SDS-PAGE between 30-60 kDa. The results of the in vitro test of E. tarda vaccine (100% ethanol precipitation) from both the viability and sterility tests showed that there were no growing colonies. The results of the RPS in tilapia (Oreochromis niloticus) reach 100% with a maintenance period of 9 days after immunization.

Fulltext View|Download
Keywords: Edwardsiella tarda; Ekstraseluler Protein (ECP); Presipitasi; SDS-PAGE

Article Metrics:

Article Info
Section: Research Articles
Language : ID
Recent articles
PENGEMBANGAN SEKTOR PERIKANAN MENUJU HILIRISASI INDUSTRI UNTUK MENDUKUNG PEMBANGUNAN EKONOMI WILAYAH KABUPATEN SERDANG BEDAGAI PROPINSI SUMATERA UTARA ANALISA PERFORMA GAYA APUNG, GAYA TENGGELAM, DAN KECEPATAN TENGGELAM PURSE SEINE CAKALANG KM BINTANG SURYA (Analysis of Buoyancy, Sinking Force and Sinking Speed of the Skipjack Purse Seines Bintang Surya) UJI IN VITRO DAN KARAKTER PROTEIN EKSTRASELULER (ECP) Edwardsiella tarda DENGAN KONSENTRASI ETHANOL BERBEDA (In Vitro Test and Character of Extracellular Protein (ECP) Edwardsiella tarda with Different Ethanol Concentration) More recent articles
Most cited articles
The Effect of Leaching Times on the Gel Properties of Catfish (Clarias gariepinus) Surimi Aplikasi Teknologi Aquaponic Pada Budidaya Ikan Air Tawar Untuk Optimalisasi Kapasitas Produksi Addition of Phytase Artificial Feed to Increase Digesting, Specific Growth and Survival Rate of Nile Tilapia fingerlings (Oreochromis niloticus) Soaking Period of Cooked Blood Cockle (Anadara granosa) Meat in Alginate Solution to Cadmium Content Reduction STRATEGI PENGEMBANGAN PARIWISATA MANGROVE DI KAWASAN KONSERVASI PERAIRAN NUSA PENIDA (Strategies of Mangrove Tourism Development in Nusa Penida Marine Protected Area) More cited articles
  1. Adriana, F., J. Koščová, D. Mudroňová. 2016 The use of probiotic bacteria against Aeromonas, infections in salmonid aquaculture. Aquaculture. 469 : 1–8. https://doi.org/10.1016/j.aquaculture.2016.11.042
  2. Aliffudin, M. 2002. Imunostimulan pada hewan akuatik. Jurnal Akuakultur Indonesia. 1 (2): 87-92. 10.19027/jai.1.87-92
  3. Aly, TM. 1981. Studies on the Effect of Different Adjuvant on the Efficiency of FMD Vaccine in Farm Animal. Ph. D. faculty of Vet. Med. Zagazig University - Egypt
  4. Anderson DP, Capstiek PB, Mowat GN. 1970. In vitro method for safety of FMD. J. hyg. Gamd. 68: 159-172. 10.1017/S0022172400028643
  5. Bai, F., Z. Li, A. Umezawa, N.Terada, S. Jin. 2018. Bacterial type III secretion system as a protein delivery tool for a broad range of biomedical applications. Biotechnology Advances. 36: 482–493. 10.1016/j.biotechadv.2018.01.016
  6. Bharadwaj, A., T. J. Abraham, S. N. Jordar. 2013. Immune effector activities in challenged rohu, Labeo rohita after vaccinating with Aeromonas bacterin. Aquaculture. 392-395: 16-22. https://doi.org/10.1016/j.aquaculture.2013.01.016
  7. Bollag, D. M and S. J. Edelstein. 1991. Protein Methods. Department of Biochemistry University of Geneva - Switzerland: Wiley-Liss
  8. Caldwell, R. B and C.T Lattemann. 2004. Simple and reliable method to precipitate proteins from bacterial culture supernatant. Applied and Environmental Microbiology. 70 (1): 610-612. 10.1128/AEM.70.1.610-612.2004
  9. Cunningham, A. L., N. Gracon, O. Leo, L. R, Friedland, R. Strugnell, B. Laupeze, M. Doherty dan P. Stern. 2016. Vaccine development: From concept to early clinical testing. Vaccine. 10. https://doi.org/10.1016/j.vaccine.2016.10.016
  10. Ebner, P. and F. Götz. 2019. Bacterial Excretion of Cytoplasmic Proteins (ECP): Occurrence, Mechanism, and Function. Trends in Microbiology. 27 ( 2): 176 -187. https://doi.org/10.1016/j.tim.2018.10.006
  11. El-Gayar, K. E. 2015. Principles of recombinant protein production, extraction and purification from bacterial strains. International Journal of Microbiology and Allied Sciences (IJOMAS). 2(2): 18-33. https://www.researchgate.net/publication/285926620_Principles_of_recombinant_protein_production_extraction_and_purification_from_bacterial_strains
  12. Ellis AE. 1988. General principles of fish vaccination. Di dalam: Ellis AE, editor. Fish vaccination. Academic Press, London, 1- 19 hal
  13. Fang, S., L. Zhang, Y. Lou, D. Yang, Q. Wang, Y. Zhang, Q. Liu. 2016. Intracellular translocation and localization of Edwardsiella tarda type III secretion system effector EseG in host cells. Microbial Pathogenesis, 97: 166-171. https://doi.org/10.1016/j.micpath.2016.05.008
  14. Green, E. R. and J. Mecsas. 2016. Bacterial Secretion Systems. Microbiol Spectr, 4(1): 1-32. 10.1128/microbiolspec.VMBF-0012-2015
  15. Griffiths, K. L. and S. A Khader. 2014. Novel vaccine approaches for protection against intracellular pathogens. Curr Opin Immunol, 0: 58–63. 10.1016/j.coi.2014.02.003
  16. Hamed, K. Y., B. A. Siame, B. J. Tenkink, R. J. Noort and Y. K. Mok. 2012. Edwardsiella tarda: Virulence mechanisms of an emerging gastroenteritis pathogen. Microbes and Infection. 14 : 26-34. https://doi.org/10.1016/j.micinf.2011.08.005
  17. Hamed, S. B., M. J.T. R. Paiva, L. Tachibana, D. de C. Dias, C. M. Ishikawa and M. A. Esteban. 2018. Fish pathogen bacteria: Adhesion, parameters influencing virulence and interaction with host cells. Fish & Shellfish Immunology. 80: 550-562. https://doi.org/10.1016/j.fsi.2018.06.053
  18. Hardi, E. H., Sukenda, E. Harris dan A. M. Lusiastuti. 2011. Toksisitas Produk Ekstrasellular (ECP) Streptococcus agalactiae pada Ikan Nila (Oreochromis niloticus). Jurnal Natur Indonesia. 13(3): 187-199. ://dx.doi.org/10.31258/jni.13.03.%25p
  19. Ibrahem, M. D., I.Shaheed, H. A. El-Yazeed and H. Korani. 2011. Assessment of the susceptibility of polyculture reared African Catfish and Nile tilapia to Edwardsiella tarda. Journal of American Science. 7(3): 779 – 786. http://www.jofamericanscience.org/journals/am-sci/am0703/93_5084am0703_779_786
  20. Jiao, X., W. Dang, Y. Hu, and L. Sun. 2009. Identification and immunoprotective analysis of an in vivo-induced Edwardsiella tarda antigen. Fish & Shellfish Immunology. 27: 633–638. 10.1016/j.fsi.2009.08.006
  21. Kamiso, H. N. 1990. Pemvaksinan Penyakit Bakteri pada Ikan. PAU – Bioteknologi, Universitas Gajah Mada, Yogyakarta.67-68 hlm
  22. Kumar, V., D. V. Nguyen, K. Baruah, and P. Bossier. 2019. Probing the mechanism of VPAHPND extracellular proteins toxicity purified from Vibrio parahaemolyticus AHPND strain in germ-free Artemia test system. Aquaculture. 504: 414-419. https://doi.org/10.1016/j.aquaculture.2019.02.029
  23. Leung, K. Y., B. A. Siame, B. J. Tenkink, R. J. Noort and Y. K. Mok. 2012. Edwardsiella tarda: Virulence mechanisms of an emerging gastroenteritis pathogen. Microbes and Infection. 14 : 26-34. https://doi.org/10.1016/j.micinf.2011.08.005
  24. Liu, X., L. Kang, Y. Liu, H. Li, X. Peng. 2013. Characterization of the Edwardsiella tarda proteome in response to different environmental stresses. Journal Of Proteomics. 8 : 320 – 333. 10.1016/j.jprot.2013.01.022
  25. Mirica, K. A., M. R. Lockett, P. W. Snyder, N. D. Shapiro, E. T. Mack, S. Nam, and G. M. Whitesides. 2012. Selective Precipitation and Purification of Monovalent Proteins Using Oligovalent Ligands and Ammonium Sulfate. Bioconjugate Chem. 23: 293−299. 10.1021/bc200390q
  26. Mohamed, L. A dan W. S. E. D. Soliman. 2013. Development and efficacy of fish vaccine used against some bacterial diseases in farmed tilapia. Nature and Science. 11 (6): 120-128. http://www.sciencepub.net/nature/ns1106/014_18280ns1106_120_128.pdf
  27. Mohanty, B. R. and P. K. Sahoo. 2007. Edwardsiellosis in fish: a brief review. J. Biosci. 32 (7) : 1331–1344. 10.1007/s12038-007-0143-8
  28. Narwiyani, S dan Kurniasih. 2011. Perbandingan patogenesitas, Edwardsiella tarda pada ikan mas koki (Charassius auratus) dan ikan celebes rainbow (Telmatherina celebensis). Jurnal Ris. Akuakultur. 6(2): 291-301. http://dx.doi.org/10.15578/jra.6.2.2011.291-301
  29. Nooralabetu, K. P.. 2014. Optimasitaion of ammonium sulfate precipitation method to achieve high throughput concentration of crude alkaline phosphatase from Brown shrimp (Metapenaeus Monoceros) hepatopancreas. International Journal Ana Bio-Sci, 2(1): 7- 16. https://plaza.umin.ac.jp/~e-jabs/2/2.7.pdf
  30. Putri, R. A., Wardiyanto dan A. Setyawan. 2013. Penyimpanan vaksin inaktif whole cell Aeromonas salmonicida dengan penambahan gliserol. Jurnal Rekayasa dan Teknologi Budidaya Perairan, 1 (2): 79-86. https://media.neliti.com/media/publications/233517-penyimpanan-vaksin-inaktif-whole-cell-ae-dcbdd020.pdf
  31. Rosidah dan W. M. Afizia. 2012. Potensi ekstrak daun jambu biji sebagai antibakterial untuk menanggulangi serangan bakteri Aeromonas hydrophila pada ikan gurame (Osphronemus Gouramy lacepede). Jurnal Akuatika. III (1): 19-27. http://jurnal.unpad.ac.id/akuatika/article/view/473
  32. Saleh, W. D. 2005. Isolation and identification of Edwardsiella tarda from infected nile tilapia fish. Bull Fac Agric. 56: 839-846. https://www.researchgate.net/publication/265168496_Isolation_and_Identification_of_Edwardsiella_tarda_from_Infected_Nile_Tilapia_Fish_Oreochromis_niloticus
  33. Sanchez, B., M. C. Urdaci and A. Margolles. 2010. Extracellular proteins secreted by probiotic bacteria as mediators of effects that promote mucosa– bacteria interactions. Microbiology. 156: 3232–3242. 10.1099/mic.0.044057-0
  34. Sanchez, B., S. Chaignepain, J.Schmitter and M. C. Urdaci. 2009. A method for the identification of proteins secreted by lactic acid bacteria grown in complex media. FEMS Microbiol Lett. 295: 226-229. 10.1111/j.1574-6968.2009.01599.x
  35. Setiawan Raden B., Dulm’iad., dan Rosidah. 2013. Efektivitas Vaksin dari Bakteri Mycobacterium fortuitum yang Diinaktivasi dengan Pemanasan untuk Pencegahan Penyakit Mycobacteriosis Pada Ikan Gurami (Osphronemus gouramy). Jurnal Perikanan Dan Kelautan. 3(1): 25-40. http://jurnal.unpad.ac.id/jpk/article/view/3528
  36. Silva, B. C., M. L. Martins, A. Jatobá, C. C. B. Neto, F. N. Vieira, G. V. Pereira, G. T. Jerônimo, W. Q. Seiffert and J. L. P. Mouriño. 2009. Hematological and immunological responses of Nile tilapia after polyvalent vaccine administration by different routes. Pesq. Vet. Bras. 29(11): 874-880. http://dx.doi.org/10.1590/S0100-736X2009001100002
  37. Song, M., J. Xie, X. Peng, and H. Li. 2013. Identification of protective immunogens from extracellular secretome of Edwardsiella tarda. Fish & Shellfish Immunology. 35: 1932-1936. 10.1016/j.fsi.2013.09.033
  38. Song, M., Y. Kang, D. Zhang, L. Chen, J. Bi, H. Zhang, L. Zhang, A. Qian, X.Shan. 2018. Immunogenicity of extracellular products from an inactivated vaccine against Aeromonas veronii TH0426 in koi, Cyprinus carpio. Fish and Shellfish Immunology. 81:176–181. https://doi.org/10.1016/j.fsi.2018.07.004
  39. Sun, Y., C.Liu, and L. Sun. 2010. Identification of an Edwardsiella tarda surface antigen and analysis of its immunoprotective potential as a purified recombinant subunit vaccine and a surface-anchored subunit vaccine expressed by a fish commensal strain. Vaccine. 28: 6603–6608. https://doi.org/10.1016/j.vaccine.2010.07.050
  40. Tizard, I. R. 1988. Pengantar Imunologi Veteriner. Terjemahan: Partadireja M. Surabaya: Airlangga University
  41. Todar, K. 2012. Bacterial protein toxins. Todar’s online textbook of bacteriology. Madison, Wisconsin
  42. Trilia, N. A. O., A. Setyawan, Y. T. Adiputra dan Wardiyanto. 2014. Imunogenisitas kombinasi vaksin inaktif whole cell Aeromonas salmonicida dan jintan hitam (Nigella sativa) pada ikan mas (Cyprinus carpio). Jurnal Rekayasa dan Teknologi Budidaya Perairan. (2): 249-257. http://jurnal.fp.unila.ac.id/index.php/bdpi/article/view/396
  43. Wibawan, I. W. T. dan R. D. Soejoedono. 2013. Intisari Imunologi medis. Fakultas Kedokteran Hewan, IPB. Bogor. 2-3 dan 26 hlm
  44. Wingfield, P. T. 2001. Protein precipitation using ammonium sulfate. Curr Protoc Protein Sci. 1-9. 10.1002/0471140864.psa03fs84
  45. Wu, L., Y. Jiang, Q. Tang, H. Lin, C. Lu, and H. Yao. 2012. Development of an Aeromonas hydrophila recombinant extracellular protease vaccine. Microbial Pathogenesis. 53. 183-188. 10.1016/j.micpath.2012.07.007
  46. Zhang, D., J. W. Pridgeon and P. H. Klesius. 2014. Vaccination of channel catfish with extracellular product of Aeromonas hydrohpyla provides protection against infection by the pathogen. Fish and Shellfish Immunology. 36: 270-275. https://doi.org/10.1016/j.fsi.2013.11.015
  47. Zhang, N and K. S. Nandakumar. 2018. Recent advances in the development of vaccines for chronic inflammatory autoimmune diseases. Vaccine. 36: 3208–3220. https://doi.org/10.1016/j.vaccine.2018.04.062
  48. Zhou, Y., L. Y. Liua, T. T. He, Z. A. Laghari, P. Nie, Q. Gao, H. X. Xie. 2016. Edwardsiella tarda EsaE (Orf19 protein) is required for the secretion of type III substrates, and pathogenesis in fish. Veterinary Microbiology. 190: 12–18. 10.1016/j.vetmic.2016.05.003

Last update:

No citation recorded.

Last update: 2025-07-28 18:25:44

No citation recorded.