Immunodetection of Adhesin Pili Protein 38.6 kDa K. pneumoniae Using Western Blot
Abstract
Abstract: Klebsiella pneumoniae is a pathogen that causes infections in communities and hospitals. These bacteria have many virulence factors that play an important role in the pathogenicity of infection and antibiotic resistance. Pili protein Klebsiella pneumoniae with 38.6 kDa as one of the virulence factors, that have roles as hemagglutinin dan adhesin protein so potentially as a vaccine candidate. This research aimed to immunodetection pili protein 38.6 kDa using Western Blot. Western Blot was used to detect that protein with specific primary antibodies. These antibodies are obtained from mice serum-induced intraperitoneally with Pili protein 38.6 kDa Klebsiella pneumoniae. Protein bands that appear on the membrane of Western Blot results are proteins with a molecular weight of 85.6 kDa, 65.5 kDa, 46.9 kDa, and 29.4 kDa. This study concludes that Pili protein 38.6 kDa Klebsiella pneumoniae as the target protein does not appear in the Western Blot result.
Abstrak: Klebsiella pneumoniae adalah patogen yang menyebabkan infeksi di komunitas maupun rumah sakit. Bakteri tersebut memiliki banyak faktor virulensi yang berperan penting dalam patogenisitas terjadinya infeksi dan resistensi antibiotik. Protein pili Klebsiella pneumoniae dengan 38,6 kDa sebagai salah satu faktor virulensi, memiliki peran sebagai protein hemagglutinin dan adhesin sehingga berpotensi sebagai kandidat vaksin. Tujuan penelitian ini untuk imunodeteksi protein pili 38,6 kDa dengan menggunakan Western Blot. Western Blot digunakan untuk mendeteksi protein itu dengan spesifik antibodi primer. Antibodi ini diperoleh dari serum tikus yang telah diinduksi secara inteperitoneal dengan protein Pili 38,6 kDa Klebsiella pneumoniae. Band protein yang muncul pada membran hasil Western Blot adalah protein dengan berat molekul 85,6 kDa; 65,5 kDa; 46,9 kDa dan 29,4 kDa. Dari penelitian ini dapat disimpulkan bahwa protein Pili 38.6 kDa Klebsiella pneumoniae sebagai protein target tidak muncul.
Keywords
Full Text:
PDFReferences
Agustina, D., Nadyatara, K., Mufida, D. C., Elfiah, U., Shodikin, M. A., & Suswati, E. (2020). Faktor Virulensi Outer Membrane Protein 20 kDa Klebsiella pneumoniae sebagai Protein Hemaglutinin dan Adhesin. EJournal Kedokteran Indonesia, 7(3), 200–204. https://doi.org/10.23886/ejki.7.10425.
Agustina, D., Sumarno, & Noorhamdani. (2014). Inhibition of Klebsiella pneumoniae adhesion in mice enterocytes by antibodies of hemagglutinin pili protein with MW 12.8 kDa of Klebsiella pneumoniae | Agustina | Journal of Tropical Life Science. Journal of Tropical Life Science, 4(1), 19–25.
Anaya, J., Shoenfeld, Y., & Rojas-Villarraga, A. (2013). Autoimmunity - NCBI Bookshelf. Retrieved September 22, 2020, from PubMed website: https://www.ncbi.nlm.nih.gov/books/NBK459447/
Ashurst, J. V., & Dawson, A. (2018). Pneumonia, Klebsiella. In StatPearls. StatPearls Publishing.
Atmani, S. M., Messai, Y., Alouache, S., Fernández, R., Estepa, V., Torres, C., & Bakour, R. (2015). Virulence characteristics and genetic background of ESBL-producing Klebsiella pneumoniae isolates from wastewater. Fresenius Environmental Bulletin, 24(1), 103–112.
Baker, M. P., Reynolds, H. M., Lumicisi, B., & Bryson, C. J. (2010, October). Immunogenicity of protein therapeutics: The key causes, consequences and challenges. Self/Nonself - Immune Recognition and Signaling, Vol. 1, pp. 314–322. https://doi.org/10.4161/self.1.4.13904
Bio Rad. (2020). Western Blot Troubleshooting: Unusual or Unexpected Bands | Bio-Rad. Retrieved November 30, 2021, from Bio-Rad Laboratories, Inc. website: https://www.bio-rad-antibodies.com/western-blot-unusual-unexpected-bands-western-blotting.html?JSESSIONID_STERLING=87077389C9241CBCD37EB2D5931A0369.ecommerce2&evCntryLang=ID-en&cntry=ID&thirdPartyCookieEnabled=true
Blancher, C., & Jones, A. (2003). SDS-PAGE and Western Blotting Techniques. In Metastasis Research Protocols (Vol. 57, pp. 145–162). Humana Press. https://doi.org/10.1385/1-59259-136-1:145
Cienfuegos-Gallet, A. V., Ocampo De Los Ríos, A. M., Sierra Viana, P., Ramirez Brinez, F., Restrepo Castro, C., Roncancio Villamil, G., … Jiménez, J. N. (2019). Risk factors and survival of patients infected with carbapenem-resistant Klebsiella pneumoniae in a KPC endemic setting: A case-control and cohort study. BMC Infectious Diseases, 19(1), 830. https://doi.org/10.1186/s12879-019-4461-x
Drenckhahn, D., Jöns, T., & Schmitz, F. (1993). Production of Polyclonal Antibodies against Proteins and Peptides. Methods in Cell Biology, 37(C), 7–56. https://doi.org/10.1016/S0091-679X(08)60242-3
Eichenberger, E. M., & Thaden, J. T. (2019, June 1). Epidemiology and mechanisms of resistance of extensively drug-resistant gram-negative bacteria. Antibiotics, Vol. 8, p. 37. MDPI AG. https://doi.org/10.3390/antibiotics8020037
Ferrer, M., & Torres, A. (2018). Epidemiology of ICU-acquired pneumonia. Current Opinion in Critical Care, 24(5), 325–331. https://doi.org/10.1097/MCC.0000000000000536
Finka, R., Agustina, D., Rachmawati, D. A., Suswati, E., Mufida, D. C., & Shodikin, A. (2019). The Role of Pili Protein 38,6 kDa Klebsiella pneumoniae as a Hemagglutinin and Adhesin Protein which Serves as a Virulence Factor. Journal of Agromedicine and Medical Sciences, 5(2), 9. https://doi.org/10.19184/ams.v5i2.9558
Gallagher, S., Winston, S. E., Fuller, S. A., & Hurrell, J. G. R. (2008). Immunoblotting and immunodetection. Current Protocols in Molecular Biology, 83(SUPPL. 83), 10.8.1-10.8.28. https://doi.org/10.1002/0471142727.mb1008s83
Gilda, J. E., Ghosh, R., Cheah, J. X., West, T. M., Bodine, S. C., & Gomes, A. V. (2015). Western blotting inaccuracies with unverified antibodies: Need for a Western Blotting Minimal Reporting Standard (WBMRS). PLoS ONE, 10(8). https://doi.org/10.1371/journal.pone.0135392
Hoppe, S., Bier, F. F., & von Nickisch-Rosenegk, M. (2012). Microarray-based method for screening of immunogenic proteins from bacteria. Journal of Nanobiotechnology, 10(1), 12. https://doi.org/10.1186/1477-3155-10-12
Khairuzzaman, M. Q. (2016). Hemaglutinin. 4(1), 64–75.
Khater, F., Balestrino, D., Charbonnel, N., Dufayard, J. F., Brisse, S., & Forestier, C. (2015). In Silico Analysis of Usher Encoding Genes in Klebsiella pneumoniae and Characterization of Their Role in Adhesion and Colonization. PLOS ONE, 10(3), e0116215. https://doi.org/10.1371/journal.pone.0116215
Kishimbo, P., Sogone, N. M., Kalokola, F., & Mshana, S. E. (2020). Prevalence of gram-negative bacteria causing community-acquired pneumonia among adults in Mwanza City, Tanzania. Pneumonia, 12(1), 7. https://doi.org/10.1186/s41479-020-00069-0
Madamanchi, N. R., & Runge, M. S. (2001). Western blotting. Methods in Molecular Medicine, 51, 245–256. https://doi.org/10.1385/1-59259-087-X:245
Mahmood, T., & Yang, P. C. (2012). Western blot: Technique, theory, and troubleshooting. North American Journal of Medical Sciences, 4(9), 429–434. https://doi.org/10.4103/1947-2714.100998
Paczosa, M. K., & Mecsas, J. (2016). Klebsiella pneumoniae: Going on the Offense with a Strong Defense. Microbiology and Molecular Biology Reviews, 80(3), 629–661. https://doi.org/10.1128/mmbr.00078-15
Thanassi, D. G., Bliska, J. B., & Christie, P. J. (2012, November 1). Surface organelles assembled by secretion systems of Gram-negative bacteria: Diversity in structure and function. FEMS Microbiology Reviews, Vol. 36, pp. 1046–1082. Oxford Academic. https://doi.org/10.1111/j.1574-6976.2012.00342.x
Torres, A., Niederman, M. S., Chastre, J., Ewig, S., Fernandez-Vandellos, P., Hanberger, H., … Wunderink, R. (2017, September 1). International ERS/ESICM/ESCMID/ALAT guidelines for the management of hospital-acquired pneumonia and ventilator-associated pneumonia. European Respiratory Journal, Vol. 50, p. 1700582. European Respiratory Society. https://doi.org/10.1183/13993003.00582-2017
Tsereteli, M., Sidamonidze, K., Tsereteli, D., Malania, L., & Vashakidze, E. (2018). Epidemiology of Carbapenem-Resistant Klebsiella Pneumoniae in Intensive Care Units of Multiprofile Hospitals in Tbilisi, Georgia. Georgian Medical News, (280–281), 164–168.
Vallecoccia, M. S., Dominedò, C., Cutuli, S. L., Martin-Loeches, I., Torres, A., & De Pascale, G. (2020, September 30). Is ventilated hospital-acquired pneumonia a worse entity than ventilator-associated pneumonia? European Respiratory Review, Vol. 29, pp. 1–8. European Respiratory Society. https://doi.org/10.1183/16000617.0023-2020
Vuotto, C., Longo, F., Balice, M., Donelli, G., & Varaldo, P. (2014). Antibiotic Resistance Related to Biofilm Formation in Klebsiella pneumoniae. Pathogens, 3(3), 743–758. https://doi.org/10.3390/pathogens3030743
WHO. (2020). WHO reveals leading causes of death and disability worldwide: 2000-2019. Retrieved December 12, 2020, from https://www.who.int/news/item/09-12-2020-who-reveals-leading-causes-of-death-and-disability-worldwide-2000-2019
Wright, K. (1989). Antibodies a Laboratory Manual. Biochemical Education, 17(4), 220. https://doi.org/10.1016/0307-4412(89)90165-9
Zhu, W. M., Yuan, Z., & Zhou, H. Y. (2020). Risk factors for carbapenem-resistant Klebsiella pneumoniae infection relative to two types of control patients: A systematic review and meta-analysis. Antimicrobial Resistance and Infection Control, 9(1). https://doi.org/10.1186/s13756-020-0686-0
DOI: http://dx.doi.org/10.22373/ekw.v7i2.10127
Refbacks
- There are currently no refbacks.
Copyright (c) 2021 Dini Agustina, Diana Chusna Mufida, Nurul Indah Saffanah
This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.
P-ISSN : 2460-8912
E-ISSN : 2460-8920
ELKAWNIE
This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.
Elkawnie: Journal of Islamic Science and Technology in 2022. Published by Faculty of Science and Technology in cooperation with Center for Research and Community Service (LP2M), UIN Ar-Raniry Banda Aceh, Aceh, Indonesia.
View full page view stats report click here