Isolation and purification of Echinococcus granulosus antigen B from hydatid cyst fluid using three different methods

Document Type : Original Articles

Authors

1 Department of Pathobiology, Faculty of Veterinary, Science and Research Branch, Islamic Azad University, Tehran, Iran

2 Proteomics and Biochemistry Department, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.

3 Department of Microbiology, School of Specialized Veterinary Science, Science and Research Branch, Islamic Azad University, Tehran, Iran.

4 Department of Pathobiology, Faculty of Veterinary, Islamic Azad University, Garmsar, Iran

Abstract

Hydatid cyst, the larval stage of cestodes Echinococcus spp., is recognized as a zoonotic infection in the world. The World Health Organization (WHO) has recently classified echinococcosis in a group of neglected tropical diseases.The prevalence of Echinococcus granulosus infection is high in Iran due to the presence of various intermediate hosts in this country. Considering the rising trend of this zoonotic parasitic disease based on national epidemiological studies, diagnosis is of great significance. WHO has suggested the use of specific antigens, especially antigen B (AgB) for serological diagnostic tests. In general, AgB is a polymeric lipoprotein, which disintegrates into 8.12, 16, and 20.24 kDa subunits. In the present study, we applied three different methods for AgB isolation from hydatid cyst fluid (HCF) and compared their efficacy in AgB isolation. Finally, the protein concentration of this antigen was measured by Bradford assay and confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The results showed that the application of polyethylene glycol (PEG 4000) as a thickener agent beside purification of HCF in dialysis bag and filtering and also dialysis against acetate buffer leading to the best quantity in purified antigen B.

Keywords

Main Subjects


Article Title [French]

Isolation et purification de l’antigène B de l’Echinococcus granulosus à partir du fluide d’un kyste hydatique par trois différentes méthodes

Abstract [French]

Le kyste hydatique est une zoonose provoquée par la larve des cestodes du genre Echinococcus dans de nombreuses régions du monde. L’Organisation Mondiale de la Santé (OMS) a récemment classifié l’échinicoccose dans le groupe des maladies tropicales jusqu’alors négligées. En Iran, la prévalence des infections à l’Echinococcus granulosus est importante en raison de l’existence d’hôtes intermédiaires variées à travers le pays. Etant donnée l’augmentation de cette zoonose parasitique soulignée par les études épidémiologiques nationales, son diagnostique est d’une grande importance. L’OMS propose l’utilisation d’antigènes spécifiques, particulièrement l’antigène B (AgB), dans l’élaboration de tests sérologiques. L’AgB est une lipoprotéine polymérique constituée de plusieurs sous-unités de 8.12, 16 et 20.24 kDa. Dans cette étude, les rendements de trois différentes méthodes dans l’isolation de l’AgB à partir du fluide du Kyste hydatique (HCF) ont été comparés. Pour cela, la concentration de l’antigène a été mesurée par la méthode de Bradford et son degré de pureté vérifié par une électrophorèse en gel de polyacrylamide contenant du laurylsulfate de sodium (SDS-PAGE). Nos résultats montrent que l’application du polyéthylène glycol (PEG 4000) comme agent épaississant, complémentée d’une filtration et d’un dialyse contre du tampon acétate (pH 8), contribue à un meilleur rendement de purification de l’HCF.

Keywords [French]

  • Antigène B
  • Echinococcus granulosus
  • Kyste hydatique
  • Isolation
Asghari, M., Mohebali, M., Kia, E.B., Farahnak, A., Aryaeipour, M., Asadian, S., Rokni, M.B., 2013. Seroepidemiology of Human Hydatidosis Using AgB-ELISA Test in Arak, Central Iran. Iranian journal of public health 42, 391-396.
Barnes, T.S., Deplazes, P., Gottstein, B., Jenkins, D.J., Mathis, A., Siles-Lucas, M., Torgerson, P.R., Ziadinov, I., Heath, D.D., 2012. Challenges for diagnosis and control of cystic hydatid disease. Acta tropica 123, 1-7.
Carmena, D., Benito, A., Eraso, E., 2006. Antigens for the immunodiagnosis of Echinococcus granulosus infection: An update. Acta tropica 98, 74-86.
Eckert, J., Gemmell, M., Meslin, F., Powloeski, Z., 2002. WHO/OIE Manual of Echinococcosis in Human and Animal. World Organization for Animal Health, 20-40.
Gillespie, S.H., Hawkey, P.M., 1995. Medical Parasitology: A Practical Approach, IRL Press at Oxford University Press.
Gonzalez-Sapienza, G., Cachau, R.E., 2003. Identification of critical residues of an immunodominant region of Echinococcus granulosus antigen B. The Journal of biological chemistry 278, 20179-20184.
Hajipirloo, H.M., Bozorgomid, A., Alinia, T., Tappeh Kh, H., Mahmodlou, R., 2013. Human cystic echinococcosis in west azerbaijan, northwest iran: a retrospective hospital based survey from 2000 to 2009. Iranian journal of parasitology 8, 323-326.
Mableson, H.E., Okello, A., Picozzi, K., Welburn, S.C., 2014. Neglected zoonotic diseases-the long and winding road to advocacy. PLoS neglected tropical diseases 8, e2800.
Mamuti, W., Sako, Y., Nakao, M., Xiao, N., Nakaya, K., Ishikawa, Y., Yamasaki, H., Lightowlers, M.W., Ito, A., 2006. Recent advances in characterization of Echinococcus antigen B. Parasitology international 55 Suppl, S57-62.
Mohammadzadeh, T., Sako, Y., Sadjjadi, S.M., Sarkari, B., Ito, A., 2012. Comparison of the usefulness of hydatid cyst fluid, native antigen B and recombinant antigen B8/1 for serological diagnosis of cystic echinococcosis. Transactions of the Royal Society of Tropical Medicine and Hygiene 106, 371-375.
Moro, P., Schantz, P.M., 2009. Echinococcosis: a review. International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases 13, 125-133.
Rafiei, A., Jahanshahi, A., Talazadeh, A., 2008. Evaluation of Specific IgG Antibody Detection in Diagnosis and Post Surgical Monitoring of Hydatic Echinococcosis. Iranian Journal Parasitology 3, 10-14.
Rahimi, H., Sadjjadi, S., Sarkari, B., 2011. Performance of antigen B isolated from different hosts and cyst locations in diagnosis of cystic echinococcosis. Iranian journal of parasitology 6, 12-19.
Rigano, R., Profumo, E., Bruschi, F., Carulli, G., Azzara, A., Ioppolo, S., Buttari, B., Ortona, E., Margutti, P., Teggi, A., Siracusano, A., 2001. Modulation of human immune response by Echinococcus granulosus antigen B and its possible role in evading host defenses. Infection and immunity 69, 288-296.
Rokni, M., 2009. Echinococcosis /hydatidosis in Iran. Iranian journal of parasitology 4, 1-16.
Sadjjadi, SM., Abidi, H., Sarkari, B., Izadpanah, A., Kazemian, S., 2007. Evaluation 0f Enzyme Linked Immunosorbant Assay, Utilizing Native Antigen B for Serodiagnosis of Human Hydatidosis. Iranian Journal of Immunology 4, 167-172.
Sarkari, B., Sadjjadi, SM., Abidi, H., Izadpanah, A., Kazemian, S., Rafiei, A., 2007. Application of Western Blotting Using Native Antigen B for Serodiagnosis of Human Cystic Echinococcosis. Iranian Journal of Parasitology 2, 7-12.
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Silva, A., 2011. Hydatid cyst/cystic echinococcosis: anatomical and surgical nomenclature and method to quantify the cyst content solidification. Chinese Medical Journal 124, 2806-2812.
Zhang, W., Li, J., Jones, M.K., Zhang, Z., Zhao, L., Blair, D., McManus, D.P., 2010. The Echinococcus granulosus antigen B gene family comprises at least 10 unique genes in five subclasses which are differentially expressed. PLoS neglected tropical diseases 4, e784.