Evaluation of Toxoplasma gondii propagated in specific pathogen free embryonated chicken egg, for diagnosis of toxoplasmosis in equids and human

  • Abeer M. Abdalhamed
  • Mohey A. Hassanain
  • Gamil S. G. ZeedanEmail author
  • Raafat M. Shaapan
Original Article


Toxoplasma gondii (T. gondii) is a worldwide distribution infects a wide variety of mammals, including humans. The present study aimed to detect the efficacy of soluble and whole T. gondii antigens propagated in specific pathogen-free of embryonated chicken egg (SPF-ECE) used to improve the potency of serological assays for diagnosis of toxoplasmosis in equids and human. Total of 220 serum samples from 170 equids (90 donkeys and 55 horses and 25 mules) and 50 humans were collected from different governorates in Egypt during the period from October 2017 to March 2018. Crude T. gondii tachyzoites antigens from low or high passages propagated in mice or SPF-ECE was used for modifying some serological tests. The experiment showed that the mortality rate of T. gondii for 103 and 104 low passages were 6/8 (75%) and 7/8 (88%) dead embryos but, lower mortality rate in high passage T. gondii were 4/8 (50%) and 5/8 (63%) dead embryos, respectively. No mortality or inflammatory signs were observed in control of negative groups. In equids sera were examined by S-ELISA using soluble T. gondii antigen propagated in SPF-ECE showed the highest positive results 26 (28.8%), followed by LAT 37 (22%) and MAGPT 36 (21.17%). While, W-ELISA and IFAT used whole T. gondii antigen prepared in SPF-ECE were 35 (20.58%) and 28 (19.41%) showed highly positive results than the same test used the whole antigen prepared in mice. The highest seroprevalence of T. gondii in human and donkeys were 19/50 (38%). and 26/90 (28.88%), more than mules were 6/25 (24%) and horses were 9/55 (16.3%) examined by S-ELISA respectively. SPF-ECE is considered an appropriate experimental model for isolation and propagation of T. gondii tachyzoites, and their soluble antigens used in serological tests (S-ELISA, LAT, and MAGPT) have sensitivity and specificity more than the whole antigen and provided reliable diagnostic tools for detection of toxoplasmosis in human and equids.




Authors contribution

Dr. GSGZ: Research idea, planned the study design, performed data, and, field animals samples collection, serological and propagation of toxoplasma in SPF-ECE, laboratory work, and drafting the paper. Dr. AMA: sharing in the conception of the research idea, field animal’s samples collection, sharing designed work, identification of T. gondii, sharing serological test and and participated in drafting the manuscript. Dr. MAH and Dr. RMS: involved in samples collection, laboratory work, interpreted the data results, and helped in manuscript preparation and designed work and participated in drafting the manuscript. All authors read and approved the final manuscript.


This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

The study was approved ethically by Medical Research Ethical Committee Research, National Statement on Ethical Conduct in Human and animals Research at National Research Centre, Egypt under registration number # 19-030#.


  1. Alanazi AD, Alyousif MS (2011) Prevalence of antibodies to Toxoplasma gondii in horses in Riyadh Province, Saudi Arabia. J Parasitol 97(5):943–945CrossRefGoogle Scholar
  2. Al-Kappany YM, Abbas IE, Devleess chauwer B, Dorny P, Jennes M, Cox E (2018) Seroprevalence of anti-Toxoplasma gondii antibodies in Egyptian sheep and goats. BMC Vet Res 14(1):120. CrossRefGoogle Scholar
  3. Boyle JP, Radke JR (2009) A history of studies that examine the interactions of Toxoplasma with its host cell: emphasis on in vitro models. Int J Parasitol 39(8):903–914CrossRefGoogle Scholar
  4. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72(1–2):248–254CrossRefGoogle Scholar
  5. Brandon-Mong GJ, Che Mat Seri NA, Sharma RS et al (2015) Seroepidemiology of toxoplasmosis among people having close contact with animals. Front Immunol 6:143CrossRefGoogle Scholar
  6. Chen S, Lou D, Lin A, Zeng X, Ding Z, Wen L, Ohta N, Wang J, Fu C (2008) Evaluation of a rapid ELISA technique for detection of circulating antigens of Toxoplasma gondii. Microbiol Immunol 52(3):180–187. CrossRefGoogle Scholar
  7. Dubey JP (2010) Toxoplasmosis of animal and humans, vol 2. CRC Press, Boca Raton, p 313. Google Scholar
  8. Ghazy AA, Shaapan RM, Abdel-Rahman EH (2007) Comparative serological diagnosis of toxoplasmosis in horses using locally isolated Toxoplasma gondii. Vet Parasitol 145(1–2):31–36. CrossRefGoogle Scholar
  9. Ghoneim NH, Shalaby SI, Hassanain NA, Zeedan GS, Soliman YA, Abdalhamed AM (2010) Comparative study between serological and molecular methods for diagnosis of toxoplasmosis in women and small ruminants in Egypt. Foodborne Pathog Dis 7:17–22. CrossRefGoogle Scholar
  10. Holliman RE, Johnson JU, Duffy K, New L (1989) Discrepant toxoplasma latex agglutination test results. J Clin Pathol 42(2):200–203CrossRefGoogle Scholar
  11. Khodakaram-Tafti A, Mansourian M, Namavari M, Hosseini A (2012) Immunohistochemical and polymerase chain reaction studies in Neospora caninum experimentally infected broiler chicken embryonated eggs. Vet Parasitol 188(1–2):10–13. CrossRefGoogle Scholar
  12. Klun I, Uzelac A, Villena I, MercierA BB, Nikolić A et al (2017) The first isolation and molecular characterization of Toxoplasma gondii from horses in Serbia. Parasit Vectors 10(167):2–9Google Scholar
  13. MacFarlane JO, Ruchman I (1948) Cultivation of Toxoplasma in the developing chick embryo. Exp Biol Med 67:1–4CrossRefGoogle Scholar
  14. Mansourian M, Namavari M, Khodakaram-Tafti A, Rahimian A (2013) Experimental Neospora caninum infection in domestic bird’s embryonated eggs. J Parasit Dis 39(2):241–244. CrossRefGoogle Scholar
  15. Mello MN, Deane MP (1976) Patterns of development of Trypanosoma cruzi in the embryonated chicken egg. Ann Trop Med Parasitol 70(4):380–388CrossRefGoogle Scholar
  16. Miao Q, Wang X, She LN et al (2013) Seroprevalence of Toxoplasma gondii in horses and donkeys in Yunnan Province, Southwestern China. Parasit Vectors 6:168CrossRefGoogle Scholar
  17. Namavari M, Mansourian M, Tafti AK, Hosseini MH, Rahimiyan A, Khordadmehr M, Lotfi M (2012) Application of chicken embryonated eggs as a new model for evaluating the virulence of Neospora caninum tachyzoites. Comp Clin Path 21(6):1665–1668CrossRefGoogle Scholar
  18. Papini RA, Buzzone G, Nardoni S, Rocchigiani G, Mancianti F (2015) Seroprevalence and genotyping of Toxoplasma gondii in horses slaughtered for human consumption in Italy. J Equine Vet Sci 35:657–661. CrossRefGoogle Scholar
  19. Piergili Fioretti D (2004) Problems and limitations of conventional and innovative methods for the diagnosis of Toxoplasmosis in humans and animals. Parassitologia 46:177–181Google Scholar
  20. Pishkari S, Shojaee S, Keshavarz H, Salimi M, Mohebali M (2017) Evaluation of Toxoplasma gondii soluble, whole and excretory/secretary antigens for diagnosis of toxoplasmosis by ELISA test. J Parasit Dis 41(1):289–291CrossRefGoogle Scholar
  21. Que X, Wunderlich A, Joiner KA, Reed SL (2004) Toxopain-1 is critical for infection in a novel chicken embryo model of congenital toxoplasmosis. Infect Immun 72(5):2915–2921CrossRefGoogle Scholar
  22. Rahbari AH, Keshavarz H, Shojaee S, Mohebali M, Rezaeian M (2012) IgG avidity ELISA test for diagnosis of acute toxoplasmosis in humans. Korean J Parasitol 50:99–102CrossRefGoogle Scholar
  23. Razmi GR, Abedi V, Yaghfoori S (2014) Serological study of Toxoplasma gondii infection in Turkoman horses in the North Khorasan Province, Iran. J Parasit Dis 40(2):515–519CrossRefGoogle Scholar
  24. Reid AJ, Vermont SJ, Cotton JA, Harris D, Hill-Cawthorne GA, Könen-Waisman S et al (2012) Comparative genomics of the apicomplexan parasites Toxoplasma gondii and Neospora caninum: coccidia differing in host range and transmission strategy. PLoS Pathog 8(3):e1002567CrossRefGoogle Scholar
  25. Saleh AM, Ali HA, Ahmed SA, Hosny SM, Morsy TA (2014) Screening of Toxoplasma gondii infection among childbearing age females and assessment of nurses’ role in prevention and control of toxoplasmosis. J Egypt Soc Parasitol 240(1416):1–4Google Scholar
  26. Setasimy A, Namavari M (2015) Use of chicken embryonated eggs for evaluating the virulence of Toxoplasma gondii. J Parasit Dis 40(4):1223–1225CrossRefGoogle Scholar
  27. Shaapan RM (2016) The common zoonotic protozoal diseases causing abortion. J Parasit Dis 40(4):1116–1129CrossRefGoogle Scholar
  28. Shappan RM, Abo-Eimaty AM, Abd-Eirazik KA, Abd-Eihafez SM (2012) PCR and serological assays for detection of Toxoplasma gondii infection in sport horses in Cairo, Egypt. Asian J Anim Vet Adv 7:158–165. CrossRefGoogle Scholar
  29. Tenter AM, Heckeroth AR, Weiss LM (2000) Toxoplasma gondii: from animals to humans. Int J Parasitol 30(12–13):1217–1258CrossRefGoogle Scholar
  30. Torrey EF, Yolken RH (2013) Toxoplasma oocytes’ as a public health problem. Trends Parasitol 29(8):380–384CrossRefGoogle Scholar
  31. Walls KW, Barnhart ER (1978) Titration of human serum antibodies to Toxoplasma gondii with a simple fluorometric assay. J Clin Microbiol 7(2):234–235Google Scholar
  32. Warren J, Russ SB (1948) Cultivation of Toxoplasma in embryonated egg An antigen derived from choriolallantoic membrane. Exp Biol Med 67:85–89CrossRefGoogle Scholar
  33. Wilson M, Ware DA, Juranek DD (1990) Serologic aspect of toxoplasmosis. J Am Vet Med Assoc 196:277–280Google Scholar

Copyright information

© Indian Society for Parasitology 2019

Authors and Affiliations

  1. 1.Department of Parasitology and Animal Diseases, Veterinary Research DivisionNational Research CentreGizaEgypt
  2. 2.Zoonotic Diseases Department, Veterinary Research DivisionNational Research CentreGizaEgypt

Personalised recommendations