ABSTRACT
Echinococcus granulosus cultivation is very important for improvement of different aspect of medical and veterinary researches. Despite many advances in this case, there is a missing link for in vitro life cycle of adult worms and it is fertilization. Regarding the researchers' observations, self-fertilization can be done in worms living in dog intestine, but despite all sorts of experimental techniques, this phenomenon has never been observed in reared worms in culture media. Furthermore cross fertilization has not been observed in vitro and even in parasites with dog intestinal origin; although it theoretically is possible. During a follow-up of cultivated adult worms, evidences of behaviors similar to self-mating [Type 2] and cross-mating were observed in our lab which will be presented here. Protoscoleces were aseptically removed from sheep hydatid cysts, washed twice with PBS and then cultivated in S.10E.H culture medium. The stages of parasite growth were observed using an inverted microscope for two months and all stages and behaviors were microscopically photographed. Different movies have also been made from these behavioral features. After around 55 days post cultivation, some evidences of behaviors similar to self-mating [Type 2] and cross-mating were observed in some of the mature adult worms. However, fertile eggs in these parasites have never been observed. Regarding the above observations, these parasites show tendency to unsuccessful self-mating/fertilization [type 2] which failure could be due to anatomical position and physiological maturation. Also lack of suitable conditions for self-fertilization causes the worms try to do unsuccessful cross- mating/fertilization in culture media
ABSTRACT
Cystic echinococcosis [CE], also known as echinococcosis/hydatidosis, is one of the most important parasitic diseases in the world. It enhances both humoral and cellular [Thl and Th2] responses in infected host. Different antigens of the worm may favor the Thl or Th2 immune responses in CE patients. To evaluate the humoral and cellular immune responses of Balb/c mice against the crude and excretory/secretory [E/S] antigens of in vitro reared Echinococcus granulosus adult worms. A total of 20 Balb/c mice divided into 5 groups of 4 mice each. Three groups of mice [n=4] were immunized with crude, E/S and an immunodominant antigen of in vitro reared Echinococcus granulosus adult worms on day 1 and 28. The fourth and the fifth groups were negative control groups and received PBS plus adjuvant, or nothing, respectively. Two weeks after the second injection, the mice were killed and their blood was collected for determining antibody responses, and their spleens were employed for proliferation assay. Total IgG were measured by indirect ELISA. Spleen cells of immunized mice were cultivated and exposed to different antigens of adult worms including E/S and crude antigens. Level of IFN-y, IL-12, IL-4 and IL-10 were measured in the recovered cell culture supernatants by capture ELISA. Total IgG assay showed the highest level of antibody produced in mice immunized with crude antigens. Proliferation assay showed a statistically significant production of cytokines in the mice immunized with crude antigens [p<0.05]. The highest levels of IFN-gamma, IL12 and IL-4 were produced in mice immunized with crude antigen of the in vitro reared Echinococcus granulosus adult worms followed by E/S antigens. Immunodomonant antigen induced the lowest levels of cytokines [IL-12, IFN-gamma, IL-4 and IL-10] in immunized mice. A significant levels of Thl related cytokines [IFN-gamma and IL-12] were produced in Balb/c mice immunized with crude antigen of the in vitro reared Echinococcus granulosus adult worms