EGPE cells are good antigen source for hydatidosis diagnostic in sheep

MAGLIOCO A; PERALTA ME; JENSEN O; MARÍA LAURA GERTISER; MELISA S BARBERY VENTURI; FRIDMAN O; GABRIELA A CANZIANI; FUCHS A

Congreso; Reunión Conjunta de Sociedades de Biociencias; 2017

Echinococcus granulosus is the causative agent of hydatidosisdisease, a zoonosis with worldwide distribution. Nowadays, diagnosisof hydatidosis in livestock happens mainly in necropsies.However, an accurate serological diagnostic test would be desirableto monitor livestock in order to prevent the dissemination of thedisease in non-endemic zones. A cellular model of E. granulosuswas established from bovine protoscoleces in our laboratory and itwas named EGPE. In these cells AgB4 was detected by immunohistochemistryand DCO1 (Echeverria et al, 2010) and antigen B2subunit was detected by PCR. Purpose: to identify whether EGPEcell proteins could be recognized by infected sheep serum. Materialsand Methods: EGPE cells were cultured in complete mediumM199 for short (7 days) or long term culture (20 days), for proteinextraction (CE). The CE were obtained in 8 mM CHAPS buffer for 2hours and 4 freeze-thaw cycles. Serum from 7 infected sheep and5 non-infected sheep were obtained from Chubut, Argentina andreactivity to EGPE proteins were analyzed by western blot and ELISA.An 18 µg of CE were used per lane for western blot and 1.5µg of CE were used per well for ELISA. A 1/125 dilution for serumsample, alkaline phosphatase conjugated rabbit anti-sheep (1:5000)and BCIP/NBT (western blot) or p-nitrophenyl phosphate in diethanolaminebuffer (ELISA) were used. Results: After 15% SDS-PAGEelectrophoresis and immunoblotting, several bands from CE wererecognized exclusively by infected sheep: 56-64 kDa and 75-83 kDain short term culture and 44-48 kDa, 56-64 kDa and 70-74 kDa inlong term culture. After ELISA, differences between control and infectedsheep were detected in short (p