Characterization of Spirometra erinaceieuropaei Plerocercoid Cysteine Protease and Potential Application for Serodiagnosis of Sparganosis

Abstract

BackgroundSparganosis is a neglected but important food-borne parasitic zoonosis. Clinical diagnosis of sparganosis is difficult because there are no specific manifestations. ELISA using plerocercoid crude or excretory–secretory (ES) antigens has high sensitivity but has cross-reactions with other helminthiases. The aim of this study was to characterize Spirometra erinaceieuropaei cysteine protease (SeCP) and to evaluate its potential application for serodiagnosis of sparganosis.Methodology/Principal FindingsThe full length SeCP gene was cloned, and recombinant SeCP (rSeCP) was expressed and purified. Western blotting showed that rSeCP was recognized by the serum of sparganum-infected mice, and anti-rSeCP serum recognized the native SeCP protein of plerocercoid crude or ES antigens. Expression of SeCP was observed at plerocercoid stages but not at the adult and egg stages. Immunolocalization identified SeCP in plerocercoid tegument and parenchymal tissue. The rSeCP had CP activity, and the optimum pH and temperature were 5.5 and 37°C, respectively. Enzymatic activity was significantly inhibited by E-64. rSeCP functions to degrade different proteins and the function was inhibited by anti-rSeCP serum and E-64. Immunization of mice with rSeCP induced Th2-predominant immune responses and anti-rSeCP antibodies had the potential capabilities to kill plerocercoids in an ADCC assay. The sensitivity of rSeCP-ELISA and ES antigen ELISA was 100% when performed on sera of patients with sparganosis. The specificity of rSeCP-ELISA and ES antigen ELISA was 98.22% (166/169) and 87.57% (148/169), respectively (P<0.05).ConclusionsThe rSeCP had the CP enzymatic activity and SeCP seems to be important for the survival of plerocercoids in host. The rSeCP is a potential diagnostic antigen for sparganosis.

Molecular Identification of Spirometra erinaceieuropaei Tapeworm in Cases of Human Sparganosis, Hong Kong

“…Diagnosis of sparganosis needs to be confirmed, normally by studying the excised lesions. Even though serologic tests for sparganosis have been described, these assays are not generally available and their performance requires more evaluation ( 9 – 13 ). In contrast, the presence of tunnel sign, conglomerated rings, bead-shaped enhancements, or images of parasites of various life stages by computerized tomography or magnetic resonance imaging are suggestive of sparganosis ( 14 ).…”

Chicken IgY-based coproantigen capture ELISA for diagnosis of human opisthorchiasis

“…These include the cysteine protease-like protein from the hookworm Ancylostoma caninum [29], plerocercoid cathepsin L–like protease of Spirometra erinaceieuropaei [30], and cathepsin B, an abundant enzyme of O. viverrini [17]. Coproantigen detection by capture ELISA has been popularly used for immunodiagnosis of many helminth infections because of its specificity [31].…”

Protective immunity in mice vaccinated with a novel elastase-1 significantly decreases Trichinella spiralis fecundity and infection

“…In this study, anti-rTsE antibody cytotoxicity on the NBL was also conducted as previously reported [39,61]. Forty NBLs were cocultivated for 48 h at 37 °C with 2 × 10 5 mouse peritoneal exudate cells (PECs) in 96-well plates in RPMI-1640 medium containing anti-rTsE serum, infection serum and preimmune serum.…”

“…Plates were developed with o-phenylenediamine dihydrochloride (OPD; Sigma-Aldrich) plus H 2 O 2 , and the reaction was terminated by using 2 M H 2 SO 4 . The absorbance at 492 nm was assayed using a microplate reader (Tecan, Schweiz, Switzerland) [38,39].…”