In Vitro Maintenance of Clonorchis sinensis Adult Worms

Clonorchis sinensis is a biological carcinogen inducing human cholangiocarcinoma, and clonorchiasis is one of the important endemic infectious diseases in East Asia. The present study investigated survival longevity of C. sinensis adult worms in various in vitro conditions to find the best way of keeping the worms longer. The worms were maintained in 0.85% NaCl, 1xPBS, 1xLocke's solution, RPMI-1640, DMEM, and IMDM media, and in 1xLocke's solution with different supplements. All of the worms died within 3 and 7 days in 0.85% NaCl and 1xPBS, respectively, but survived up to 57 days in 1xLocke's solution. The worms lived for 106 days in DMEM, and 114 days in both RPMI-1640 and IMDM media. The survival rate in RPMI-1640 medium was the highest (50%) compared to that in DMEM (20+/-10%) and in IMDM (33.3+/-25.2%) after 3 months. The 1xLocke's solution with 0.005% bovine bile supplement showed increased duration of maximum survival from 42 days to 70 days. Higher concentration of bile supplements than 0.005% or addition of glucose were disadvantageous for the worm survival. The worms died rapidly in solutions containing L-aspartic acid, L-glutamic acid, and adenine compared to L-arginine, L-serine, and L-tryptophan. In conclusion, the 1xLocke's solution best supports the worms alive among inorganic solutions for 57 days, and the RPMI-1640 medium maintains living C. sinensis adults better and longer up to 114 days in vitro than other media.

Functional Genes and Proteins of Clonorchis sinensis

During the past several decades, researches on parasite genetics have progressed from biochemical and serodiagnostic studies to protein chemistry, molecular biology, and functional gene studies. Nowadays, bioinformatics, genomics, and proteomics approaches are being applied by Korean parasitology researchers. As for Clonorchis sinensis, investigations have been carried out to identify its functional genes using forward and reverse genetic approaches and to characterize the biochemical and biological properties of its gene products. The authors review the proteins of cloned genes, which include antigenic proteins, physiologic and metabolic enzymes, and the gene expression profile of Clonorchis sinensis.

Resistance to reinfection in rats induced by irradiated metacercariae of Clonorchis sinensis

A study was made to observe the association between the resistance to reinfection induced by irradiated metacercariae (MC) of Clonorchis sinensis and antigen specific Th1- and Th2-type cytokine productions in rats. Rats were infected with 20 MC of C. sinensis, previously exposed to a single dose of gamma irradiation, which varied from 0 to 100 Gy. All of them, single dose of 12 Gy showed higher IgG antibody titer with lowest worm recovery. Thus, 50 MC were used to challenge infection in rats previously infected with 20 MC irradiated at 12 Gy and the highest resistance to challenge infection was observed. The results of lymphocyte proliferation with specific antigen, ES Ag were shown no difference of proliferative responses as compared with primary and challenge infection at 12 Gy irradiation dose. In the case of cytokines production were observed that interferon (IFN-gamma) and interlukin (IL-2) were significantly enhanced, while IL-4 and IL-10 was almost unchanged to make comparison between primary and secondary infection at 12 Gy irradiation dose. In conclusion, the single dose of 12 Gy could be adopted for induction of the highest resistance to challenge infection. Up-regulation of Th1 type cytokines, IFN-gamma and IL-2 may be affected to develop vaccine by irradiated MC.

Bile duct changes in rats reinfected with Clonorchis sinensis

This study describes an evaluation of the sonographic, cholangiographic, pathological, and immunological findings, and the protective effect shown by rats reinfected with Clonorchis sinensis. Eight experimental rat groups were, namely, a normal control, a primary infection control, a reinfection I (reinfection 7 week after treatment following 3-week infection), a reinfection II (reinfection 2 week after treatment following 8-week infection), a reinfection III (exploration of the intrahepatic bile ducts 1 week after reinfection 4 week after treatment following 4-week infection), a superinfection, a secondary infection control, and an infection following immunization group. Sonographic and cholangiographic findings showed moderate or marked dilatation of the bile duct confluence in the primary infection control, reinfection II, and secondary infection control groups. Juvenile worms survived in the intrahepatic bile ducts 1 week after reinfection following treatment in the reinfection III group. It was concluded that reinfecting juvenile worms found during the first week following reinfection failed to survive or grow further. Anatomical, pathophysiological, or immunological changes may induce protection from reinfection in rats.

Clonorchis sinensis metacercarial infection in the pond smelt Hypomesus olidus and the minnow Zacco platypus collected from the Soyang and Daechung Lakes

The pond smelt Hypomesus olidus and minnow Zacco platypus were collected from the Soyang and Daechung Lakes in January 2003, and their metacercarial infections was examined by the muscle compression and artificial digestion techniques. In the Soyang Lake, 161 metacercariae of Clonorchis sinensis (0.35 per fish) were harvested from 459 pond smelts examined. Also, 13 metacercariae of C. sinensis (0.43 per fish), 1 of Metagonimus sp., 4 of Echinostoma sp., 148 of Centrocestus armatus and 44 unidentified species were collected from 30 minnows. In the Daechung Lake, 369 metacercariae of C. sinensis (3.69 per fish) and 51 unidentified species were recovered from 100 pond smelts. The metacercariae of C. sinensis were fed to experimental rats, in which the adult flukes were identified. The pond smelts and minnows collected from the Soyang and Daechung Lakes were verified to be the second intermediate hosts and the sources of human C. sinensis infection.