Gene expression of Clonorchis sinensis metacercaria induced by gamma irradiation.

Gamma-rays are a form of ionizing radiation and produce serious cellular damage to nuclei and organelles. Gamma irradiation induces the expressions of genes involved in DNA repair. Clonorchis sinensis resides in and provokes pathophysiologic changes in the bile ducts of mammals. The C. sinensis metacercariae are unsusceptible or resistant to gamma irradiation with LD50 of 16.5 Gy. Using the annealing control primer-based polymerase chain reaction (PCR) method, 19 genes were found to be up-regulated in C. sinensis metacercariae exposed to gamma rays. Contigs of up-regulated genes (URGs) were retrieved in a C. sinensis expressed sequence tag pool and extended by DNA-walking. Of the 13 URGs annotated putatively as functional genes, five URGs were associated with energy metabolism, six with protein processing, and the other two with DNA repair protein RAD23 and inhibitor of apoptosis protein. Four URGs were confirmed up-regulated by gamma irradiation by quantitative real-time PCR. One unknown gene, which was up-regulated to the greatest extent, might contribute to early recovery from gamma-irradiation-induced damage. The up-regulations of genes encoding DNA repair, protein processing, and energy metabolism proteins suggests that increases in gene products orchestrate DNA lesion repair and recover cellular functions in gamma-irradiated C. sinensis metacercariae.

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.

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.

Effects of gamma-irradiation on the infectivity and chromosome aberration of Clonorchis sinensis.

Effects of gamma irradiation on the worm survival and chromosomal aberration of Clonorchis sinensis were studied. The metacercariae irradiated with various amounts of gamma radiation (ranging from 5 Gy to 50 Gy) were fed to rats, and the effects were compared with those of non-irradiated controls. Recovery rates of adult worms in irradiated groups were reduced gradually as increasing of the irradiation doses. No worm was recovered from rats which were fed with 50 Gy irradiated metacercariae. The chromosome number was 2n = 56 in all worms from all experimental groups. However, the groups irradiated with 20 Gy, 25 Gy or 30 Gy showed variations in the chromosome number, depending on different cells in the same individual. Radiation doses used in this study did not appear to induce chromosome aberrations, however, irradiation with 30 Gy showed slightly reduced chromosome size.

[Effect of gamma-irradiation on infectivity of Clonorchis sinensis metacercariae].

The purpose of the present study was to observe the survival and development of Clonorchis sinensis metacercariae in their final hosts after Co-60 gamma irradiation exerting on both metacercariae isolated or in fish. Guinea pigs or albino rats were orally infected by gavage. Bio-assay, fecal examination for ova and dissection of infected animals were used for the estimation of minimum effective dose of gamma irradiation to control infectivity of Clonorchis sinensis metacercariae. Results showed that the minimum effective irradiation dose for isolated metacercariae was 0.05 kGy. The LD50 of the irradiation dose for metacercariae in fish was 0.05 kGy, and the minimum effective dose was 0.15 kGy. No significant difference in radiation susceptibility to Co-60 gamma irradiation was found among Clonorchis sinensis metacercariae in fishes collected at different localities in the northern, middle or southern parts of China. The present finding suggests that irradiation of the fish at a dose of 0.15 kGy could control the infectivity of Clonorchis sinensis metacercariae and thus be adopted as a control measure in preventive medicine.

Studies on the use of cobalt-60 gamma irradiation to control infectivity of Clonorchis sinensis metacercariae.

The purpose of the present study was to observe the survival and development of Clonorchis sinensis metacercariae in their final hosts after Cobalt-60 gamma irradiation of isolated metacercariae or the parasite in fish. Guinea pigs or albino rats were orally infected with irradiated metacercariae by gavage. Bioassay, fecal examination for ova, and necropsy of infected animals, were the methods used for the estimation of minimum effective dose of gamma irradiation to control infectivity of the metacercariae. Results showed that the minimum effective irradiation dose for isolated metacercariae was 0.05 kGy. The LD50 of the irradiation dose for metacercariae in fish was 0.05 kGy, and the minimum effective dose was 0.15 kGy. No significant difference in radiation susceptibility to Co-60 gamma irradiation was found among C. sinensis metacercariae in fishes collected at different localities in northern, middle or southern parts of China. The present finding suggests that irradiation of fish by 0.15 kGy could control infectivity of C. Sinensis metacercariae and thus be adopted as a control measure in preventing infections.

The effects of gamma irradiation on the survival and development of Clonorchis sinensis metacercariae.

The effects of gamma irradiation on the survival and development of C. sinensis metacercariae were studied to evaluate the feasibility of irradiation as a control measure for clonorchiasis. Pseudorasbora parva were collected at an endemic river of clonorchiasis and were used for irradiation of the fluke in three schemes. The first (Scheme 1) was irradiation of the isolated metacercariae from the fish followed by infection to experimental rats. The second (Scheme 2) was irradiation of the fish, and then the metacercariae were isolated and infected to rats. The third (Scheme 3) was irradiation on the rat livers after infection with normal metacercariae. Irradiation doses varied from 5 to 100 Gy for Schemes 1 and 2, and 10 to 25 Gy for Scheme 3. The rats were sacrificed 2 to 6 weeks after infection. In Scheme 1, the metacercariae irradiated at 50 Gy failed to survive in the rats after 2 or 6 weeks. However, 1 to 44% of the metacercariae irradiated at 5-30 Gy survived. The estimated LD50 of Scheme 1 was 16.5 Gy. The flukes irradiated in Scheme 2 survived better than those in Scheme 1. The average worm recovery rate in 50 Gy was 28%(7-39% individually). Increasing the dose up to 100 Gy brought a remarkably low survival rate of an average 1%(0-3% individually). The LD50 of Scheme 2 was 47.5 Gy. Worm recovery rates in the 10 Gy group of Scheme 3 were 21-39%, and those in the 25 Gy group were 2% and 34%. Although the metacercariae were irradiated, all of the recovered worms were morphologically normal.(ABSTRACT TRUNCATED AT 250 WORDS)