Gamma선이 鉤蟲幼蟲 및 宿主의 抵抗力에 미치는 영향

Abstract

[한글]

Influences of Gamma-ray Irradiation upon the Resistance of Hookworm and the Hosts

Jae-Ho Cho, M.D.

Department of Parasitology

Yonsei University College of Medicine, Seoul, Korea

(Director: Professor Chin Thack Soh, M.D.

There have been many reports about the influence of irradiation upon the eggs and

larvae of nematodes.

Parthes (1904) first reported that growth and maturity of ascaris eggs were

inhibited by X-ray irradiation. Tyzzer and Honeij(1916) and Zaiman et al.(1655)

reported that Trichinella spiralis larvae attenuated by exposure to a suitable dose

of gamma-irradiation induced a certain degree of resistance to challenging

infection with normal larvae, and stimulated production of antibodies. Miller(1964,

1965 b, c) found that a single subcutaneous infection with 40-KR-irradiated larvae

showed a highly significant resistance to a subsequent challenge of normal larvae

and a sterilizing effect on the female worm of Ancylostoma caninum. Naiman(1944)

reported that X-ray irradiation of rats increased the severity of Trypanosoma

lewisi infection, and diminished the ablastic antibody production.

The present investigation was designed to study on the influence of

gamma-irradiation upon the following areas; infectivity and reproductivity of

hookworm, pathogenicity and productivity of antibodies in the host

Ancylostoma caninum eggs were obtained from a dog feces and cultured for 7 days

at 25∼30。C. The filariform larvae were inoculated subcutanequslyon the thigh

region of experimental animals. Parasites free dogs and rats were used for

experiments.

Serum collection: The dog serum collected after 15min centrifugation at 3,000rpm

with 2 to 3 ml of their blood which was placed at room temperature for 20 minutes.

The blood of rats was taken from their cut tails with the capillary tube, and the

sera were obtained after 5 minutes centrifugation at 11,000rpm by Adams Autocrit.

Total serum protein was measured by Micro-Kjeldahl method and the serum protein

fraction was checked using paper electrophoresis in which serum was run to the

Whatman No. 1 filter paper for 16 hours at 70 volt. The reading was done by Junior

spectrophotometer in 520 mμ light length. Irradiation: Gamma-ray irradiation was

performed with a "Gamma cell 220" Cobalt-60 irradiator. Each of the larval pools

and rats were exposed to the following dosages;

Pool 0: No irradiation Pool 1: 5,000 R

Pool 2: 50,000 R Pool 3: 100,000 R

Pool 4: 1,000,000 R

Control rat : No irradiation Rat Ⅰ: 100 R

Rat Ⅱ: 500 R Rat Ⅲ: 1,000 R

Egg productivity: E.P.G(Eggs per gram of feces) was counted 21 to 46 days after

the infection by Stoll's eggs counting method at five day interval. Infectivity:

The rats were sacrificed 3 to 30 days after skin infection of the irradiated

larvae, and the migrating larvae were collected by Baermann's larvae collection

method from lungs, liver and carcase of the experimental animals. In dogs, adult

worms were collected from in he small bowel of the host 5 weeks after the

infestation.

Pathogenicity: The routine pathological examinations of lungs, liver and skin of

the sacrificed rats were carried out.

The results are summarize as follow:

Infectivity and reproductivity on gamma-irradiated larvae of Ancylostoma caninum:

In rats the proper host for A.caninum, the migrating larvae were collected from

lung, liver and muscles of the sacrificed animals.

In the 5,000 R-group, 25.7% the larvae were recovered three days after infection

with 1,000 irradiated larvae, and 29.1% in 100,000 R-group and 14.9% in 1,000,000

R-group respectively, while in the control group showed 38.3%b recovery.

In the control group, 74 larvae were recovered 30 days after infection with 1,000

non-irradiated filariform larvae. None was seen in both 50,000R and 100,000R

groups.

Whereas in 1,000,000 R-group the larvae was not found at 15 days after infection.

In the experimental groups for the challenge infection of non-irradiated larvae,

247 larvae were recovered in 5,000 R-group, 165 in 50,000 R-group, 254 in 100,000

R-group and 348 in the control group respectively.

In dogs, the proper host to A.caninum, 25 adult worms were recovered in 5,000

R-group after infected with 100 irradiated filariform larvae, 41 adults in the

control group. However, the adult was not found at all in 100,000 R-group.

In 5,000 R-group 41 adults were recovered after the challenging infection on

control larvae, but 57 adults in 100,000 R-group and 76 adults in the control

group. A single infection of 5,000 to 50,000 R-irradiated larvae showed a higher

resistance to a subsequent challenge of normal larvae.

The E.P.G. of 5,000 R-group in the dog decreased compared with control group

during 36 days observation after infection of irradiated filariform larvae, and

none was seen in 100,000 R-irradiated group.

Serum γ-globulin of rats and dogs infected with gamma-irradiated larvae of

A.caninum:

In all groups of rats, the abnormal host to A.caninum, serum γ-globulin

increased remarkably from 10.2% to 22.5% in comparison with the normal control

(10.2% to 19.6%) and to 100,000 R-group (10.0% to 21.1%). In dogs, the normal host

to A.caninum, γ-globulin tended to be similar to the rat groups. On the contrary,

A/G ratio decreased in al infection group during 5 weeks after infection of

hookworm.

Pathogenicity of rats infected with gamma-irradiated larvae of A.caninum:

In the rats infected with 5,ooo R-irradiated larvae, some inflammatory process

and hemorrhages were found in the lung but tissues were similar to the normal rats.

The pathological changes of the skin were similar with the picture of the lungs.

Infectivity and serum γ-globulin on gamma-irradiated rats infected with

non-irradiated larvae of A.caninum:

In infectivity slightly increased in 1,00 R-group in comparison to the normal

control rats. Serum γ-globulin decreased slightly in 100 R-group three weeks after

infection.

Whereas 1,000 R-irradiated rats which were infected with the normal larvae were

died within two weeks after infection.

The above results indicate that:

1. The infectivity of the gamma-ray irradiated filariform larvae of A.caninum was

inhibited at the dosage of 50,000R

2. The reproductivity of the gamma-ray irradiated filariform larvae of A.caninum

decreased at dosage of 5,000R

3. The pathogenicity of the gamma-ray irradiated filariform larvae wa milder than

contry group. The more the dosage of gamma-ray irradiation increased, the milder

pathological changes were occured.

4. The γ-globulin in serum of the host enhanced remarkably in the group of

50,000 R irradiated filariform larvae, and then the A/G ratio was decreased

according to the increase of γ-globulin.

5. In the irradiated host the discovery of the migrating larvae increased

slightly at the dosage of 1,000R. on the other hand the γ-globulin decreased

slightly in 100 R-irradiated host.

The above results indicate that gamma-ray irradiation decreases the infectivity

and reproductivity of the parasite. On the other hand, the irradiation upon the

host decreases the resistance of the host against parasite infection.

[영문]

There have been many reports about the influence of irradiation upon the eggs and larvae of nematodes.

Parthes (1904) first reported that growth and maturity of ascaris eggs were inhibited by X-ray irradiation. Tyzzer and Honeij(1916) and Zaiman et al.(1655) reported that Trichinella spiralis larvae attenuated by exposure to a suitable dose

of gamma-irradiation induced a certain degree of resistance to challenging infection with normal larvae, and stimulated production of antibodies. Miller(1964, 1965 b, c) found that a single subcutaneous infection with 40-KR-irradiated larvae

showed a highly significant resistance to a subsequent challenge of normal larvae and a sterilizing effect on the female worm of Ancylostoma caninum. Naiman(1944) reported that X-ray irradiation of rats increased the severity of Trypanosoma lewisi infection, and diminished the ablastic antibody production.

The present investigation was designed to study on the influence of gamma-irradiation upon the following areas; infectivity and reproductivity of hookworm, pathogenicity and productivity of antibodies in the host

Ancylostoma caninum eggs were obtained from a dog feces and cultured for 7 days at 25∼30。C. The filariform larvae were inoculated subcutanequslyon the thigh region of experimental animals. Parasites free dogs and rats were used for

experiments.

Serum collection: The dog serum collected after 15min centrifugation at 3,000rpm with 2 to 3 ml of their blood which was placed at room temperature for 20 minutes.

The blood of rats was taken from their cut tails with the capillary tube, and the sera were obtained after 5 minutes centrifugation at 11,000rpm by Adams Autocrit.

Total serum protein was measured by Micro-Kjeldahl method and the serum protein fraction was checked using paper electrophoresis in which serum was run to the Whatman No. 1 filter paper for 16 hours at 70 volt. The reading was done by Junior

spectrophotometer in 520 mμ light length. Irradiation: Gamma-ray irradiation was performed with a "Gamma cell 220" Cobalt-60 irradiator. Each of the larval pools and rats were exposed to the following dosages;

Pool 0: No irradiation Pool 1: 5,000 R

Pool 2: 50,000 R Pool 3: 100,000 R

Pool 4: 1,000,000 R

Control rat : No irradiation Rat Ⅰ: 100 R

Rat Ⅱ: 500 R Rat Ⅲ: 1,000 R

Egg productivity: E.P.G(Eggs per gram of feces) was counted 21 to 46 days after the infection by Stoll's eggs counting method at five day interval. Infectivity: The rats were sacrificed 3 to 30 days after skin infection of the irradiated larvae, and the migrating larvae were collected by Baermann's larvae collection

method from lungs, liver and carcase of the experimental animals. In dogs, adult worms were collected from in he small bowel of the host 5 weeks after the infestation.

Pathogenicity: The routine pathological examinations of lungs, liver and skin of the sacrificed rats were carried out.

The results are summarize as follow:

Infectivity and reproductivity on gamma-irradiated larvae of Ancylostoma caninum:

In rats the proper host for A.caninum, the migrating larvae were collected from lung, liver and muscles of the sacrificed animals.

In the 5,000 R-group, 25.7% the larvae were recovered three days after infection with 1,000 irradiated larvae, and 29.1% in 100,000 R-group and 14.9% in 1,000,000 R-group respectively, while in the control group showed 38.3%b recovery.

In the control group, 74 larvae were recovered 30 days after infection with 1,000 non-irradiated filariform larvae. None was seen in both 50,000R and 100,000R groups.

Whereas in 1,000,000 R-group the larvae was not found at 15 days after infection.

In the experimental groups for the challenge infection of non-irradiated larvae, 247 larvae were recovered in 5,000 R-group, 165 in 50,000 R-group, 254 in 100,000 R-group and 348 in the control group respectively.

In dogs, the proper host to A.caninum, 25 adult worms were recovered in 5,000 R-group after infected with 100 irradiated filariform larvae, 41 adults in the control group. However, the adult was not found at all in 100,000 R-group.

In 5,000 R-group 41 adults were recovered after the challenging infection on control larvae, but 57 adults in 100,000 R-group and 76 adults in the control group. A single infection of 5,000 to 50,000 R-irradiated larvae showed a higher resistance to a subsequent challenge of normal larvae.

The E.P.G. of 5,000 R-group in the dog decreased compared with control group during 36 days observation after infection of irradiated filariform larvae, and none was seen in 100,000 R-irradiated group.

Serum γ-globulin of rats and dogs infected with gamma-irradiated larvae of A.caninum:

In all groups of rats, the abnormal host to A.caninum, serum γ-globulin increased remarkably from 10.2% to 22.5% in comparison with the normal control (10.2% to 19.6%) and to 100,000 R-group (10.0% to 21.1%). In dogs, the normal host

to A.caninum, γ-globulin tended to be similar to the rat groups. On the contrary, A/G ratio decreased in al infection group during 5 weeks after infection of hookworm.

Pathogenicity of rats infected with gamma-irradiated larvae of A.caninum:

In the rats infected with 5,ooo R-irradiated larvae, some inflammatory process and hemorrhages were found in the lung but tissues were similar to the normal rats.

The pathological changes of the skin were similar with the picture of the lungs.

Infectivity and serum γ-globulin on gamma-irradiated rats infected with non-irradiated larvae of A.caninum:

In infectivity slightly increased in 1,00 R-group in comparison to the normal control rats. Serum γ-globulin decreased slightly in 100 R-group three weeks after infection.

Whereas 1,000 R-irradiated rats which were infected with the normal larvae were died within two weeks after infection.

The above results indicate that:

1. The infectivity of the gamma-ray irradiated filariform larvae of A.caninum was inhibited at the dosage of 50,000R

2. The reproductivity of the gamma-ray irradiated filariform larvae of A.caninum decreased at dosage of 5,000R

3. The pathogenicity of the gamma-ray irradiated filariform larvae wa milder than contry group. The more the dosage of gamma-ray irradiation increased, the milder pathological changes were occured.

4. The γ-globulin in serum of the host enhanced remarkably in the group of 50,000 R irradiated filariform larvae, and then the A/G ratio was decreased according to the increase of γ-globulin.

5. In the irradiated host the discovery of the migrating larvae increased slightly at the dosage of 1,000R. on the other hand the γ-globulin decreased slightly in 100 R-irradiated host.

The above results indicate that gamma-ray irradiation decreases the infectivity and reproductivity of the parasite. On the other hand, the irradiation upon the host decreases the resistance of the host against parasite infection.