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Korean J Parasitol > Volume 27(4):1989 > Article

Original Article
Korean J Parasitol. 1989 Dec;27(4):231-248. English.
Published online Mar 20, 1994.  http://dx.doi.org/10.3347/kjp.1989.27.4.231
Copyright © 1989 by The Korean Society for Parasitology
Comparative growth and development of the metacercariae of Fibricola seoulensis (Trematoda: Diplostomidae) in vitro, in vivo and on the chick chorioallantois
B S Seo
Department of Parasitology, College of Medicine, Seoul National University, Seoul, Korea.
Abstract

The growth and development of the metacercariae of F. seoulensis cultivated in vitro or on the chick chorioallantois were assessed by comparison with the optimum process of maturation in albino rats and new born chickens. The process of maturation was divided for convenience into six stages: Stage 1; cell multiplication, Stage 2; body shaping, Stage 3; separation of genital anlagen, Stage 4; organogeny, Stage 5; gametogony, and Stage 6; oviposition. In Hank's and Tyrode's solutions, the metacercariae were alive up to 200 days or more at 4 degrees C without any development. The in vivo maturation process in rats or chicks was as follows: stage 1 from 6 hours; stage 2 from 24 hours; stage 3 from 48 to 72 hours; stage 4 from 3 to 4 days; stage 5 from 4 to 5 days; and stage 6 from 5 to 8 days. Despite unsuccessful infection of the metacercariae to 12 day old chicks, fully mature worms of stage 5 or 6 were recovered from new born chicks (1 to 2 days old). The metacercariae of F. seoulensis grown in vitro were up to stage 3 and no further maturation was observed. Of various media employed, the medium NCTC 109 (Gibco) or NCTC 135 (Gibco) supplemented with 20% egg yolk or 20% whole egg macerate or 0.5% yeast was basically required for the earlier development of the fluke. It took 16.1 days (in average) to reach the stage 3 after cultivation. The metacercariae cultivated on the chorioallantoic membranes of 6-13 day old chick embryo at 37-38 degrees C showed their full development up to stage 5 or 6. However, the worms were in general remarkably retarded, compared with those grown in rats or chickens. In the experiments of worm transplant, although the transfer was failed from in vitro culture to in vivo of rats (per os), the transplants from in vitro culture to the chorioallantois and from the chorioallantois to in vivo of rat host were successful with or without development of the transferred worms. In the present study, it was observed that the metacercariae of F. seoulensis can be maintained in vitro media with poor development as well as fully matured in 1 to 2 day-old chicks or on the chorioallantois at a very low rate.

Figures


Fig. 1
Growth and development of F. seoulensis metacercariae in vitro, in vivo and on the chorioallantois according to the length measurements.


Fig. 2
Growth and development of F. seoulensis metacercariae in vitro, in vivo and on the chorioallantois according to the stage determination.


Fig. 3
Growth and development of F. seoulensis metacercariae in vitro, in vivo and on the chorioallantois by stages.


Plate I
Worms recovered from rats.

1. Stage 1, showing increased number of glandular cells in tribocytic organ.

2. Stage 1, 6 hours after infection, increaced g.c. and g.a. in a sectioned preparation.

3-4. Stage 2, one day old worm, showing elongation of hindbody, much increased g.c. and g.a. in a sectioned preparation.

5-6. Stage 3, 2 day old worm, showing separation of developing genital anlagen in toto-mounted and sectioned preparations.

7-8. Stage 4, 4 day old worm, showing organogeny, and early stage of spermatogenesis in a sectioned preparation.

9-10. Stage 5 (6 days old) and stage 6 (9 days old) respectively. showing preovigerous and ovigerous stage.

(a.t.; anterior testis, p.t.; posterior testis, g.a.; genital anlagen, g.c.; glandular cells, o.v.; ovary t.o.; tribocytic organ)



Plate II
Worms recovered from chicks.

1. Stage 1, 6 hour old worm.

2. Stage 2, 1 day old worm.

3 & 4. Stage 4, 3~4 day old worm.

5. Stage 5, 6 day old worm.

6. Stage 6, 9 day old worm.



Plate III
Worms recovered from in vitro cultures.

1. Stage 1.

2. Stage 2.

3. Stage 3.

4. Enlarged from Fig. 3, showing separation of genital primordia.

5. Developed glandular cells around tribocytic organ in stage 3 in a sectioned preparation.

6. Sectioned slide of stage 3, showing enlarged mass of g.a. and increased number of g.c. in tribocytic organ.



Plate IV
Worms recovered from the chorioallantoic membranes.

1 & 2. Stage 3 and enlarged posterior portion.

3 & 4. Stage 4 and enlarged posterior portion.

5 & 6. Stage 4 and enlarged posterior portion.

7. Stage 2.

8. Sectioned slide of stage 5, showing early stage of oogenesis (ov) and spermatogenesis (a.t. & p.t.).

9. Stage 6, showing ovigerous stage.


Tables


Table 1
Development of the metacercariae of F. seoulensis in vivo of albino rats


Table 2
Measurements of the worms from albino rats and chicks (fixd in 10% formalin)


Table 3
Development of the metacercariae of F. seoulensis in vivo of chickens


Table 4
Survival of the metacercariae of F. seoulensis in salt solutions at 4℃


Table 5
Development of the metacercariae of F. seoulensis in vitro media (37℃)


Table 6
Measurements of worms collected from in vitro cultures or from chorioallantoic membranes


Table 7
Development of the metacercariae of F. seoulensis on the chick chorioallantoic membranes (chick embryo, 6~13 days old)


Table 8
Results of transplant experiment

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