Hostname: page-component-7b7567c8c-mnklg Total loading time: 0 Render date: 2026-11-12T07:25:47.180Z Has data issue: false hasContentIssue false
  • English
  • Français

Development of Fasciola hepatica in Lymnaea columella infected with miracidia derived from cattle and marmoset infections

Published online by Cambridge University Press:  01 March 2008

E.A. Mendes,
W.S. Lima and
A.L. de Melo
E.A. Mendes
Affiliation:
Departamento de Parasitologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Av. Antonio Carlos, 6627, CP 486, CEP 30123 970, Minas Gerais, Belo Horizonte, Brazil
W.S. Lima
Affiliation:
Departamento de Parasitologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Av. Antonio Carlos, 6627, CP 486, CEP 30123 970, Minas Gerais, Belo Horizonte, Brazil
A.L. de Melo*
Affiliation:
Departamento de Parasitologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Av. Antonio Carlos, 6627, CP 486, CEP 30123 970, Minas Gerais, Belo Horizonte, Brazil
*
*Author for correspondence Fax: (+55 31) 3409 2970 E-mail: aldemelo@icb.ufmg.br
Rights & Permissions [Opens in a new window]

Abstract

The development of Fasciola hepatica from two species of definitive hosts, i.e. cattle (Bos taurus) and a marmoset (Callithrix penicillata) in the snail Lymnaea columella was determined based on the production of rediae and cercariae and snail survival rate. More rediae and cercariae at 60–74 days post-infection were produced by snails infected by cattle-derived miracidia (cattle group) than by those infected by marmoset-derived miracidia (marmoset group). Among the L. columella parasitized by the marmoset group, the survival rate and the percentage of positive snails were higher than among those parasitized by the cattle group. Eggs of F. hepatica released in cattle faeces were significantly bigger than those released in marmoset faeces. Miracidia originating from parasites that completed their development in cattle were more efficient in infecting the intermediate host. These results suggest that vertebrate-host origin influences the eggs produced by the parasite and the infection rates in the snail host L. columella.

Information

Type
Research Papers
Information
Journal of Helminthology , Volume 82 , Issue 1 , March 2008, pp. 81 - 84
Copyright
Copyright © Cambridge University Press 2007

Introduction

The liver fluke, Fasciola hepatica is a helminth parasite of the bile ducts of vertebrates, both wild and domestic. It is associated with great economic losses, mainly through reduced meat, milk and wool production, condemnation of livers, veterinary medication costs and high mortality rates that affect herds of domestic animals such as cattle and sheep. Wild animals are important reservoirs in some regions, contributing to the maintenance and dissemination of the parasite.

The worldwide distribution of F. hepatica is related to climatic factors such as availability of humid habitats subject to flooding as well as the presence of definitive and especially intermediate hosts, the snails of the genus Lymnaea (Boray, Reference Boray1969; Costa et al., Reference Costa, Silva Santos and Botelho1987). The definitive host species in which the parasite develops plays an important role in determining the success of infection in the intermediate host (Vignoles et al., Reference Vignoles, Ménard, Rondelaud, Agoulon and Dreyfuss2004). Definitive host species eliminate viable eggs in their faeces and the development of miracidia occurs in the environment. The miracidium must then penetrate into a susceptible snail, the intermediate host. In the snail tissues the larvae undergo some changes, to develop to rediae and later to cercariae. Soon after their emergence from the host, the cercaria changes to metacercaria. To complete the life cycle metacercariae must be ingested by definitive hosts.

Morphological characterization of F. hepatica has produced significant knowledge about its biology, since the pioneering research relative to many aspects of the parasite and its host (Leuckart, Reference Leuckart1882; Dawes, Reference Dawes1961, Reference Dawes1962). Since then, some criteria, such as the full somatic development and the ability to eliminate eggs in faeces, have been used to establish the correlation of development and maturation of the worm to effectiveness in infecting experimental laboratory or wild mammals (Boray, Reference Boray, Hillyer and Hopla1982).

Several studies have shown that some morphological characteristics of F. hepatica may vary, depending on the host species or the parasite strain as well as the intermediate host species (Vignoles et al., Reference Vignoles, Ménard, Rondelaud, Chauvin and Dreyfuss2001).

In the present study the influence of the definitive host on the development of F. hepatica in L. columella was investigated by infecting snails with miracidia derived from cattle or marmoset origin. Morphometric analysis of the parasite eggs released from each host species was also performed.

Material and methods

Parasite and animals

Eggs of F. hepatica were obtained from faeces of naturally infected cattle (Bos taurus) from farms in the municipality of Itajubá, state of Minas Gerais, Brazil, and faeces of marmosets (Callithrix penicillata) from the breeding marmoset colony of the Federal University of Minas Gerais, experimentally infected according to Girão & Ueno (Reference Girão and Ueno1985). The faeces were transferred to Petri dishes containing dechlorinated water, in which they were incubated at 27°C for 14 days to allow miracidia to develop.

Infection of snails

Specimens (n = 700) of laboratory-reared L. columella (Itajubá strain), measuring 4–8 mm, were used for experimental infection. Six hundred snails were divided in groups of 100 specimens which were individually placed in wells of culture plates (Plates Kartel S.P.A.) with 2 ml dechlorinated water and four miracidia. The plates were covered and left under the light of a 60 W lamp placed 50 cm away for 24 h. At the end of this period each well was inspected with the aid of a stereomicroscope to confirm infection. Three hundred snails were infected by F. hepatica miracidia emerging from eggs in cattle faeces (cattle group). The other 300 were infected by miracidia originating from marmoset infections (marmoset group). The control group was formed by 100 snails not exposed to miracidia. Snails were maintained in plastic containers at room temperature and fed on lettuce Lactuca sativa. Infected snails were separated in groups of 100 individuals. Five snails of each group were placed in individual Petri dishes and dissected under a stereomicroscope to search for developmental stages of F. hepatica daily from 60 to 74 days post-infection (DPI). The rediae and cercariae obtained from the living snails after dissection at the end of the experimental period were not included in the results.

Parameters

Survival of snails in each group was monitored daily. The percentage of positive snails was calculated in relation to the number of snails examined in each observation-interval group. The relationship between developmental stage productions by L. columella infected by miracidia hatching from eggs in the faeces of each host was evaluated by the Mann–Whitney U-test. The Poisson model and statistical software R 2.2.1 were used to analyse the variables of infection time (DPI) and numbers of cercariae and rediae produced each day by infections derived from each host.

Morphological differences were verified by measuring the length and width of 100 F. hepatica eggs from each host. Morphometrical analyses were carried out based on measurements made with an optical microscope with eyepiece micrometer under the 10 ×  objective. Analysis of variance (ANOVA) was used to compare the mean lengths and widths of eggs of each host.

Results

The survival rate of snails infected by cattle-derived miracidia (cattle group) was lower (41.0%) than that of snails infected by marmoset-derived miracidia (marmoset group) (56.0%). The differences among all groups ranged from 9% to 26%. The snails infected by the marmoset group and examined at 60–64 DPI presented higher survival rates than the other groups. The survival rates at 70–74 DPI for the cattle group snails (9.0%) and the marmoset group (11.0%) showed similar values. For both types of hosts the survival rate at 60–64 DPI was higher than that at 70–74 DPI (table 1).

Table 1

Numbers and percentage of positive and surviving snails and number and means of developmental stages produced by Lymnaea columella infected by Fasciola hepatica miracidia of cattle or marmoset origin at different days post-infection (DPI).

The percentage of positive snails from the marmoset group infections (28.0%) was higher than in those infected by the cattle group (25.3%). The percentage of positive snails in all groups varied from 16.0% at 70–74 DPI to 36.0% in the group examined at 65–69 DPI. The highest value for positive snails was obtained at 65–69 DPI for both groups (table 1). From those snails surviving, only two out of 41 from the cattle group and only three out of 56 from the marmoset group were positive for larval stages.

The production of rediae and cercariae is shown in table 1. The rediae produced in the cattle group infection (929), were similar to those obtained in the marmoset group infections (913); however, while the cattle group snails dissected produced more rediae (394) at 65–69 DPI, the mean number of rediae per snail was highest (83.0 ± 30.3) at 70–74 DPI for the marmoset group.

Lymnaea columella produced more cercariae (13,588) when infected by the cattle group miracidia than those infected by the marmoset group (12,060). The cercariae obtained during the experimental period varied from 3345 to 5641 for the cattle group and from 2040 to 5486 for the marmoset group. In the cattle group infected L. columella, the snails examined between 65 and 69 DPI presented the highest number of cercariae. In the marmoset group infected L. columella, the highest number of cercariae was obtained at 60–64 DPI. The highest mean numbers of cercariae were produced by both the cattle group and the marmoset group infected L. columella at 60–64 DPI (767.0 ± 608.7 per snail and 685.7 ± 346.9 per snail, respectively) (table 1).

During the daily examination it was verified that the cattle group infections produced more cercariae and rediae at 68 DPI. The greatest numbers of cercariae and rediae from the marmoset group infections were obtained at 64 DPI and 70 DPI, respectively (data not shown). After 71 DPI, significant drops were observed in the number of developmental stages produced by L. columella infected by both groups of miracidia, which continued until the end of the experiment at day 74. Moreover, during the observation period, the developmental stages produced by L. columella tended to diminish. Significant differences (P < 0.05), occurred in relation to the initial and final numbers of cercariae and rediae produced by L. columella infected by miracidia derived from both hosts.

The measurements of 100 F. hepatica eggs released in faeces from each host showed significant difference (P < 0.00001) in mean length. The eggs from cattle faeces were longer (138.4 μm ± 0.1) than those derived from marmoset faeces (133.6 μm ± 0.1), while no difference was observed for width values (P = 0.07) from both hosts (marmosets 71.6 ± 0.1 μm; cattle 73.1 ± 0.9 μm).

Discussion

The larval development of the parasite into the snail host and egg morphometric analyses were different for cattle and for marmosets, suggesting worm variability depending on peculiarities such as the definitive host. It has been established that the host influences the phenotype of some parasites and this influence may be attributed also to the helminth physiology, the worm burden and the host immune system (Senft et al., Reference Senft, Gibler and Knopf1978). From the data obtained in this study, snails that had been parasitized by miracidia derived from naturally infected cattle produce more cercariae than those infected by miracidia from experimentally infected marmosets, suggesting that bovines were more competent in transmitting the parasite. Rondelaud & Dreyfuss (Reference Rondelaud and Dreyfuss1995) showed that the success of experimental infection of snails differs strongly between liver-fluke miracidia obtained from different definitive hosts. In Brazil, ruminants are the common hosts of F. hepatica and the environment in which they are being parasitized plays a significant role in maintaining the parasite life cycle. Nevertheless, F. hepatica from experimentally infected marmosets also originated viable eggs, the miracidia being able to infect L. columella and to produce rediae and cercariae. The survival rate of snails harbouring developmental stages from cattle was lower than that for those resulting from marmoset infections (41% vs. 56%), maybe because of mechanical and chemical tissue damage provoked by sporocyst migration, nutrition and maturation of rediae (Dawes, Reference Dawes1956; Kendall, Reference Kendall1965), which were in higher numbers than those from marmosets, resulting in more cercariae. The percentage of positive snails harbouring cercariae from the cattle group infection suggests that miracidia from such origin are more adapted to invade and develop into L. columella. The results of the present study seem to demonstrate that the definitive host in which the parasite develops plays an important role in the infection of the intermediate host, with implications for dispersal of the parasite. Several wild animal species have been incriminated as responsible for maintenance and transmission of F. hepatica (Hurtrez-Boussés et al., Reference Hurtrez-Boussés, Meunier, Durand and Renaud2001). Costa et al. (Reference Costa, Silva Santos and Botelho1987) verified that the coypu Myocastor coypus can act as one of the principal reservoirs and probable disseminator of F. hepatica at the ‘Estação Experimental Zootécnica de Dom Pedrito’, in the Brazilian state of Rio Grande do Sul. The occurrence of morphological variability among eggs of F. hepatica derived from two host species, as demonstrated in the present study, could be considered an example of a host-mediated effect on development. These differences could indicate different levels of susceptibility of different hosts to infection (Abrous et al., Reference Abrous, Comes, Gasnier, Rondelaud, Dreyfuss, Chauvin, Ménard, Agoulon and Cabaret1998). In fact, the eggs laid by adult parasites were directly affected by the conditions and type of definitive host in which they develop (Dixon, Reference Dixon1964; Haroun & Hillyer, Reference Haroun and Hillyer1986). Based on the comparison of developmental stage productivity and of morphological variability in F. hepatica eggs derived from the cattle and the marmoset group, it appears that factors imposed on each of these hosts could be acting directly on the adult parasite. These may be related to genetic or epigenetic effects, resulting in several phenotypes (Abrous et al., Reference Abrous, Comes, Gasnier, Rondelaud, Dreyfuss, Chauvin, Ménard, Agoulon and Cabaret1998; Vignoles et al., Reference Vignoles, Ménard, Rondelaud, Agoulon and Dreyfuss2004). A significant reduction verified at 71 DPI and similar reductions in the numbers of developmental stages in L. columella parasitized by miracidia from the cattle and the marmoset group may be related to their viability and origin, as well as the ability of snails to maintain infections for long periods. The numbers of cercariae and rediae produced by the cattle group miracidia at 70–74 DPI were lower than those obtained during the other intervals, suggesting a rapid parasite development in the early stages, probably associated with their viability. The number of cercariae originated from marmoset infections in the same interval (70–74 DPI) was also lower than that obtained in other intervals, although the number of rediae produced was higher. This suggests that marmoset-derived miracidia showed poorer development, with prolonged cercaria formation and maturation time. Thus, it can be concluded that there was some factor related to the type of definitive host that stimulated or inhibited worm development and egg production and, in consequence, morphometric variation of the eggs produced that influenced parasite development in the intermediate host and its subsequent dissemination.

Acknowledgements

Thanks are due to the anonymous referees for helpful comments on the manuscript.

References

Abrous, M., Comes, A.M., Gasnier, N., Rondelaud, D., Dreyfuss, G., Chauvin, A., Ménard, A., Agoulon, A. & Cabaret, J. (1998) Morphological variability in Fasciola hepatica eggs in ruminants, rodents and lagomorphs. Journal of Helminthology 72, 313–317.CrossRefGoogle ScholarPubMed
Boray, J.C. (1969) Experimental fascioliasis in Australia. Advances in Parasitology 7, 95–210.CrossRefGoogle ScholarPubMed
Boray, J.C. (1982) Fascioliasis. pp. 71–88in Hillyer, G.V. & Hopla, C.E. (Eds) Handbook series in zoonoses Section C. Parasitic zoonoses. Vol. III. Boca Raton, Florida, CRC Press.Google Scholar
Costa, N.C., Silva Santos, I.C. & Botelho, G.A. (1987) Myocastor coypus Molina, 1782 (Rodentia, Echymidae) naturalmente infectado com Fasciola hepatica (Linnaeus 1758 (Trematoda, Fasciolidae). A Hora Veterinária 36, 24–26.Google Scholar
Dawes, B. (1956) The Trematoda. 644 pp. Cambridge, Cambridge University Press.Google Scholar
Dawes, B. (1961) On the early stages of Fasciola hepatica penetrating into the liver of an experimental host, the mouse: a histological picture. Journal of Helminthology, R.T. Leiper Supplement, pp. 41–52.CrossRefGoogle Scholar
Dawes, B. (1962) On the growth and maturation of Fasciola hepatica L. in mouse. Journal of Helminthology 36, 11–38.CrossRefGoogle Scholar
Dixon, K.E. (1964) The relative suitability of sheep and cattle as hosts for the liver fluke, Fasciola hepatica L. Journal of Helminthology 38, 203–212.CrossRefGoogle ScholarPubMed
Girão, E.S. & Ueno, H. (1985) Técnica de quatro tamises para o diagnóstico coprológico quantitativo da Fasciolose dos ruminantes. Pesquisa Agropecuária Brasileira 20, 905–912.Google Scholar
Haroun, E.M. & Hillyer, G.V. (1986) Resistance to fascioliasis. A review. Veterinary Parasitology 20, 63–93.CrossRefGoogle ScholarPubMed
Hurtrez-Boussés, S., Meunier, C., Durand, P. & Renaud, F. (2001) Dynamics of host–parasite interactions: the example of population biology of the liver fluke (Fasciola hepatica). Microbes and Infection 3, 841–849.CrossRefGoogle ScholarPubMed
Kendall, S.B. (1965) Relationships between the species of Fasciola and molluscan host. Advances in Parasitology 3, 59–98.CrossRefGoogle Scholar
Leuckart, R. (1882) Zur Entwicklungsgeschichte des Leberegels (Distomum hepaticum). Archive für Naturgeschichte 48, 80–119.Google Scholar
Rondelaud, D. & Dreyfuss, G. (1995) Fasciola hepatica: the influence of the definitive host on the characteristics of infection in the snail Lymnaea truncatula. Parasite 2, 275–280.CrossRefGoogle Scholar
Senft, A.W., Gibler, W.B. & Knopf, P.M. (1978) Scanning electron microscope observations on tegumental maturation in Schistosoma mansoni grown in permissive and non-permissive hosts. American Journal of Tropical Medicine and Hygiene 27, 258–266.CrossRefGoogle ScholarPubMed
Vignoles, P., Ménard, A., Rondelaud, D., Chauvin, A. & Dreyfuss, G. (2001) Fasciola hepatica: the characteristics of experimental infections in Lymnaea truncatula subjected to miracidia differing in their mammalian origin. Parasitology Research 87, 945–949.CrossRefGoogle ScholarPubMed
Vignoles, P., Ménard, A., Rondelaud, D., Agoulon, A. & Dreyfuss, G. (2004) Fasciola hepatica: the growth and larval productivity of redial generations in Galba truncatula subjected to miracidia differing in their mammalian origin. Journal of Parasitology 90, 430–433.CrossRefGoogle ScholarPubMed
Figure 0

Table 1Numbers and percentage of positive and surviving snails and number and means of developmental stages produced by Lymnaea columella infected by Fasciola hepatica miracidia of cattle or marmoset origin at different days post-infection (DPI).