0020-7519/90 S3.00 + 0.00 Pergamon Press plc Aurrrolim Societyfor Porariro/ogy
Inkmotional Journalfor Parasitology Vol. 20, No. 2, pp. 263-264, 1990 Printed in Grmt Britain
RESEARCH NOTE
EXPERIMENTAL INFECTION OF MICE WITH TOXOCARA LARVAE OBTAINED FROM JAPANESE QUAILS
CANIS
T. K. PAHARI and N. K. SASMAL* Department
of Parasitology,
Faculty of Veterinary and Animal Sciences, Bidhan Chandra Agricultural University, Mohanpur 741252, W.B., India (Received 12 December 1989; accepted 20 December 1989)
Ahstracf-PAHAaI T. K. and SASMAL N. K. 1990. Experimental infection of mice with Toxocara canis larvae obtained from Japanese quails. International Journal for Parasitology 20: 263-264. The migration and distribution of Toxocara canis larvae in the tissues of Japanese quails, infected orally with 5 x 10’ infective eggs, were studied, as well as the re-infectivity of these larvae in mice, inoculated with 50 larvae obtained from the liver of these quails. Post-infection, the highest concentrations of larvae were found to be present in the liver of quails while only a few migrated to other tissues like lungs, heart, muscle and brain. The migration and distribution of the larvae in the tissues of mice were studied by necropsy on days 6 and 12 postinfection. On both days the highest number of larvae, 11 and 10, were recovered from the carcase followed by six and seven from the leg muscles and four and eight from the brain, respectively. A few larvae were recovered from the liver, lungs and viscera. This implies that the larvae had a special affinity for the muscle and brain tissue of mice, unlike in the quails. The role of these larvae in relation to paratenism is discussed.
INDEX KEY WORDS: Toxocaru cunis; migration; Japanese quails; mice; paratenism.
behaviour of larvae in different abnormal hosts have been performed (Sprent, 1952, 1955; Beaver, 1956; Done, EXPERIMENTALstudies on the migratory T. cunis
Richardson KcGibson, 1960; Schaeffler, 1960; Galvin, 1964) and it is presumed that larvae are transferred in nature from animal to animal through paratenism (Sprent, 1954). The experimental re-infectivity of Toxocara was reported to be comparatively higher
than any other ascarids and the infections were retained for a longer period in the abnormal hosts (Ishii, 1959; Okoshi & Usui, 1968). Several previous reports have discussed the migratory behaviour of T. cunis larvae in laboratory mammalian hosts, but none have examined this question in Japanese quails (Coturnix coturnix juponica). Accordingly the present study was undertaken to clarify this aspect and to verify the re-infectivity of these larvae to mice. Eggs were obtained by dissecting gravid T. canis females, incubated in0.5% formalin at 27°C f 1°C for 46 weeks, and washed in water to remove formalin before use. Healthy 5-6-week-old male laboratoryreared Japanese quails, weighing 17&l 80 g, and 46week-old albino mice of either sex, weighing 25-30 g were used in experiments. They were maintained in small metal cages and provided with a commercial diet and water ad lib. Strict infection-free status was maintained throughout the experiment. The calcu-
* To whom all correspondence should be addressed.
lated dose of embryonated eggs and infective larvae in water suspension was determined by the method of Oshima (1961) and the animals were dosed by stomach intubation. The quails and the mice were starved for 6 h prior to and 2 h post-infection. Twelve Japanese quails were each infected with 5 x lo3 infective eggs and were necropsied in groups of four on days lo,20 and 30 post-infection. Similarly, three uninfected control birds were also necropsied at intervals ranging from 10 to 30 days. Eight mice were each inoculated with 50 actively motile second-stage larvae isolated from the liver of Japanese quails on day 30 postinfection and were necropsied in groups of two on days 6 and 12 post-infection. Similarly, two uninfected control mice were also killed at intervals ranging from 6 to 12 days. Quails and mice were killed with chloroform. Immediately following death, the alimentary tract, liver, lungs, heart, spleen, kidney, muscle, carcase and brain were removed and rinsed in saline. Larvae were recovered from organs and tissues by digestion in pepsin as described by Sprent (1952) with some modification, and by Baermann isolation from minced fresh tissues. Larvae in the brain were counted directly by squashing the tissues between two glass slides. The larvae were identified according to Nichols (1956) and Sprent (1955). The results have been summarized in Table 1. On days 10, 20 and 30, 93-95% of all recovered larvae were in the liver of quail and very few in muscle, brain and heart. In the mice on both days 6 and 12 postinfection, the highest number of larvae (11 and 10, 263
264
T. K. PAHARI and N. K. SASMAL
TABLE I-NUMBER OF LARVAERECOVERED FROMTISSUESOF JAPANESEQUAILINOCULATED WITH5 x 10 3 INFECTIVE EGGSOF Toxocara canis No. of quails
Average
Days infected Stomach*
4 4 4
Intestine
285 296 304
10 20 30
lProventriculus
Liver
Lungs 1
larvae recovered Heart 4 5 5
from
Thigh muscle
Breast muscle
Neck muscle
3 4 5
2 4 6
5 2 4
Brain
2 4
Total larvae recovered 300 313 328
and gizzard tissues.
respectively) were recovered from the carcase followed by the leg muscle (six and seven) and brain (four and eight). In comparison to these, a few larvae were
recovered from the liver, lungs and viscera; two and none, four and two, one and none, respectively. The percentage inoculum recovered at necropsy in the first case was 56 and the second case, 54. No morphological differences were found among the larvae from Japanese quails and mice. They were all second-stage larvae. No larvae were recovered from the control groups at necropsy. The infection of mice with T. canis larvae of quail origin indicates that Japanese quail may possibly serve as a paratenic host of T. canis, like chickens and pigeons (Beaver, 1956; Galvin, 1964; Okoshi & Usui, 1968). It is evident that almost all larvae accumulated in the liver of quails. This special affinity of larvae for avian liver tissue was suggested by Galvin (1964) and Okoshi & Usui (1968) in chickens and pigeons. Migration to other tissues in pigeons has also been reported by Galvin (1964). Larvae recovered from the liver of quail were used for infection to mice. In the mice most of these larvae migrated to organs beyond the liver. In the present experiment there is some suggestion that more larvae accumulated in the brain with increasing time as reported by Sprent (1955) and Dunsmore, Thompson & Bates (1983). It is clear from the present investigation that larvae of T. canis can establish in both the abnormal hosts, quails and mice, and the larvae of quail origin retain their infectivity to mice even after day 30 post-infection. However, in respect to larval recovery the adaptability of the larvae to mammalian tissue was found to be greater than to the avian tissue which is in conformity with Okoshi & Usui (1968) and Mittra & Sasmal (1985). The larvae from both sources were morphologically identical as reported by Nichols (1956) Ishii (1959) and Okoshi & Usui (1968) in their studies on the transfer of larvae from mouse to mouse and chicken to mouse. These findings are in agreement with those of Sprent (1954) and Okoshi & Usui (1968) who reported that effective transfer of larvae from animal to animal may always be possible in the natural world by means of paratenism.
Acknowledgemenrs-The authors are grateful to the Bidhan Chandra Agricultural University for financial assistance and express their gratitude to Prof. H. L. Shah, Department of Parasitology, Jabalpur Veterinary College, for his scientific advice.
REFERENCES BEAVER P. C. 1956. Larva migrans. Experimental Parasirology 5: 587-621. DONE J. T., RICHARDSON M. D. & GIBSON T. E. 1960. Experimental visceral larva migrans in the pig. Research in Veterinary Science 23: 109-l 14. DUNSMOREJ. D., THOMPSONR. C. A. & BATESI. A. 1983. The accumulation of Toxocara canis larvae in the brains of mice. International Journal for Parasitology 13: 5 17-521. GALVINT. J. 1964. Experimental Toxocara canis infections in chickens and pigeons. Journal of Parasitology SO: 124127. ISHII T. 1959. Studies on larva migrans. 2. Comparative studies on migratory behaviour and larval morphology of Toxocara canis and Ascaris suilla, with regard to their reinfectivity. Japanese Journal of Parasitology 8: 558-566. MI~~RA S. & SASMALN. K. 1985. Exnerimental infection of pupswith Ancylostoma caninum larvae from an abnormal host, the chicken. Journal of Helminthology 59: 303-306. NICHOLS R. L. 1956. The etioloev of visceral larva miarans. I. Diagnostic morphology of infective second-stage Tixocara larvae. Journal of Parasitology 42: 349-362. OKOSHI S. & USUI M. 1968. Experimental studies of Toxascaris leonina VI. Experimental infection of mice, chickens and earthworms with Toxascaris leonina, Toxocara canis and Toxocara cati. Japanese Journal of Veterinary Science 30: 151-166. OSHIMA T. 1961. Standardization of techniques for infecting mice with Toxocara canis and observations on the normal migration routes of the larvae. Journal of Parasitology 47: 652-656. SCHAEFFLERW. F. 1960. Experimental infection of sheep with the dog ascarid, Toxocara canis. Journal of Parasitology 46 (suppl.): 17. SPRENTJ. F. A. 1952. On the migratory behaviour of the larvae of various Ascaris species in white mice. I. Distribution of larvae in tissues. Journal of Infectious Diseases 901 165-l 76. SPRENT J. F. A. 1954. The life cycles of nematodes in the Family Ascarididae Blanchard 1896. Journal of Parasitology 40: 608-617. SPRENTJ. F. A. 1955. On the invasion of the central nervous system by nematodes II. Invasion of the nervous system in Ascariasis. Parasitology 45: 41-55.
Inkmotional Journalfor Parasitology Vol. 20, No. 2, pp. 263-264, 1990 Printed in Grmt Britain
RESEARCH NOTE
EXPERIMENTAL INFECTION OF MICE WITH TOXOCARA LARVAE OBTAINED FROM JAPANESE QUAILS
CANIS
T. K. PAHARI and N. K. SASMAL* Department
of Parasitology,
Faculty of Veterinary and Animal Sciences, Bidhan Chandra Agricultural University, Mohanpur 741252, W.B., India (Received 12 December 1989; accepted 20 December 1989)
Ahstracf-PAHAaI T. K. and SASMAL N. K. 1990. Experimental infection of mice with Toxocara canis larvae obtained from Japanese quails. International Journal for Parasitology 20: 263-264. The migration and distribution of Toxocara canis larvae in the tissues of Japanese quails, infected orally with 5 x 10’ infective eggs, were studied, as well as the re-infectivity of these larvae in mice, inoculated with 50 larvae obtained from the liver of these quails. Post-infection, the highest concentrations of larvae were found to be present in the liver of quails while only a few migrated to other tissues like lungs, heart, muscle and brain. The migration and distribution of the larvae in the tissues of mice were studied by necropsy on days 6 and 12 postinfection. On both days the highest number of larvae, 11 and 10, were recovered from the carcase followed by six and seven from the leg muscles and four and eight from the brain, respectively. A few larvae were recovered from the liver, lungs and viscera. This implies that the larvae had a special affinity for the muscle and brain tissue of mice, unlike in the quails. The role of these larvae in relation to paratenism is discussed.
INDEX KEY WORDS: Toxocaru cunis; migration; Japanese quails; mice; paratenism.
behaviour of larvae in different abnormal hosts have been performed (Sprent, 1952, 1955; Beaver, 1956; Done, EXPERIMENTALstudies on the migratory T. cunis
Richardson KcGibson, 1960; Schaeffler, 1960; Galvin, 1964) and it is presumed that larvae are transferred in nature from animal to animal through paratenism (Sprent, 1954). The experimental re-infectivity of Toxocara was reported to be comparatively higher
than any other ascarids and the infections were retained for a longer period in the abnormal hosts (Ishii, 1959; Okoshi & Usui, 1968). Several previous reports have discussed the migratory behaviour of T. cunis larvae in laboratory mammalian hosts, but none have examined this question in Japanese quails (Coturnix coturnix juponica). Accordingly the present study was undertaken to clarify this aspect and to verify the re-infectivity of these larvae to mice. Eggs were obtained by dissecting gravid T. canis females, incubated in0.5% formalin at 27°C f 1°C for 46 weeks, and washed in water to remove formalin before use. Healthy 5-6-week-old male laboratoryreared Japanese quails, weighing 17&l 80 g, and 46week-old albino mice of either sex, weighing 25-30 g were used in experiments. They were maintained in small metal cages and provided with a commercial diet and water ad lib. Strict infection-free status was maintained throughout the experiment. The calcu-
* To whom all correspondence should be addressed.
lated dose of embryonated eggs and infective larvae in water suspension was determined by the method of Oshima (1961) and the animals were dosed by stomach intubation. The quails and the mice were starved for 6 h prior to and 2 h post-infection. Twelve Japanese quails were each infected with 5 x lo3 infective eggs and were necropsied in groups of four on days lo,20 and 30 post-infection. Similarly, three uninfected control birds were also necropsied at intervals ranging from 10 to 30 days. Eight mice were each inoculated with 50 actively motile second-stage larvae isolated from the liver of Japanese quails on day 30 postinfection and were necropsied in groups of two on days 6 and 12 post-infection. Similarly, two uninfected control mice were also killed at intervals ranging from 6 to 12 days. Quails and mice were killed with chloroform. Immediately following death, the alimentary tract, liver, lungs, heart, spleen, kidney, muscle, carcase and brain were removed and rinsed in saline. Larvae were recovered from organs and tissues by digestion in pepsin as described by Sprent (1952) with some modification, and by Baermann isolation from minced fresh tissues. Larvae in the brain were counted directly by squashing the tissues between two glass slides. The larvae were identified according to Nichols (1956) and Sprent (1955). The results have been summarized in Table 1. On days 10, 20 and 30, 93-95% of all recovered larvae were in the liver of quail and very few in muscle, brain and heart. In the mice on both days 6 and 12 postinfection, the highest number of larvae (11 and 10, 263
264
T. K. PAHARI and N. K. SASMAL
TABLE I-NUMBER OF LARVAERECOVERED FROMTISSUESOF JAPANESEQUAILINOCULATED WITH5 x 10 3 INFECTIVE EGGSOF Toxocara canis No. of quails
Average
Days infected Stomach*
4 4 4
Intestine
285 296 304
10 20 30
lProventriculus
Liver
Lungs 1
larvae recovered Heart 4 5 5
from
Thigh muscle
Breast muscle
Neck muscle
3 4 5
2 4 6
5 2 4
Brain
2 4
Total larvae recovered 300 313 328
and gizzard tissues.
respectively) were recovered from the carcase followed by the leg muscle (six and seven) and brain (four and eight). In comparison to these, a few larvae were
recovered from the liver, lungs and viscera; two and none, four and two, one and none, respectively. The percentage inoculum recovered at necropsy in the first case was 56 and the second case, 54. No morphological differences were found among the larvae from Japanese quails and mice. They were all second-stage larvae. No larvae were recovered from the control groups at necropsy. The infection of mice with T. canis larvae of quail origin indicates that Japanese quail may possibly serve as a paratenic host of T. canis, like chickens and pigeons (Beaver, 1956; Galvin, 1964; Okoshi & Usui, 1968). It is evident that almost all larvae accumulated in the liver of quails. This special affinity of larvae for avian liver tissue was suggested by Galvin (1964) and Okoshi & Usui (1968) in chickens and pigeons. Migration to other tissues in pigeons has also been reported by Galvin (1964). Larvae recovered from the liver of quail were used for infection to mice. In the mice most of these larvae migrated to organs beyond the liver. In the present experiment there is some suggestion that more larvae accumulated in the brain with increasing time as reported by Sprent (1955) and Dunsmore, Thompson & Bates (1983). It is clear from the present investigation that larvae of T. canis can establish in both the abnormal hosts, quails and mice, and the larvae of quail origin retain their infectivity to mice even after day 30 post-infection. However, in respect to larval recovery the adaptability of the larvae to mammalian tissue was found to be greater than to the avian tissue which is in conformity with Okoshi & Usui (1968) and Mittra & Sasmal (1985). The larvae from both sources were morphologically identical as reported by Nichols (1956) Ishii (1959) and Okoshi & Usui (1968) in their studies on the transfer of larvae from mouse to mouse and chicken to mouse. These findings are in agreement with those of Sprent (1954) and Okoshi & Usui (1968) who reported that effective transfer of larvae from animal to animal may always be possible in the natural world by means of paratenism.
Acknowledgemenrs-The authors are grateful to the Bidhan Chandra Agricultural University for financial assistance and express their gratitude to Prof. H. L. Shah, Department of Parasitology, Jabalpur Veterinary College, for his scientific advice.
REFERENCES BEAVER P. C. 1956. Larva migrans. Experimental Parasirology 5: 587-621. DONE J. T., RICHARDSON M. D. & GIBSON T. E. 1960. Experimental visceral larva migrans in the pig. Research in Veterinary Science 23: 109-l 14. DUNSMOREJ. D., THOMPSONR. C. A. & BATESI. A. 1983. The accumulation of Toxocara canis larvae in the brains of mice. International Journal for Parasitology 13: 5 17-521. GALVINT. J. 1964. Experimental Toxocara canis infections in chickens and pigeons. Journal of Parasitology SO: 124127. ISHII T. 1959. Studies on larva migrans. 2. Comparative studies on migratory behaviour and larval morphology of Toxocara canis and Ascaris suilla, with regard to their reinfectivity. Japanese Journal of Parasitology 8: 558-566. MI~~RA S. & SASMALN. K. 1985. Exnerimental infection of pupswith Ancylostoma caninum larvae from an abnormal host, the chicken. Journal of Helminthology 59: 303-306. NICHOLS R. L. 1956. The etioloev of visceral larva miarans. I. Diagnostic morphology of infective second-stage Tixocara larvae. Journal of Parasitology 42: 349-362. OKOSHI S. & USUI M. 1968. Experimental studies of Toxascaris leonina VI. Experimental infection of mice, chickens and earthworms with Toxascaris leonina, Toxocara canis and Toxocara cati. Japanese Journal of Veterinary Science 30: 151-166. OSHIMA T. 1961. Standardization of techniques for infecting mice with Toxocara canis and observations on the normal migration routes of the larvae. Journal of Parasitology 47: 652-656. SCHAEFFLERW. F. 1960. Experimental infection of sheep with the dog ascarid, Toxocara canis. Journal of Parasitology 46 (suppl.): 17. SPRENTJ. F. A. 1952. On the migratory behaviour of the larvae of various Ascaris species in white mice. I. Distribution of larvae in tissues. Journal of Infectious Diseases 901 165-l 76. SPRENT J. F. A. 1954. The life cycles of nematodes in the Family Ascarididae Blanchard 1896. Journal of Parasitology 40: 608-617. SPRENTJ. F. A. 1955. On the invasion of the central nervous system by nematodes II. Invasion of the nervous system in Ascariasis. Parasitology 45: 41-55.
