Veterinary Parasitology, 35 (1990) 189-193 Elsevier Science Publishers B.V., Amsterdam - - Printed in The Netherlands
189
T h e E s t a b l i s h m e n t and D e v e l o p m e n t of H a e m o n c h u s c o n t o r t u s in Goats WAHAB A. R A H M A N 1 and G.H. C O L L I N S 2
'School of Biological Sciences, University Sains Malaysia, Minden, 11800 Penang (Malaysia) 2Department of Veterinary Pathology, University of Sydney, N.S. W. 2006 (Australia) (Accepted for publication 25 September 1989)
ABSTRACT Rahman, W.A. and Collins, G.H., 1990. The establishment and development of Haemonchus contortus in goats. Vet. Parasitol., 35: 189-193. Twelve goats were inoculated with 40 000 third-stage Haemonchus contortus larvae and two were killed on each of Days 4, 7, 11, 14, 18 and 21 after inoculation (DAI). The number of worms that established, and the site of development were recorded. More worms established in the fundic, than in the middle or pyloric thirds of the abomasum. Early development occurred within the mucosa; emergence into the lumen started between 7 and 11 days after infection. By 4 DAI, all worms had completed the third moult to the L4 stage. At 11 DAI the majority of the worms were adults. A mean of 13.2% of the female worms had eggs in their uteri at 18 DAI; by 21 DAI more than half of the female worms had eggs in their uteri. The development of H. contortus was essentially similar to that described in sheep.
INTRODUCTION
The establishment of H. contortus in goats has only been described briefly by Sahai and Deo (1966) and A1-Quaisy et al. (1987). Otherwise, it has been assumed that the development of the parasitic stages of H. contortus in the goat is similar to that in sheep. The present study was carried out with the aim of extending the findings of Sahai and Deo (1966) and A1-Quaisy et al. (1987) and comparing the establishment and development of H. contortus in goats with that in sheep as described by other workers. M A T E R I A L S AND M E T H O D S
Experimental animals Newly born goats were obtained from farms near Sydney, Australia. The majority were Saanens or Nubians, whilst a few were Toggenburgs or British 0304-4017/90/$03.50
© 1990 Elsevier Science Publishers B.V.
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W.A. RAHMAN AND G.H. COLLINS
Alpines. All goats were reared worm-free, on wire-mesh floors, indoors, until the age of 2-3 months. The goats were fed a milk replacer (Denkavit; Daken Corporation Pty. Ltd., New South Wales, Australia) first by bottle, later ad libitum; chaffed lucerne was introduced into the diet from 3 weeks of age.
Infective larvae Infective larvae of H. contortus were obtained from the McMaster laboratory, CSIRO, Sydney from a strain (McMaster susceptible) which had been regularly passed through sheep for several years. The larvae were less than 1 week old when used and had been stored at 4 ° C. Larvae were checked for motility before used.
Experimental procedure Twelve goats were fasted for 12 h and each inoculated with 40 000 thirdstage H. contortus larvae. Two goats were killed by barbiturate overdose on each of Days 4, 7, 11, 14, 18 and 21 after inoculation (DAI) directly into the oesophagus from a syringe attached to a short rubber tube.
Recovery of worms At necropsy, the abomasum was opened and divided into three parts (fundic, middle and pyloric ) by cutting at one-third intervals along the greater curvature. The contents and the washings from the mucosa of each part were collected into separate containers containing 10% formalin. Each part was then cut into smaller pieces and digested in an aqueous solution of 1.0% pepsin with hydrochloric acid at 37 °C for 45 min. The digested tissues were washed in a sieve (mesh size 0.0625 mm). Large pieces of tissue were discarded and the sieved material was preserved in 10% formalin. Worm counts were performed on 3 X 5% aliquots of the contents and of the digested material from each third of the abomasum. Where worm counts were low, all of the contents were examined. One hundred larvae from a mixture, comprising equal numbers of larvae from the fundic, middle and pyloric parts of both the mucosa and lumen of the abomasum, were identified according to their stage in the life cycle (Veglia, 1915). RESULTS
The number of larvae which established and developed was low in all goats (Table 1 ). More worms established in the fundic than in the middle or pyloric thirds of the abomasum (Table 2). By 4 DAI, all worms had completed the third moult to the L4 stage. At 11 DAI the majority of the worms were adults.
HA EMONCHUS CONTORTUS IN GOATS
191
TABLE 1
Total number of worms found in the abomasum of goats at various times after infection with 40 000 infective larvae of third-stage Haemonchus contortus larvae DAI
4 7 11 14 18 21
Goat
1 2 1 2 1 2 1 2 1 2 1 2
Number of worms recovered Mucosa
Lumen
1754 1259 4724 3823 2309 2439 3337 4616 663 584 469 386
0 0 0 0 959 3457 856 997 7402 6832 4657 3270
Total
1754 1259 4724 3823 3268 5896 3422 5613 8065 7416 5126 3656
Percent
Mean %
estab, (%)
estab. (n=2)
4.4 3.1 11.8 9.6 8.2 14.7 10.5 14.0 20.2 19.5 12.8 3.1
3.8 10.4 11.5 12.3 19.8 11.0
TABLE 2 Mean number (n = 2 ) of worms in the three parts of the abomasum of goats at various times after infection with 40 000 infective larvae of H a e m o n c h u s c o n t o r t u s DAI
Fundic third
Middle third
Pyloric third
4 7 11 14 18 21
1210 3344 3920 3719 4544 3183
28l 875 856 1085 2427 898
17 56 86 115 771 311
A mean of 13.2% of the female worms had eggs in their uteri at 18 DAI; by 21 DAI more than half of female worms had eggs in their uteri. DISCUSSION
There was a large variation in the percentage of H. contortus larvae that were able to establish in individual goats, but in general, the establishment was low. The larvae used for infecting the goats were obtained from a strain of H. contortus that has always been maintained in sheep and which may be less successful in establishing in goats. This possibility is supported by the results of Sahai and Deo (1966) and A1-Quaisy et al. (1987) who also obtained low recovery rates in goats infected with a sheep-derived strain of H. contortus. A
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more recent experiment has shown that a goat-derived strain of H. contortus established at a higher rate in goats (Rahman and Collins, unpublished data, 1988). The larvae established mainly in the fundic third of the abomasum; the middle and pyloric thirds contained only small numbers. As this pattern was found at all times after infection, it is likely that there is little redistribution after establishment and that little displacement of worms occurred during necropsy. In the passage of the larvae through the abomasum, the first part of the abomasum encountered is the fundic third, and this may explain the higher numbers found there. Alternatively, the surface of the abomasal mucosa is folded and most of the folds are concentrated in the fundic area, with fewer folds in the middle, and the least number of folds in the pyloric third. It is possible that larvae may only be able to establish in the mucosa if they are first trapped in a fold. The distribution of H. contortus in goats was essentially the same as reported by Sommerville (1963) and Dash (1985) for the early fourth stage in the abomasum of sheep. In sheep, larvae were numerous in the fundic area but almost completely absent from the posterior pyloric area. In contrast, for species other than H. contortus, the reported distribution has varied in different experiments. The early parasitic larval stages of Ostertagia circumcincta in sheep were found to be confined mainly to the pyloric region by Sommerville (1963) and to both the fundic and pyloric area by Dash (1985); Trichostrongylus axei were found to parasitize both the fundic and pyloric regions of the abomasum (Dash, 1985). Variation in the distribution of O. circumcincta and T. axei in the abomasum of sheep has been described by Dunsmore (1966) and Sommerville (1963). Thus, of the three species parasitizing the abomasum of both goats and sheep, it seems that H. contortus is more restricted than O. circumcincta and T. axei with regard to the regions of the mucosa which can be invaded. Migration of the late fourth and adult stages of H. contortus from the anterior towards the posterior part of the abomasum has been reported in sheep (Dash, 1985). A similar posterior migration has been noted in the later larval stages of O. circumcincta (Dash, 1985). Dash (1985) suggested that the posterior migration of the late fourth and adult stages of H. contortus in sheep indicates selection by the blood-feeding stages of a more favourable feeding site, or avoidance of a local host reaction in the area parasitized by the earlier larval stages. However, in this investigation no discernible migration occurred. By 4 DAI all the worms were at the L4 stage and the third stage larvae were totally absent; the third moult occurred earlier than 4 DAI. The worms spent between 7 to 11 DAI in the mucosa before emerging, mostly as late L4 larvae, and establishing themselves in the lumen (Table 1 ). At 18 DAI, some of the worms in the lumen were mature adults. The pre-patent period was 20 DAI. However, it should be pointed out that the minimal time to reach patency could have been as early as 15 DAI, because at 18 DAI 13.2% of the female worms already had eggs in their uteri.
HAEMONCHUS CONTORTUS IN GOATS
193
In sheep, the third moult occurs by 2 to 3 DA! (Veglia, 1915; Silverman, 1965; Coadwell and Ward, 1975 ). The fourth moult can occur as early as 6 DAI (Silverman 1965), but more usually occurs between 9 and 11 DAI (Veglia, 1915; Coadwell and Ward, 1975). Eggs are present in the faeces as early as 12 DAI (Silverman and Patterson, 1960). ACKNOWLEDGEMENTS
The authors wish to thank Mr. David Griffith and Ms. Sally Pope from the Department of Veterinary Pathology, University of Sydney, Australia, for their able technical help. The Australian Development Assistance Bureau (ADAB) is deeply acknowledged for providing a grant for the above research.
REFERENCES A1-Quaisy, H.H.K., Altaif, K.I., Al-Zubaidy, A.J. and Makkawi, T.A., 1987. The pathogenecity oi haemonchosis due to H. contortus in sheep and goats in Iran. Vet. Parasitol., 24: 221-228. Coadwell, W.J. and Ward, P.F.V., 1975. Observations on the development ofH. contortus in young sheep given a single infection. Parasitology, 71:505-515. Dash, K.M., 1985. Distribution of trichostrongylid nematodes in the abomasum of sheep. Int. J. Parasitol., 15: 505-510. Dunsmore, J.D., 1966. Topographical distribution of Ostertagia spp. in the abomasum of sheep. J. Helminthol., 40: 39-44. Sahai, B.N. and Deo, P.G., 1966. Studies on cross infection of H. contortus in sheep and goats with a note on their infectivity in suckling cow-calves and buffalo-calves Indian Vet. J., 43: 969-972. Silverman, P.H., 1965. Some immunologic aspects of parasitic helminth infection. Am. Zool., 5: 153-163. Silverman, P.H. and Patterson, J.E., 1960. Histotrophic (parasitic) stages in H. contortus. Nature (London), 185: 54-55. SommerviUe, R.I., 1963. Distribution of some parasitic nematodes in the alimentary tract of sheep, cattle and rabbits. J. Parasitol., 49: 593-599. Veglia, F., 1915. The anatomy and life-history of the H. contortus (Rud.). Report of the Director of Veterinary Research, Department of Agriculture, Union of South Africa: pp. 349-500.
189
T h e E s t a b l i s h m e n t and D e v e l o p m e n t of H a e m o n c h u s c o n t o r t u s in Goats WAHAB A. R A H M A N 1 and G.H. C O L L I N S 2
'School of Biological Sciences, University Sains Malaysia, Minden, 11800 Penang (Malaysia) 2Department of Veterinary Pathology, University of Sydney, N.S. W. 2006 (Australia) (Accepted for publication 25 September 1989)
ABSTRACT Rahman, W.A. and Collins, G.H., 1990. The establishment and development of Haemonchus contortus in goats. Vet. Parasitol., 35: 189-193. Twelve goats were inoculated with 40 000 third-stage Haemonchus contortus larvae and two were killed on each of Days 4, 7, 11, 14, 18 and 21 after inoculation (DAI). The number of worms that established, and the site of development were recorded. More worms established in the fundic, than in the middle or pyloric thirds of the abomasum. Early development occurred within the mucosa; emergence into the lumen started between 7 and 11 days after infection. By 4 DAI, all worms had completed the third moult to the L4 stage. At 11 DAI the majority of the worms were adults. A mean of 13.2% of the female worms had eggs in their uteri at 18 DAI; by 21 DAI more than half of the female worms had eggs in their uteri. The development of H. contortus was essentially similar to that described in sheep.
INTRODUCTION
The establishment of H. contortus in goats has only been described briefly by Sahai and Deo (1966) and A1-Quaisy et al. (1987). Otherwise, it has been assumed that the development of the parasitic stages of H. contortus in the goat is similar to that in sheep. The present study was carried out with the aim of extending the findings of Sahai and Deo (1966) and A1-Quaisy et al. (1987) and comparing the establishment and development of H. contortus in goats with that in sheep as described by other workers. M A T E R I A L S AND M E T H O D S
Experimental animals Newly born goats were obtained from farms near Sydney, Australia. The majority were Saanens or Nubians, whilst a few were Toggenburgs or British 0304-4017/90/$03.50
© 1990 Elsevier Science Publishers B.V.
190
W.A. RAHMAN AND G.H. COLLINS
Alpines. All goats were reared worm-free, on wire-mesh floors, indoors, until the age of 2-3 months. The goats were fed a milk replacer (Denkavit; Daken Corporation Pty. Ltd., New South Wales, Australia) first by bottle, later ad libitum; chaffed lucerne was introduced into the diet from 3 weeks of age.
Infective larvae Infective larvae of H. contortus were obtained from the McMaster laboratory, CSIRO, Sydney from a strain (McMaster susceptible) which had been regularly passed through sheep for several years. The larvae were less than 1 week old when used and had been stored at 4 ° C. Larvae were checked for motility before used.
Experimental procedure Twelve goats were fasted for 12 h and each inoculated with 40 000 thirdstage H. contortus larvae. Two goats were killed by barbiturate overdose on each of Days 4, 7, 11, 14, 18 and 21 after inoculation (DAI) directly into the oesophagus from a syringe attached to a short rubber tube.
Recovery of worms At necropsy, the abomasum was opened and divided into three parts (fundic, middle and pyloric ) by cutting at one-third intervals along the greater curvature. The contents and the washings from the mucosa of each part were collected into separate containers containing 10% formalin. Each part was then cut into smaller pieces and digested in an aqueous solution of 1.0% pepsin with hydrochloric acid at 37 °C for 45 min. The digested tissues were washed in a sieve (mesh size 0.0625 mm). Large pieces of tissue were discarded and the sieved material was preserved in 10% formalin. Worm counts were performed on 3 X 5% aliquots of the contents and of the digested material from each third of the abomasum. Where worm counts were low, all of the contents were examined. One hundred larvae from a mixture, comprising equal numbers of larvae from the fundic, middle and pyloric parts of both the mucosa and lumen of the abomasum, were identified according to their stage in the life cycle (Veglia, 1915). RESULTS
The number of larvae which established and developed was low in all goats (Table 1 ). More worms established in the fundic than in the middle or pyloric thirds of the abomasum (Table 2). By 4 DAI, all worms had completed the third moult to the L4 stage. At 11 DAI the majority of the worms were adults.
HA EMONCHUS CONTORTUS IN GOATS
191
TABLE 1
Total number of worms found in the abomasum of goats at various times after infection with 40 000 infective larvae of third-stage Haemonchus contortus larvae DAI
4 7 11 14 18 21
Goat
1 2 1 2 1 2 1 2 1 2 1 2
Number of worms recovered Mucosa
Lumen
1754 1259 4724 3823 2309 2439 3337 4616 663 584 469 386
0 0 0 0 959 3457 856 997 7402 6832 4657 3270
Total
1754 1259 4724 3823 3268 5896 3422 5613 8065 7416 5126 3656
Percent
Mean %
estab, (%)
estab. (n=2)
4.4 3.1 11.8 9.6 8.2 14.7 10.5 14.0 20.2 19.5 12.8 3.1
3.8 10.4 11.5 12.3 19.8 11.0
TABLE 2 Mean number (n = 2 ) of worms in the three parts of the abomasum of goats at various times after infection with 40 000 infective larvae of H a e m o n c h u s c o n t o r t u s DAI
Fundic third
Middle third
Pyloric third
4 7 11 14 18 21
1210 3344 3920 3719 4544 3183
28l 875 856 1085 2427 898
17 56 86 115 771 311
A mean of 13.2% of the female worms had eggs in their uteri at 18 DAI; by 21 DAI more than half of female worms had eggs in their uteri. DISCUSSION
There was a large variation in the percentage of H. contortus larvae that were able to establish in individual goats, but in general, the establishment was low. The larvae used for infecting the goats were obtained from a strain of H. contortus that has always been maintained in sheep and which may be less successful in establishing in goats. This possibility is supported by the results of Sahai and Deo (1966) and A1-Quaisy et al. (1987) who also obtained low recovery rates in goats infected with a sheep-derived strain of H. contortus. A
192
W.A. RAHMAN AND (I.H. COLLINS
more recent experiment has shown that a goat-derived strain of H. contortus established at a higher rate in goats (Rahman and Collins, unpublished data, 1988). The larvae established mainly in the fundic third of the abomasum; the middle and pyloric thirds contained only small numbers. As this pattern was found at all times after infection, it is likely that there is little redistribution after establishment and that little displacement of worms occurred during necropsy. In the passage of the larvae through the abomasum, the first part of the abomasum encountered is the fundic third, and this may explain the higher numbers found there. Alternatively, the surface of the abomasal mucosa is folded and most of the folds are concentrated in the fundic area, with fewer folds in the middle, and the least number of folds in the pyloric third. It is possible that larvae may only be able to establish in the mucosa if they are first trapped in a fold. The distribution of H. contortus in goats was essentially the same as reported by Sommerville (1963) and Dash (1985) for the early fourth stage in the abomasum of sheep. In sheep, larvae were numerous in the fundic area but almost completely absent from the posterior pyloric area. In contrast, for species other than H. contortus, the reported distribution has varied in different experiments. The early parasitic larval stages of Ostertagia circumcincta in sheep were found to be confined mainly to the pyloric region by Sommerville (1963) and to both the fundic and pyloric area by Dash (1985); Trichostrongylus axei were found to parasitize both the fundic and pyloric regions of the abomasum (Dash, 1985). Variation in the distribution of O. circumcincta and T. axei in the abomasum of sheep has been described by Dunsmore (1966) and Sommerville (1963). Thus, of the three species parasitizing the abomasum of both goats and sheep, it seems that H. contortus is more restricted than O. circumcincta and T. axei with regard to the regions of the mucosa which can be invaded. Migration of the late fourth and adult stages of H. contortus from the anterior towards the posterior part of the abomasum has been reported in sheep (Dash, 1985). A similar posterior migration has been noted in the later larval stages of O. circumcincta (Dash, 1985). Dash (1985) suggested that the posterior migration of the late fourth and adult stages of H. contortus in sheep indicates selection by the blood-feeding stages of a more favourable feeding site, or avoidance of a local host reaction in the area parasitized by the earlier larval stages. However, in this investigation no discernible migration occurred. By 4 DAI all the worms were at the L4 stage and the third stage larvae were totally absent; the third moult occurred earlier than 4 DAI. The worms spent between 7 to 11 DAI in the mucosa before emerging, mostly as late L4 larvae, and establishing themselves in the lumen (Table 1 ). At 18 DAI, some of the worms in the lumen were mature adults. The pre-patent period was 20 DAI. However, it should be pointed out that the minimal time to reach patency could have been as early as 15 DAI, because at 18 DAI 13.2% of the female worms already had eggs in their uteri.
HAEMONCHUS CONTORTUS IN GOATS
193
In sheep, the third moult occurs by 2 to 3 DA! (Veglia, 1915; Silverman, 1965; Coadwell and Ward, 1975 ). The fourth moult can occur as early as 6 DAI (Silverman 1965), but more usually occurs between 9 and 11 DAI (Veglia, 1915; Coadwell and Ward, 1975). Eggs are present in the faeces as early as 12 DAI (Silverman and Patterson, 1960). ACKNOWLEDGEMENTS
The authors wish to thank Mr. David Griffith and Ms. Sally Pope from the Department of Veterinary Pathology, University of Sydney, Australia, for their able technical help. The Australian Development Assistance Bureau (ADAB) is deeply acknowledged for providing a grant for the above research.
REFERENCES A1-Quaisy, H.H.K., Altaif, K.I., Al-Zubaidy, A.J. and Makkawi, T.A., 1987. The pathogenecity oi haemonchosis due to H. contortus in sheep and goats in Iran. Vet. Parasitol., 24: 221-228. Coadwell, W.J. and Ward, P.F.V., 1975. Observations on the development ofH. contortus in young sheep given a single infection. Parasitology, 71:505-515. Dash, K.M., 1985. Distribution of trichostrongylid nematodes in the abomasum of sheep. Int. J. Parasitol., 15: 505-510. Dunsmore, J.D., 1966. Topographical distribution of Ostertagia spp. in the abomasum of sheep. J. Helminthol., 40: 39-44. Sahai, B.N. and Deo, P.G., 1966. Studies on cross infection of H. contortus in sheep and goats with a note on their infectivity in suckling cow-calves and buffalo-calves Indian Vet. J., 43: 969-972. Silverman, P.H., 1965. Some immunologic aspects of parasitic helminth infection. Am. Zool., 5: 153-163. Silverman, P.H. and Patterson, J.E., 1960. Histotrophic (parasitic) stages in H. contortus. Nature (London), 185: 54-55. SommerviUe, R.I., 1963. Distribution of some parasitic nematodes in the alimentary tract of sheep, cattle and rabbits. J. Parasitol., 49: 593-599. Veglia, F., 1915. The anatomy and life-history of the H. contortus (Rud.). Report of the Director of Veterinary Research, Department of Agriculture, Union of South Africa: pp. 349-500.
