Resistance dynamics of calves to infection with Trichostrongylus axei

Veterinary Parasitology, 4 (1978) 153--160 153 © Elsevier Scientific Publishing Company, Amsterdam -- Printed in The Netherlands

RESISTANCE DYNAMICS OF CALVES TO INFECTION WITH TRICHOSTRONGYLUS A X E I

H. HERLICH

Agricultural Research Services, Beltsville Agricultural Research Center, Beltsville, Md. 20705 (U.S.A.) (Received 7 September 1977)

ABSTRACT Herlich, H., 1978. Resistance dynamics of calves to infection with Trichostrongylus axei. Vet. Parasitol., 4: 153--160. Calves were immunized with 10 000 infective larvae each of Haemonchus contortus, Ostertagia ostertagi, and Trichostrongylus axei and challenged with 100 000 of each species. When compared with nonimmunized controls, the immunized calves had 95% fewer H. contortus, 10% fewer O. ostertagi, and 20% fewer T. axei; the latter two reductions were not significant. In four other experiments single or multiple immunizing inoculations of T. axei followed by challenge at various intervals after immunization failed to result in significant resistance as measured by the number of worms of the challenge inoculation that became established, by post-challenge fecal egg count, and by gross pathologic alterations of abomasa at necropsy.

INTRODUCTION

Three species of abomasal parasites, Haemonchus placei, Ostertagia ostertagi, and Trichostrongylus axei, may cause severe disease in cattle. Extensive studies have been made of the ability of calves to develop resistance to infection with H. placei and O. ostertagi under natural and experimental conditions. Little or no resistance develops against O. ostertagi (Michel, 1953; Anderson et al., 1967; Herlich, 1976) except when calves are exposed to low level daffy doses of infective larvae for at least 180 days (Michel et al., 1973). On the other hand, either a single moderate (50 000 larvae) exposure or short term repeated low level exposure of calves to H. placei produced strong resistance to reinfection (Roberts et al., 1952; Roberts, 1957). A single oral exposure to 10 000 infective T. axei larvae resulted in a 50% reduction of a challenge infection in a single unreplicated experiment with very few calves (Ross, 1970). It seemed appropriate to investigate in greater detail the ability of calves to acquire resistance to T. axei infection.

154

MATERIALS AND METHODS

Handling and maintenance of calves, preparation and administration of infective larvae, fecal sampling and examination, necropsy procedures, and worm population determinations were as previously described (Herlich, 1976). One change in procedure was that the abomasa were exposed to pepsin--HC1 digestion fluid (Herlich, 1956) at 38°C for 6 h instead of 16 h. In five experiments, some of the "immunizing" and challenge regimens used in the cited studies on H. placei and O. ostertagi were replicated. In experiment 1, the three abomasal parasite species were used; in experiments 2--5, only T. axei was used. Specific details of immunizing and challenge exposure, intervals between exposures, and time of necropsy are shown in Tables I--V. In experiment 5, the effect of chemically abbreviated immunizing inoculations was compared with that of nonabbreviated immunizing inoculations. The immunizing inoculations were ended on day 10 by administration of the anthelmintic levamisole at 8 mg/kg. The strains of parasites used were: H. contortus, the BPL ovine strain continuously passaged through sheep; O. ostertagia, the OOH strain continuously passaged through cattle; and T. axei, the BAA strain isolated from cattle and maintained by alternate passage through cattle and sheep. The BPL strain of H. contortus was used as it is readily established in calves (Herlich, 1973) and because it is maintained at the Institute. All differences in worm burdens were analyzed for significance by Student's " t " test. Wherever the terms "immunizing" and " i m m u n i z e d " calves are used in this paper, they are intended to describe the inoculations and the experimental calves prior to administration of challenge inoculations, respectively. "Challenged-controls" is used to describe all calves n o t immunized and challenged concurrently with the immunized calves. Wherever EPG (eggs per gram of feces) is used it is given as the average for the calf group. RESULTS

Experiment 1 (Table I). -- All calves of group A (immunized) developed patent infections after their immunizing inoculations. EPG peaked at 312 on day 36 and was still 226 at necropsy on day 57. None of group B (challenged-controls) had patent infections; all animals were killed 13 days after challenge, an interval less than the shortest prepatent period for the three parasite species. Consequently, populations of adult worms were regarded as arising from the immunizing inoculations, and all other stages were considered a result of the challenge. On the basis of numbers of worms of challenge inoculation origin, group A calves had 95% fewer H. contortus, 10% fewer O. ostertagi, and 20% fewer T. axei, than group B calves. Only the difference in numbers of H. contortus is significant (P < 0.001).

155 TABLE I Treatments and worm burdens in calf groups* of experiment 1 Schedule of treatments (Days)

0

44

57 Species

H. contortus O. ostertagi T. axei

Protocols Group A immunized-challenged

Group B challenged-control

Single dose of 10000 L3 each of H. contortus, O. ostertagi, T. axei

NIL

Challenge single dose of 100000 each of three species

As for group A

¢

Necropsy all calves

Worms recovered Adult**

Immature

Adult

Immature

720 3900 6500

180 9500 10100

0 0 0

3800 10500 12600

*Five calves per group. **Adult = sexually mature; all other stages recorded as immature.

Experiment 2 (Table II). -- All calves of groups C (immunized-challenged) and D (immunized-not challenged) developed patent infections after their immunizing inoculations. When the calves were challenged on day 27, EPG was 170 for group C and 204 for group D. On day 48 (21 days post-challenge), EPG was 600 for group C, 194 for group D, and 240 for group E (challengedcontrol); however, the EPG was virtually equal in the three groups by necropsy on day 56. Group C had significantly (P < 0.01) more worms than groups D and E. However, if the number of worms in group D is considered a reflection of the residuum of the immunizing inoculation in group C, the difference in worm numbers between immunized-challenged (group C) and challengedcontrol (group E) is not significant.

Experiment 3 (Table III). -- All calves of groups F (immunized-challenged) and G (immunized-not challenged) developed patent infections after their immunizing inoculations; when the calves were challenged on day 58, EPG was 420 for group F and 388 for group G. At necropsy on day 87, the EPG was 764 for group F, 378 for group G, and 602 for group H (challengedcontrol). Group F had significantly (P < 0.05) more worms than groups G and H. However, if the number of worms in group G is considered a reflection of the residuum of the immunizing inoculations in group F, the differ-

~56

TABLE II Treatments and T. a x e i numbers in calf groups* of experiment 2 Schedule of treatments

(Days) 0 27 56

Protocols Group C immunized-challenged

Group D immuni~.ed-not challenged

Group E challenged-control

50000 L3 single dose 50000 L~ single dose

50000 L3 single dose NIL

NIL

~

50000 Lj

N e c r o p s y all calves

Worms recovered per calf

Mean

34400 43600 40400 37600

25200 18600 12500 10800

24000 24300 16200 12200

39000

16775

19175

*Four calves per group. TABLE III Treatments and T. axei numbers with calf groups* of experiment 3 Schedule of treatments

(Days) 0--24 58 87

Protocols Group F immunized-challenged

Group G immunized-not challenged

C~oup H challenged-control

10000 L3 every 72 h 150000 L3

10000 L3 every 72 h NIL

NIL 150000 L~

~

N e c r o p s y all calves

Worms recovered per calf

Mean

126100 138100 119400 131300

59600 38800 42100 23900

96600 86000 71300 94600

128725

41100

87125

*Four calves per group.

157 ence in worm numbers between group F (immunized-challenged) and group H (challenged-control) is n o t significant.

Experiment 4 (Table IV). -- All calves of groups J to O developed patent infections after their immunizing inoculations. The course of infections in respect to EPG is depicted in Fig. 1. EPG of immunized-challenged groups (J, L, and N) rose after challenge inoculations and at necropsy appeared to be still rising in challenged-controls and immunized-challenged groups, except in group N which showed a drop. EPG of immunized-not challenged groups (K, M, and O) remained at a rather constant low level throughout the experiment. One calf in each immunized challenged group and in the challenged-control group died during the 4th week post-challenge. Groups J, L, and N (immunized-challenged) had either slightly more or slightly less T. axei than group P (challenged-controls). However, if the numbers of worms in groups K, M, and O (immunized-not challenged) are considered a reflection of the residuum of the immunizing inoculations of groups J, L, and N, only group L had significantly (P<0.05) fewer (35%) T. axei than group P. Experiment 5 (Table V). -- Levamisole was apparently very effective against 10-day old T. axei in group Q. A few eggs ( < 6 EPG) appeared sporadically throughout the immunizing period, and four of five calves were negative for eggs at time of challenge inoculation. Group R calves (immunized-not anthelmintic dosed) had an EPG of 314 at time of challenge. At necropsy on day 103, EPG were: group Q - 1840; group R -- 1820, and group S (challengedcontrols) -- 1560. The three groups had substantial worm burdens; group Q had significantly (P < 0.05) fewer worms than groups R (44%) and S (26%), and group 1~ had significantly (P < 0.05) more (30%) worms than group S. Experiments 2--5. In all experiments and in all groups, 98% or more of all worms recovered at necropsy were sexually mature adults. Differences between groups in the numbers of immature (fourth-stage larvae) worms were not significant. At necropsy, within experiments, there were no gross differences among the various treatment groups in the degree of congestion and extent and severity of lesions in the abomasa of the calves. DISCUSSION The results of experiment 1 show that the calves used for this investigation were able to develop resistance to challenge infections (reinfections). The reduction of H. contortus in the immunized calf group was 95%. This finding agrees with the level of protection afforded to calves against H. placei (Roberts, 1957). Failure of the immunizing regimens to protect against O. ostertagi was n o t unexpected because others (Michel, 1953; Herlich, 1976) have amply shown that calves do n o t readily develop resistance to that parasite. The insignificant reduction (20%) in numbers of T. axei does n o t

Mean

Group K immunizednot challenged

Group L immunizedchallenged

Group M immunizednot challenged

149400

155400 147200** 145600 15266

20700 12100 13100

Worms recovered per calf

105933

97400 117400 103000** 9200

12600 6200 8800

1000 L3/Day 6000 L3 one dose/week; 5 weeks Days 0--30 Days 0, 7, 14, 21, 28 100000 L3 NIL 100000 L3 NIL 100000 L 3 NIL 100000 L3 NIL 100000 L 3 NIL 100000 L3 NIL < Necropsy all survivors; one calf* * in groups J, L, N,

Group J immunizedchallenged

Protocols

*Three calves per group

43 44 45 85

0--30

Schedule of treatments (Days)

Treatment and T. axei in calf groups* of experiment 4

TABLE IV

Group O immunizednot challenged

124800

73200** 162400 138800

12866

9800 17800 11000

30000 L3 single dose Day 0 100000 L3 NIL 100000 L 3 NIL 100000 L 3 NIL and P died during days 65--68

Group N immunizedchallenged

147466

129400 166800** 146200

NIL 100000 L3 100000 L 3 100000 L 3 )

Group P challengedcontrol

159

24-

A

-24

;

B

t

i

,i

C

I t I

18-

t I ! -18

0 )¢

0

12-

L idu

6-

! 0

42

5'3 8'5

0

42 53

85

0

42

53

85

TIMe (DAYS)

Fig. 1. Fecal egg counts of calf groups, experiment 4. T. axei immunization regimen: A -- 1000/day for 30 days; B -- 6000/week for 5 weeks; C -- 30000 single dose. Legend: immunized-challenged group ; immunized-not challenged group . . . . . . ; challengedcontrol group +++++. Challenge inoculations on days 43, 44, and 45 (~) for groups A, B, and C.

TABLE V Treatments and To axei in calf groups* of experiment 5 Schedule of treatments (Days)

Protocols Group Q immunized-challenged anthelmintic dosed

Group R immunized-challenged not anthelmintic dosed

Group S challenged-control

0 10 14 24 28 38 58 103

50000 L3 Levamisole (8 mg/kg) 50000 L3 Levamisole (8 mg/kg) 50000 L~ Levamisole (8 mg/kg) ( ~.

50000 L3 NIL 50000 L3 NIL 50000 L3 NIL 185000 L3 all calves Necropsy all calves

NIL NIL NIL NIL NIL NIL )

70600 50300 37500 44400 61200

100500 88400 101800 88400 91800

72700 70800 63800 64000 84600

52800

94180

71180

Worms recovered per calf

Mean

*Five calves per group.

160 agree w i t h R o s s ' ( 1 9 7 0 ) finding o f 50% p r o t e c t i o n . T h e d i f f e r e n c e in results b e t w e e n R o s s ' a n d t h e p r e s e n t s t u d y m a y involve v a r i a t i o n s in i m m u n o g e n i c i t y o f t h e T. axei strains used. R o s s u s e d a strain o b t a i n e d f r o m Dr. T. G i b s o n , w h i c h suggests a strain o f o v i n e origin; w h e r e a s , a b o v i n e isolate was u s e d in the present study. T h e results o f e x p e r i m e n t s 2 - - 5 i n d i c a t e t h a t i n o c u l a t i o n o f calves w i t h T. axei b y a v a r i e t y o f s c h e d u l e d d o s e s a n d t i m e s d o e s n o t p r o d u c e a high degree o f r e s i s t a n c e t o challenge i n f e c t i o n w i t h t h a t species. T h u s , t h e resist a n c e r e s p o n s e - - m o r e p r e c i s e l y , t h e general l a c k o f resistance - - is similar, if n o t identical, t o t h e s i t u a t i o n w i t h O. ostertagi. O n l y in e x p e r i m e n t s 4 a n d 5 was t h e r e d u c t i o n in an i m m u n i z e d g r o u p statistically significant even so, in e x p e r i m e n t 4, 32% o f t h e larvae in t h e challenge i n o c u l u m h a d b e c o m e established as a d u l t w o r m s as c o m p a r e d w i t h 49% in t h e c o n t r o l s . In e x p e r i m e n t 5, 29% o f t h e larvae in t h e challenge i n o c u l u m b e c a m e e s t a b l i s h e d as a d u l t w o r m s in t h e c h e m i c a l l y a b b r e v i a t e d i m m u n i z e d g r o u p as c o m p a r e d w i t h 38% in t h e c o n t r o l s . In a d d i t i o n t o a b s e n c e o f r e s i s t a n c e t o T. axei as m e a s u r e d b y w o r m b u r d e n s resulting f r o m challenge i n o c u l a t i o n s , o t h e r criteria f r e q u e n t l y u s e d t o m e a s u r e r e s i s t a n c e w e r e negative. T r i c h o s t o n g y l u s axei egg p r o d u c t i o n was n o t d r a m a t i c a l l y o r c o n s i s t e n t l y r e d u c e d a m o n g a n y o f t h e i m m u n i z e d calf groups. Similarly, gross p a t h o l o g i c a l t e r a t i o n s , c h a r a c t e r i z e d b y c o n g e s t i o n , e d e m a , a n d e r o s i o n , o f t h e calves' a b o m a s a at n e c r o p s y w e r e c o m p a r a b l e f o r i m m u n i z e d a n d c o n t r o l g r o u p s w{thin e x p e r i m e n t s .

REFERENCES

Anderson, N., Armour, J., Jarrett, W.F.H., Jennings, F.W., Ritchie, J.S.D. and Urquhart, G.M., 1967. Experimental infections of Ostertagia ostertagi in calves: Results of two regimens of multiple inoculations. Am. J. Vet. Res., 28: 1073--1077. Herlich, H., 1956. A digestion method for post-mortem recovery of nematodes from ruminants. Proc. Helminthol. Soc. Wash., 23: 102--103. Herlich, H., 1973. Effect of thiabendazole on a drug-resistant ovine isolate of Haemonchus contortus in cattle. Proc. Helminthol. Soc. Wash., 40: 165. Herlich, H., 1976. Attempts to produce protection against Ostertagia ostertagi in cattle. Am. J. Vet. Res., 37: 61--64. Michel, J.F., 1953. The phenomena of host resistance and the course of infection of Ostertagia o.stertagi in calves. Parasitology, 53: 63--84. Michel, J.F., Lancaster, M.B. and Hong, C., 1973. Ostertagia ostertagi: Protective immunity in calves. The development in calves of a protective immunity to infection with Ostertagia ostertagi. Exp. Parasitol., 33: 179--186. Roberts, F.H.S., 1957. Reactions of calves to infestation with the stomach worm, Haemonchus placei (Place, 1893) Random, 1911. Aust. J. Agric. Res., 8: 740--767. Roberts, F.H.S., O'Sullivan, P.J.O. and Riek, R.F., 1952. The epidemiology of parasitic gastro-enteritis of cattle. Aust. J. Agric. Res., 3: 187--226. Ross, J.G., 1970. Investigations of acquired immunity to T. axei infections in calves. Br. Vet. J., 126: 21--23.