Multiple forms of bovine pepsinogen: isolation and identification in serum from calves with ostertagiasis

By using fast protein liquid chromatography (FPLC) a new form of bovine pepsinogen has been identified. Compared with the previously isolated pepsinogen (PG I) the new pepsinogen (PG II) is less mobile on agarose electrophoresis, is eluted earlier from an anion-exchange column and is 30 times less abundant. Agarose electrophoresis identified PG I in the serum of calves given third stage Ostertagia ostertagi larvae, while FPLC identified both PG I and PG II in such serum.     

Further studies on the response to transplanted adult Ostertagia ostertagi in calves

Calves infected by surgical transplantation of adult Ostertagia ostertagi had raised levels of plasma pepsinogen and those in which the largest number of worms established also had elevated plasma gastrin concentrations. Despite the elevated plasma pepsinogen values, the abomasal pH of the animals did not change significantly, and there was no significant difference in the percentage establishment of adult parasites in calves previously infected with O ostertagi third stage larvae and those which had been maintained parasite-naive before transplant.     

Effect of Ostertagia ostertagi on lamb performance and cross resistance to O circumcincta

Fifteen worm-free lambs (two-and-a-half to three months or four to four-and-a-half months old) were infected with 3500 or 4000 Ostertagia ostertagi larvae on five days each week for six weeks, and their performance compared to that of controls. Eleven lambs were killed after eight weeks and four were challenged with O circumcincta to determine whether any cross resistance had developed. A feature of the O ostertagi infection was the considerable variation in response. Overall liveweight gain was lowered by 24 per cent in the two-and-a-half to three-month-old infected lambs, mainly due to reductions of 27 to 40 per cent in four of the seven lambs. There was no consistent effect in the older lambs. The worm populations consisted mainly of early fourth stage larvae and developing worms, but a small percentage reached sexual maturity and these produced a low faecal egg count (1 to 63 eggs per gram). Numerous intraluminal refractive crystals were present in the gut of both adult worms and developing stages, possibly reflecting degenerative changes. Hypertrophy of the abomasal mucosa with patchy loss of differentiation was a feature of the infection, and in four lambs serum pepsinogen concentrations were elevated. Exposure to O ostertagi did confer some protection against challenge with O circumcincta in that worm counts were reduced to about 60 per cent of those in controls, although no increase was observed in the numbers of arrested larvae. The successful passage of O ostertagi through young lambs could be important in mixed or alternate grazing systems by providing a reservoir of infection for the alternate host.     

Immune modulation by Ostertagia ostertagi and the effects of diet

IgG1 antibody responses to Ostertagia ostertagi third stage larvae (L3) and the third party antigen, keyhole limpet haemocyanin (KLH), and faecal egg counts were determined in calves infected with a single dose of O. ostertagi and in uninfected, pair-fed calves. The infected and uninfected calves were given diets either high (H) or low (L) in protein and energy. The diets were within the normal range of husbandry practice in the UK. IgG1 antibody responses to L3 antigen were significantly greater from 6 weeks post-infection in infected calves given the L diet than in infected calves given the H diet (P less than 0.05). The effects of diet and infection on anti-KLH IgG1 responses were independent of each other. IgG1 responses to KLH were decreased by infection and by the L diet compared with the H diet.     

Gastrin and pepsinogen changes during an Ostertagia ostertagi challenge infection in calves

Daily changes in serum gastrin and pepsinogen concentration have been studied during two types of infection with Ostertagia ostertagi in calves. In a first experiment two calves were trickle infected (10 times 10,000 L3 Ostertagia) and two animals received a single infection of 100,000 L3 Ostertagia. Gastrin and pepsinogen changes are discussed in relation to adult wormburdens. The second experiment involved 8 calves and was designed to investigate pepsinogen and gastrin changes following a challenge infection in previously sensitized calves. The high dosed group was infected with 5,000 L3 O. ostertagi during 30 days, the low dosed group received 500 L3 O. ostertagi and group 3 served as uninfected control. At day 41 post infection all animals were treated with oxfendazole and on day 61 challenged with 100,000 L3 O. ostertagi. Only in the high dosed group a distinct pepsinogen and gastrin reaction was noticed. Both parameters dropped to almost preinfection levels after treatment. Two days post challenge a moderate rise (+/- 1,000 mU tyr) of the pepsinogen concentration was observed in the previously infected animals and gastrin showed a temporary slight increase in several animals 8 to 10 days post challenge. The effect of treatment and challenge infection is discussed in relation to gastrin and pepsinogen changes and immunity.     

Concurrent daily infection of calves with Fasciola hepatica and Ostertagia ostertagi.

Three groups of calves were infected daily with either 1500 Ostertagia ostertagi larvae, 20 Fasciola hepatica metacercariae, or 1500 O ostertagi plus 20 F hepatica metacercariae. Weekly measurements were taken of calf weight, faecal egg output, plasma concentrations of albumin, plasma activities of sorbitol dehydrogenase, gamma glutamyl transpeptidase and pepsinogen and standard haematological indices. Calves were killed either 10 or 21 weeks after daily infections began. F hepatica infection had little influence on the size and structure of the O ostertagi worm population or vice versa. Mean worm burdens found at 20 weeks in those animals infected with both F hepatica and O ostertagi were 293 flukes and 20,641 nematodes. While this level of infection is similar to that seen in the disease complex in the field, there was no evidence of clinical disease or any difference in weight gain between the groups in this experiment. Factors other than additive worm burdens are obviously important for the expression of disease under field conditions.     

Synergistic influence of Ostertagia ostertagi and Trichostrongylus axei on Ostertagia ostertagi larval inhibition and abomasal lesions in cattle

Parasite-free 4-month-old calves were inoculated with Ostertagia ostertagi and/or Trichostrongylus axei followed 6 weeks later by increasing doses of O ostertagi for 8 weeks. Clinical signs of parasitism, fecal egg counts, and plasma pepsinogen concentrations were monitored, and gross lesions and parasite burdens were determined postmortem. Clinical signs of parasitism were not observed and weight gains were not affected in experimentally infected calves. In calves infected with O ostertagi, mean plasma pepsinogen concentrations were greater than for control calves and were diagnostically significant 4 weeks after inoculation and during the last 4 weeks of serial inoculations with O ostertagi. In calves that were given O ostertagi and T axei, abomasal pH was significantly increased, and abomasal lesions were more pronounced than in control calves or in calves inoculated with only O ostertagi or T axei. Abomasal lymph nodes were enlarged in all parasitized calves; other lymph nodes in the calves inoculated with both O ostertagi and T axei were usually smaller than in calves inoculated with only O ostertagi or T axei. Numbers of O ostertagi-inhibited larvae were small in all inoculated calves, but the percentage inhibition was significantly greater in calves inoculated with both O ostertagi and T axei. The percentage inhibition was 3.53% for the O ostertagi-inoculated calves and 7.07% for calves inoculated with both O ostertagi and T axei. These percentages indicated a synergistic effect of concurrent abomasal parasitism, whereas a synergistic effect on T axei worm burden was not observed. The low percentage of larval inhibition indicated that factors other than host resistance are involved in naturally occurring pretype II ostertagiosis.     

Effects of Ostertagia ostertagi infection on secretion of metabolic hormones in calves.

Effects of Ostertagia ostertagi infection on secretion of insulin, pancreatic glucagon, cortisol, gastrin, and pepsinogen were studied in calves inoculated with 100,000 (group 1) or 10,000 (group 2) O ostertagi infective larvae weekly for 14 weeks. Plasma insulin concentrations in both inoculated groups were lower than those in a non-infected (group 3) control group. The differences between group 1 and group 3 were significant (P < 0.05) at 2 and 12 weeks after initial inoculation. Plasma pancreatic glucagon and cortisol concentrations of groups 1 and 2 did not differ significantly from those of the control group, although plasma pancreatic glucagon concentration was consistently lower in group-1 calves from 4 weeks to end of the study. Plasma pepsinogen and serum gastrin concentrations also increased significantly (P < 0.05) in both groups that received inoculations. We concluded that decreased plasma insulin concentrations are contributory to changes in postabsorptive protein metabolism, and that serum gastrin concentrations are more representative of the pathologic changes in the abomasum than are plasma pepsinogen concentrations.     

Persistence of dead Ostertagia ostertagi in the abomasal mucosa following anthelmintic treatment

Anthelmintic trails, conducted with albendazole, fenbendazole and ivermectin for efficacy against gastrointestinal nematodes, principally inhibited early fourth larval stages of Ostertagia ostertagi in naturally infected cattle. Cattle wee slaughtered seven to 20 days after treatment. O ostertagi was the predominant abomasal nematode recovered with occasional small numbers of Haemonchus species and Trichostrongylus axei. Control calves uniformly had very large O ostertagi infections, primarily early fourth stage larvae. Viable surviving worms and variable numbers of dead and degenerate worms were recovered in abomasal contents and washings. These O ostertagi larvae and adults were characterised by adherent debris or proteinaceous material, degenerated cuticles and distortion of internal structures. This study demonstrated the necessity for proper timing of slaughter for anthelmintic trial evaluation to allow clearance of dead nematodes, specifically O ostertagi larvae which are sequestered in the abomasal glandular tissue. Nematode collection within seven to 12 days after treatment will include dead and degenerate larval nematodes. The peripheral coating of larvae was suggestive of the Splendore-Hoeppli effect which has been associated with immunological responsiveness. The antigenic stimulus for this material and the lymphocyte and eosinophil infiltration was suspected to be early fourth stage O ostertagi larvae within the mucosa but was not identified definitively.     

Pathogenesis of simulated natural infections with Ostertagia ostertagi in calves

The possibility of a mucosal hypersensitivity reaction and its relationship to the pathogenesis of simulated natural infections with Ostertagia ostertagi were studied in calves. Four groups of 4 calves each were used. One group was used as noninfected control; a 2nd group was given increasing doses of infective larvae; a 3rd group was given increasing doses of larvae and these were removed by succeeding treatment with an anthelmintic; and a 4th group was given an initial dose of larvae which was then eliminated with an anthelmintic. All calves given larvae became sensitized, as shown by an intradermal skin test. The continuously infected calves had significantly (P less than 0.05) higher fecal egg counts, eosinophil counts, plasma pepsinogen values, and worm burdens and significantly (P less than 0.05) lower lymphocyte counts than did the other groups of calves. These animals also had the most extensive mucosal pathologic changes. The group given intermittent larval challenge exposures followed by an anthelmintic showed decreased lymphocyte values, but these were not significant. Plasma pepsinogen values of this group increased between every challenge exposure and treatment, a 3-day period. This indicated that a mucosal hypersensitivity reaction had occurred in these calves at these times, because they were shown to have been sensitized, and challenge-exposure infections were not present for sufficient time to have produced direct pathologic effects. It therefore seems that a part of the pathologic changes in O ostertagi infections may be the result of the continuous challenge exposure experienced by the animals through a constant intake of larvae from pasture and the intestinal reaction to this challenge exposure.     

Modulation of calf immune responses by Ostertagia ostertagi: the effect of diet during trickle infection.

The effects of Ostertagia ostertagi infection and diet on antibody responses to O. ostertagi third stage larval (L3) antigen and to an unrelated antigen, Keyhole Limpet Haemocyanin (KLH) were determined in calves experimentally infected with 3000 L3 on alternate days for 6 weeks. Calves were given one of two diets, and were either infected or not infected with O. ostertagi L3. The diets were either high (H) or low (L) in protein/energy and were within the range of normal husbandry practice in the UK. Both IgG1 and IgG2, but not IgA, responses to L3 antigen were increased in the L-diet compared with the H-diet. IgA responses to L3 antigen were not affected by dietary treatment. The effects of diet and infection on anti-KLH IgG1 were independent of each other; IgG1 anti-KLH responses were decreased by infection and by the L-diet compared with the H-diet. The data suggest that there is a strong interrelationship between diet and immunity during nematode infections.     

The immune response of sheep to larval challenge with Ostertagia circumcincta and O. ostertagi.

One of three groups of sheep was challenged twice-weekly with infective-stage larvae (L3) of the sheep parasite O. circumcincta, another with the cattle parasite O. ostertagi while the third received no larval challenge. Positive faecal egg counts (FEC) and a rise in plasma pepsinogen levels were observed only in those animals given O. circumcincta. Anti-O. circumcincta L3 IgG titres were rapidly elevated during parasite challenge with either O. circumcincta or O. ostertagi. Throughout the experiment, no rise in anti-adult IgG titres or eosinophil numbers was observed in peripheral blood in any group. On evidence of self-cure of the trickle-infection, determined by a reduction in FEC, all groups were drenched and challenged with 15,000 O. circumcincta L3. No effect of previous challenge on parasite establishment or FEC was observed, although egg viability was significantly reduced in both groups given prior challenge. Significant differences in adult female worm length were observed between groups. Those recovered from animals previously challenged with O. circumcincta were shorter than from those given O. ostertagi which were in turn shorter than those from previously unchallenged animals. In utero egg counts were significantly lower in worms from animals previously challenged with O. circumcincta than in those from unchallenged control animals. The results indicate that a level of immunity to O. circumcincta can be conferred by exposure to O. ostertagi.     

Efficacy of ivermectin against inhibited larvae of Ostertagia ostertagi

The efficacy of ivermectin against inhibited early 4th-stage larvae of ostertagia ostertagi and other nematodes of the abomasum and intestinal tract was determined in naturally infected yearling beef cattle. The time when large numbers of inhibited larvae were acquired was determined by monthly slaughter of monitor cattle, beginning in January. In April, 12 animals were removed from pasture and maintained free of further helminth exposure until slaughter (21 days). At 9 days after the cattle were removed from pasture, ivermectin was administered to the principals by subcutaneous injection (200 micrograms/kg); the other 6 animals were given subcutaneous injections of the ivermectin vehicle. both groups were klled and necropsied at 12 days after treatment. Mean numbers of O ostertagi in the 6 controls were: adults, 41,906; developing 4th stage, 73,813; and early 4th stage, 334,965. The mean proportion of early 4th-stage larvae was 73.7%. In the 6 principals (treated with ivermectin), the following reductions were observed: O ostertagi adults, 100%; developing 4th stage, 99.8%; and early 4th stage, 99,9%. Small numbers of dead and degenerated O ostertagi of all developmental stages were recovered from abomasal washings before fixation; few viable worms were recovered.     

Efficacy of doramectin against naturally acquired adult and inhibited larval infections by some nematode parasites in cattle in New Zealand

A study was conducted to determine the therapeutic efficacy of 1% doramectin injected subcutaneously at 200 microg/kg into cattle harbouring naturally acquired infections of inhibited Ostertagia ostertagi larvae. Sixteen yearling Friesian bulls, grazed without anthelmintic treatment throughout the autumn-winter, were selected on the basis of similar body weights and serum pepsinogen activities. After removal from pasture on day -23 they were weighed and randomly assigned to two treatment groups on the basis of this weight. On day 0, one group was given saline (1 ml/50 kg) while the second was treated with doramectin (200 microg/kg). Both treatments were given by subcutaneous injection. All stock were slaughtered 14-15 days after treatment. Moderate to high levels of adult O. ostertagi and Trichostrongylus axei and early and late 4th larval stages of O. ostertagi were recovered from saline-treated calves at necropsy. Doramectin was highly effective in eliminating all stages of O. ostertagi (99.9%; p<0.0001) and T. axei (100%; p<0.0001). No evidence of lesions were detected at the injection sites at necropsy. These results confirm that doramectin is an extremely effective broad-spectrum avermectin anthelmintic with efficacy against inhibited as well as maturing larval and adult forms of O. ostertagi.     

Efficacy of fenbendazole against inhibited larvae of Ostertagia ostertagi in yearling cattle

Efficacy of fenbendazole, at doses of 7.5 and 10.0 mg/kg of body weight, against inhibited early 4th-stage larvae of Ostertagia ostertagi and other nematodes of the abomasum and intestinal tract, was investigated in naturally infected yearling heifers in late May 1982. In Louisiana, this is near the end of the period (March to May) in which maximal numbers of inhibition-prone larvae are acquired. The mean numbers of O ostertagi in 10 untreated control cattle were: adults, 4,880; developing 4th-stage larvae, 12,546; and inhibited early 4th-stage larvae, 167,931. At the 7.5 mg/kg dose level (10% liquid suspension) in 10 cattle, percentage reduction of O ostertagi in comparison with controls was: adults, 95.7%; developing 4th stages, 91.1%; and inhibited 4th stage, 55.0%. Percentage reductions of other genera were as follows: abomasum--Trichostrongylus axei, 99.6%; Haemonchus sp, 95.1%; intestinal tract--Cooperia spp, 97.8%; Trichostrongylus colubriformis, 100.0%; and Oesophagostomum radiatum 4th stage and adults, 100.0%. At the 10.0 mg/kg dose (10% liquid suspension) in 11 cattle, the percentage reduction of O ostertagi in comparison with controls was: adults, 98.6%; developing 4th stages, 92.9%; and inhibited 4th stage, 80.0%. Percentage reductions of other genera were: abomasum--T axei, 99.9%; Haemonchus sp, 98.8%; intestinal tract--Cooperia spp, 99.3%; T colubriformis, 100.0%; and Oes radiatum 4th stage and adults, 100.0%. Variability of efficacy against inhibited larvae was observed, particularly at the 7.5 mg/kg dose; at this dose, 7 of the 10 heifers in the group yielded in excess of 54,000 surviving larvae.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serum pepsinogen levels and Ostertagia ostertagi populations in clinically normal adult beef cattle

Serum pepsinogen levels and Ostertagia ostertagi populations in clinically normal grass-fed bullocks were investigated in three groups of 10 prime cattle aged between 2.5 and 2.75 years slaughtered in late summer (February), early autumn (March) and late autumn (May) respectively. Apart from occasional foci of mucosal hyperplasia abomasa were grossly normal. Serum pepsinogen levels ranged between 0.2 and 2.5 i.u./l with group means of 1.4, l.5 and 1.3 i.u./l. O. ostertagi counts ranged between 0 and 5,194 with group means of 734,630 and 701 worms. The composition of the worm populations varied with a higher proportion of adults recovered in February and very few worms from most cattle in March, suggesting the termination of a parasite generation. An increase in numbers of early fourth-stage larvae in May indicated exposure to a new generation. These changes were not reflected in the pepsinogen levels. The findings are discussed in relation to the adequacy of the pepsinogen assay as a diagnostic aid in field infections, animal age, and correlations between pepsinogen levels and parasite populations.     

Experimentally induced ostertagiosis in rabbits inoculated with Ostertagia ostertagi of bovine origin

Weanling specific-pathogen-free rabbits were orally inoculated with 50,000 Ostertagia ostertagi 3rd-stage infective larvae cultured from bovine feces and were killed 42 days after inoculation. Two preliminary trials were done, using conventionally reared weanling rabbits inoculated with 2,500, 5,000, and 50,000 O ostertagi 3rd-stage larvae and dexamathasone in 1 of the trials; the rabbits were killed 14 and 28 days after inoculation. Fecal egg counts were monitored, and worm burdens were determined. The pH of the gastric contents was measured at necropsy. Recoveries of the parasite from the gastric mucosa comprised early 4th-stage larvae and larvae that had developed beyond the early 4th stage. The maximum worm burden established at 42 days after inoculation was 1,668 immature nematodes or 3.34% of the inoculum. Gross and microscopic gastric lesions were present. Gross nodular mucosal elevations, which tended to be confluent in the cardiac region, were observed. Microscopic changes were principally mucosal hyperplasia, focal lymphocyte accumulation with moderate glandular dilation, and slight variable eosinophil and plasma cell accumulation. The pathologic changes had characteristics similar to those in cattle infected with O ostertagi and other conditions characterized by gastric mucosal hyperplasia.     

Studies of the immunomodulatory effects of low-level infection with Ostertagia ostertagi in calves

Possible immunomodulation by low-level infection with Ostertagia ostertagi was studied in 4-month-old calves. Six groups of 4 calves each were subjected to the following regimens: group 1--nonparasitized controls; group 2--nonparasitized, but challenge exposed at day 64 with Brucella abortus strain 19 vaccine (BA) and at day 78 with IV administration of a soluble third-stage larval (L3) antigen preparation of O ostertagi (OAG); group 3--nonparasitized, but challenge exposed at day 78 with 75 x 10(3) L3 of O ostertagi; group 4--continuously parasitized by weekly dosing with 30 x 10(3) L3 of O ostertagi; group 5--continuously parasitized by weekly dosing with 30 x 10(3) L3 of O ostertagi, then challenge exposed on day 64 with BA and on day 78 with IV inoculation of OAG; and group 6--continuously parasitized by weekly dosing with 30 x 10(3) L3 of O ostertagi, then challenge exposed on day 78 with 75 x 10(3) L3 of O ostertagi. Over the initial 10 weeks of the study, nonparasitized calves, (groups 1, 2, and 3) had higher body weight, blood lymphocyte (BL) response to phytohemagglutinin (PHA), and significantly (P less than 0.05) higher feed consumption and lymphocyte numbers, whereas parasitized calves (groups 4, 5, and 6) had higher BL responses to pokeweed mitogen (PWM) and significantly (P less than 0.05) higher neutrophil and eosinophil numbers, plasma pepsinogen (PP) values, and BL response to OAG.(ABSTRACT TRUNCATED AT 250 WORDS)     

Blood gastrin and pepsinogen responses to subclinical infection with Ostertagia ostertagi in adult dairy cattle

Blood gastrin and pepsinogen responses to a single infection with 100,000 Ostertagia ostertagi infective larvae in lactating dairy cows were investigated. None of the infected cows showed signs of clinical ostertagiasis, nor was there any difference in live weight gain, milk yield or faecal egg count between groups. Pepsinogen levels of the infected group were significantly elevated between days 3 and 24 after infection (peak 1041 mU tyrosine; day 14). In contrast, there was no significant difference in blood gastrin levels between infected and control animals suggesting that few adult worms had become established in the former group. These data are compared with the increases in both gastrin and pepsinogen levels recorded in susceptible calves exposed to the same level, pattern and strain of ostertagia infection in a previous experiment. It is suggested that gastrin assay may be of value in adult cattle for indicating when elevated pepsinogen levels are merely associated with a rise in larval intake and not with the establishment of large adult worm burdens.     

Antigenic differences between the life cycle stages of Cooperia oncophora

The antigenic differences among the life cycle stages of Cooperia oncophora were studied by SDS-gel electrophoresis and Western blotting. Sodium dodecylsulphate polyacrylamide gel electrophoresis of the different life cycle stages of C oncophora revealed a complex protein pattern with a decreasing number of protein bands towards the adult stages. Several bands of the fourth stage larvae were in common with both the third stage and the adult nematode. Western blotting with sera from C oncophora monoinfected calves showed that the antigens of the fourth stage larvae were recognised predominantly and the presence of stage specific antigens in all stages. Strong cross reactivity was demonstrated when serum from Ostertagia ostertagi infected calves was used.