The use of detector dogs in the diagnosis of nematode infections in sheep feces

This study was conducted to assess a dog's ability to differentiate between nematode-infected and uninfected sheep feces. Two German shepherd bitches were trained for scent detection over a 6-month period using operant/clicker conditioning. On completion of the training, testing was undertaken with 9 paper bags containing uninfected and 1 with infected feces, placed randomly around a circle. The dog and handler were not able to observe the placement of the bags. The 10th bag contained feces from sheep infected with either Teladorsagia circumcincta, Trichostrongylus vitrinus, Haemonchus contortus, or a mixed infection of all 3 species. Over 80 trials the dog had a mean success rate of greater than 80% in the detection of T. circumcincta- or T. vitrinus-infected feces and H. contortus-infected feces was detected with a slightly lower reliability of 76%, but mixed infections were detected at 92% reliability (one-proportion binomial analysis, P < 0.05). Trials were then undertaken to determine the time after administration of infective larvae that the dog was first able to differentiate T. circumcincta infection in sheep feces. At 7 days postinfection (dpi), the dog was capable of identifying T. circumcincta at least 85% of the time. These trials demonstrate that dogs are able to detect the common parasite infections in sheep with reliability equal to immunological assays. The results suggest that diagnostics based on odor detection using dogs to identify signature odors which could then be used to create sensitive detection devices might allow parasite detection on-farm and on all sheep in a flock.     

Egg output of Ostertagia circumcincta in sheep given single infections of varying size

The faecal egg counts of groups of sheep given 5 000, 20 000 or 40 000 infective larvae of Ostertagia circumcincta reached a peak at about 28 days after infection; the height of the peak varying directly with the number of larvae given. At 35 days the ratios of total worm burdens and the total eggs in all females amongst the groups were similar. Between 35 and 70 days post infection the faecal egg counts fell to a low level which was roughly the same regardless of the number of larvae administered. At 70 days the two groups of lambs given the two higher doses of larvae had similar worm burdens, but those given 5 000 larvae harboured about half as many. At this time the total eggs in all females was approximately the same in each group. Whatever factor, in addition to worm numbers, which may have limited egg output appeared to be ineffective until some time after the peak egg count was reached.     

Prevalence of gastrointestinal helminths,lungworms and liver fluke in sheep and goats in Norway

The present study describes the occurrence of various gastrointestinal helminths, lungworms and liver flukes in Norwegian sheep and goats as assessed from faecal samples and post mortem examinations performed between 2007 and 2010. Faecal samples for gastrointestinal nematode egg counts were collected from 77 sheep flocks and 30 dairy goat flocks from three geographical regions in Norway. Additionally, thirty-two lambs and 16 adult goats were euthanized for necropsy examination and for identification of adult gastrointestinal nematodes and tapeworms, lungworms and liver flukes. The survey showed that there was a higher mean excretion of trichostrongyle eggs in sheep than in goats at the individual level (392 EPG vs. 154 EPG, p < 0.001). For both host species, the mean prevalence and intensity of excreted trichostrongyle eggs were significantly higher in the southern coastal region compared with the inland and northern regions (p < 0.001). Third stage larvae of Trichostrongylus/Teladorsagia, Haemonchus and Nematodirus type were the most prevalent ones in the coprocultures from sheep, whereas larvae of Trichostrongylus/Teladorsagia and Nematodirus type dominated in goats. The most prevalent gastrointestinal nematode species found at necropsy was Teladorsagia circumcincta (75.0 and 81.2% respectively in sheep and goats), while the largest mean worm burdens were recorded for Haemonchus contortus in sheep (724 ± 623) and T. circumcincta in goats (377 ± 529). Other gastrointestinal nematode species were present at low prevalence or in low numbers.     

Development and spatial distribution of the free-living stages of Teladorsagia circumcincta and Trichostrongylus colubriformis on pasture: A pilot study

Abstract

AIMS: To measure the development of Teladorsagia (=Ostertagia) circumcincta and Trichostrongylus colubriformis eggs to third-stage infective larvae (L3) at different times of the year. Also, to measure the spatial distribution of L3 across herbage, soil and faeces, in order to assess whether spatial issues could be important in larval dynamics on pasture.

METHODS: Field plots were contaminated with sheep faeces containing approximately 20,000 eggs of each of T. circumcincta and T. colubriformis on five separate occasions, viz 01 December 1996 (summer), 18 March 1998 (autumn), 17 June 1998 (winter), 15 October 1998 (spring), and 23 July 1999 (winter). Replicate plots (n=10) were harvested at intervals for up to 12 months after deposition of faeces, and the number and distribution of L3 were measured. Larvae were sampled from faeces (where these remained), herbage, and three soil zones to a depth of 145 mm.

RESULTS: There were large differences between contamination dates in the percentage of eggs that developed to L3. For both species the highest percentage development was for eggs deposited in December (7.8% and 25.9% for T. circumcincta and T. colubriformis, respectively) and the lowest for June (0.4% and 0.03% T. circumcincta and T. colubriformis, respectively). Development in winter was often delayed, and this was always associated with a low yield of larvae, probably due to compounding mortalities associated with long periods of exposure to low temperatures.

The relative distribution of L3 present on herbage, in faeces or in the soil varied between sampling times. However, overall the most L3 were recovered from soil (74% and 66% for T.circumcincta and T. colubriformis, respectively, averaged over all samples), and the lowest recoveries were from the herbage.

CONCLUSIONS: Although the data are limited, the results indicated that the highest percentage of eggs developed to infective larvae in summer and only minimal development occurred in winter. The data do not support the view that substantial contamination of pastures with sheep parasites occurs over winter. Large numbers of larvae were recovered from soil, which indicates that, assuming they can subsequently migrate onto herbage, soil is a potentially important reservoir ofinfective larvae in New Zealand. Therefore, the spatial distribution of L3 on pasture may affect both the dynamics and transmission of parasite populations. Further work on both these issues is warranted.     

Study on the pathogenicity of ostertagiosis due to Ostertagia circumcincta in sheep,in Iraq

The response of Awassi sheep to Ostertagia circumcincta, the most prevalent ovine strongylid species in Iraq, was studied.A dose of 100000 larvae of O. circumcincta (Iraqi strain) induced moderate clinical symptoms of ostertagiosis. These symptoms were correlated with elevated pH of the abomasal fluid and increased plasma pepsinogen levels. There was no evidence of larval inhibition since the majority of the fourth stage larvae (L₄) continued their development. No appreciable loss of worm population was observed and most of the parasites survived and exhibited a prolific egg-laying potential six moths after initial infection. Assessments of the pathophysiological changes were performed and correlated with parasitological and clinical observations.The Awassi breed of sheep, the most prevalent in the Arab Middle East, seems to be more susceptible to ostertagiosis than other breeds and there was no spontaneous self-cure in this breed.     

Dynamics of the free-living stages of sheep intestinal parasites on pasture in the North Island of New Zealand. 1. Patterns of seasonal development

Abstract

AIM: To describe the seasonal pattern of development of third-stage infective larvae (L3) from eggs of Teladorsagia (=Ostertagia) circumcincta, Trichostrongylus colubriformis and Haemonchus contortus on pasture in the North Island of New Zealand.

METHODS: Sheep faeces containing known numbers of eggs of all three nematode species were deposited on, or buried in, pasture plots at three sites, viz coastal Manawatu, Upper Hutt Valley, and East Cape hill country. Development was measured by recovering L3 from faeces, herbage and soil 28–31 days after deposition on 13–18 occasions, between January 2005 and July 2006. Analysis of the number of larvae recovered used a mixed model including number of eggs deposited, weight of faeces recovered (an assumed indicator of earthworm activity), site, contamination date, and position of deposited faeces, i.e. on the surface or buried.

RESULTS: There was a significant effect of contamination date on development of all three species, with maximum numbers ofL3 developing between late spring (November) and early autumn (March), and minimum numbers in June and July. There were large differences between species, with H. contortus exhibiting a long period (April to October) where development was close to zero, whereas T. circumcincta developed to some extent all year round. Development of T. colubriformis was intermediate between the other two species.

Burying faeces containing nematode eggs increased the number of L3 recovered compared with surface deposition (p≤0.001), although there were a small number of exceptions involving only T. colubriformis. The weight of faeces recovered at harvest, which was assumed to be an indication of earthworm activity, was correlated with the number of L3 recovered for all species (p<0.001). In a separate analysis, earthworms were assumed tohave been active if <5 g faeces remained at harvest. Where this occurred, the number of L3 of T. colubriformis and T.circumcincta recovered was reduced by 56% and 58%, respectively (p<0.001).

CONCLUSIONS: A marked seasonal pattern of development was observed for all three species, with the most larvae developing in spring-early autumn and the least in winter. This seasonal pattern was most pronounced in H. contortus and least obvious in T. circumcincta. Burying faeces containing eggs generally resulted in more L3 being recovered, whilst the apparent activity of earthworms resulted in fewer larvae being recovered.     

Self-cure and immunity following infection and reinfection in ovine haemonchosis

Egg expulsion and post mortem examination were used to compare self-cure and resistance to reinfection with Haemonchus contortus in female Merino sheep, AA and BB hemoglobin types. Animals were infected by 500 or 5000 larvae of H. concortus once, or several times, before being challenged by 500 (low dose) or 5000 (high dose) larvae.In AA sheep infected once by 500 larvae self-cure was detected by a challenge of either 500 or 5000 larvae while in multi-infected sheep only a challenge of 5000 L₃ induced the self-cure. In BB sheep a mild self-cure was detected only by a challenge of 5000 larvae.In AA sheep receiving several infections, challenge by 500 larvae was not effective in producing resistance while 5000 L₃ was effective in producing a state of resistance for up to 35 days post challenge.At post mortem examination, a strong state of resistance was evidenced in multi-infected animals by the low number of L₄ and adults present in the abomasum up to the 35th day after challenge.In BB sheep no state of resistance was demonstrated by challenge with either 500 or 5000 L₃, even in multi-infected animals.     

Ostertagia ostertagi infections in angora goats

Angora goats grazing on a paddock contaminated with Ostertagia ostertagi and O. circumcincta were found to have infections consisting of equal numbers of both species. Subsequently, goats were infected with O. ostertagi larvae of bovine origin. The infection became patent by day 28 and egg counts reached a plateau about 7 weeks after infection. Worm numbers did not decline during the 12 weeks of the experiment. It was concluded that Angora goats are suitable hosts for O. ostertagi.     

Immunization of neonatal lambs against the larvae of Taenia hydatigena, using viable eggs followed by chemotherapy

The subcutaneous injection of viable eggs of Taenia hydatigena into neonatal lambs induced 100% protection against the development of viable T. hydatigena larvae from an oral challenge. However, contrary to some published results, no protection was induced against a simultaneous infection with eggs of T. ovis and Echinococcus granulosus.The short-acting partial resistance to oral infection with T. hydatigena eggs, transferred from infected ewes to their lambs, was not enhanced by hyperimmunization of the ewe. In the lamb, this resistance did not interfere with the development of subcutaneously injected T. hydatigena eggs into immature larvae, or with the consequent induction of resistance to an oral challenge with T. hydatigena eggs. The immunizing lesion regressed rapidly after treatment of lambs with Mebendazole, and after 6 months the lesions were negligible in both treated and untreated lambs.     

A multi-species model to assess the effect of refugia on worm control and anthelmintic resistance in sheep grazing systems

Objective Develop a computer simulation model that uses daily meteorological data and farm management practices to predict populations of Trichostrongylus colubriformis, Haemonchus contortus and Teladorsagia (Ostertagia) circumcincta and the evolution of anthelmintic resistance within a sheep flock. Use the model to explore if increased refugia, provided by leaving some adult sheep untreated, would delay development of anthelmintic resistance without compromising nematode control. Procedures Compare model predictions with field observations from a breeding flock in Armidale, NSW. Simulate the impact of leaving 1–10% of adult sheep untreated in diverse sheep‐grazing systems. Results Predicted populations of Tr. colubriformis and T. circumcincta were less than those observed in the field, attributed to nutritional stress experienced by the sheep during drought and not accounted for by the model. Observed variation in faecal egg counts explained by the model (R²) for these species was 40–50%. The H. contortus populations and R² were both low. Leaving some sheep untreated worked best in situations where animals were already grazing or were moved onto pastures with low populations of infective larvae. In those cases, anthelmintic resistance was delayed and nematode control was maintained when 1–4% of adult stock remained untreated. Conclusions In general, the model predicted that leaving more than 4% of adults untreated did not sufficiently delay the development of anthelmintic resistance to justify the increased production risk from such a strategy. The choice of a drug rotation strategy had an equal or larger effect on nematode control, and selection for resistance, than leaving 1–10% of adults untreated.     

Experimental in vitro transmission of Babesia sp. (EU1) by Ixodes ricinus

Babesia sp. (EU1), first characterized in 2003, has been implicated in human cases of babesiosis in Italy, Austria and Germany. It has been identified in roe deer and in its suspected tick vector, Ixodes ricinus, in several European countries. The aim of the present study was to validate the competence of I. ricinus as a vector of Babesia sp. (EU1) via experimental infections. For this purpose, a parasite strain isolated from roe deer was cloned in sheep erythrocytes. After experimental infections, parasite DNA was successfully amplified by PCR in both eggs and larvae originating from infected I. ricinus females and in the salivary glands of females exposed to Babesia sp. (EU1) as nymphs. We also demonstrate that infected females were able to transmit parasite DNA during a new blood meal. Together with previous epidemiological studies, these results validate I. ricinus as a competent vector for Babesia sp. (EU1).     

The Effect of Anthelmintic Capsules on the Egg Output and Larval Viability of Drug-resistant Parasites

Challenge with an equal mix of drug-resistant and drug-susceptible larvae of Teladorsagia circumcincta resulted in infections in groups of lambs (n = 6) either untreated or given controlled-release capsules, containing either albendazole or ivermectin. Lambs treated with albendazole capsules contained similar numbers of adult worms at necropsy to the other groups but had no detectable faecal egg count. Animals treated with ivermectin capsules had similar worm burdens and faecal egg counts to the control group but the worms had significantly higher numbers of eggs in utero. These results provide evidence for suppression of egg production by both anthelmintic treatments. The observation that albendazole caused a significant reduction in the developmental success of parasite eggs also has implications for the use of faecal egg count as an indicator for pasture contamination with resistant parasites. In two further groups of lambs, either untreated or given albendazole capsules, treatment caused a significant reduction in egg count and adult worm burden of Trichostrongylus colubriformis. No significant effects were observed on in utero egg counts or egg viability and the apparent effect on the number of eggs produced in faeces per adult female was not significant (p = 0.077). There was, therefore, no evidence that albendazole controlled-release capsules caused suppression of egg output in this species.     

Assessing the efficacy of Duddingtonia flagrans chlamydospores per gram of faeces to control Haemonchus contortus larvae

The aims were (a) to quantify the number of Duddingtonia flagrans chlamydospores per gram of faeces (CPG) recovered from sheep administered with different oral doses and, (b) to describe the relationship between CPG and eggs per gram of faeces (EPG) on the efficacy to reduce Haemonchus contortus infective larvae. Three doses of chlamydospores per kg BW were orally administered during seven days: (T1) non treated control group, (T2) 1 × 10⁶, (T3) 2.5 × 10⁶ and (T4) 5 × 10⁶. Three lambs, infected with H. contortus, were used per group. Faeces were obtained from the rectum of each lamb during the fungal administration period (days 0–6) and for six days after that period. Four coproculture replicates were made from each animal in days 2, 4, 6, 8 and 10. A higher chlamydospore dose produced higher CPG in faeces (p < 0.05), but a clear dose dependent effect was not found either in the larvae reduction or in the CPG:EPG ratio. When ratios were re-analyzed, independently of the treatment groups of origin, a better efficacy was obtained with a ratio from 5 to 10 CPG:EPG and a higher ratio (>10 per egg) showed a lower reduction efficacy (p < 0.05). The binomial analysis showed that for each unit of increment in CPG:EPG ratio there was a reduction of larvae number until a point (between 5 and 10 CPG:EPG) where no further reduction was detected. The surface response test indicated that the number of larvae was reduced by CPG until possible saturation. The highest CPG:EPG ratios did not necessarily improve efficacy of D. flagrans.     

捻转血矛线虫虫卵的大量分离纯化

笔者采用虫体取卵和孵化后感染性幼虫直接在培养瓶瓶盖内收集的方法,获得了大量纯净单一的感染性幼虫,并且感染绵羊成功。再对感染绵羊直肠采粪收集虫卵,就可收集到大量纯种单一虫卵,进行三期幼虫的大量孵化。该试验解决了在寄生性线虫分子生物学研究中需要大量单一虫种的纯化分离问题。     

Serodiagnosis of Metastrongylus apri infection of pigs by immunofluorescence

The lyophilised first stage larvae of Metastrongylus apri were used as antigen in the indirect fluorescent antibody test for the diagnosis of the infection of pigs. The larvae were recovered from the embryonated eggs discharged by the gravid females of the nematode in vitro. Ten pigs were used in the test. Serial serum samples were taken of seven pigs, experimentally infected with varying doses of M. apri larvae, all of which were found to discharge eggs of the nematode in their faeces after infection was established. The other three pigs were uninfected and used as controls.Positive cuticular fluorescence of the larvae was first detected with the serum samples obtained between 14 and 33 days post-infection. This fluorescence persisted with subsequent serum samples up to 85 days post-infection. Pronounced and uniform cuticular fluorescence was generally observed with the serum dilutions of up to $\text{1}\text{10}$. Earlier post-infection serum samples either exhibited no cuticular fluorescence or gave non-brilliant fluorescence. The pre-infection serum of these pigs and also serum samples obtained from the three uninfected control pigs did not give cuticular fluorescence even at the initial serum dilution of $\text{1}\text{5}$.     

Efficacy of an Arthrobotrys oligospora N mutant in nematode-trapping larvae after passage through the digestive tract of sheep

For oral applications, biocontrol of animal parasitic nematodes in ruminants, ion beam implanted nematode-trapping fungi must have the capacity to survive the passage through the digestive tract and be efficient in reducing infective larvae of nematodes in the faeces.Ion beams induced mutation in the spores of nematode-trapping Arthrobotrys oligospora. Mutants with genetic stability were bred. The fungi were cultured in bottles with corn kernels as growth media, and spores of different doses were, respectively, administered orally to each group of sheep naturally infected with gastrointestinal nematodes. The control group did not receive fungi. The faeces of these experimental animals were collected and faecal cultivations carried out. The fungal germination, growth, reproduction and predation of livestock parasitic nematode larvae were tested in laboratory. The efficacy of an A. oligospora N mutant in nematode-trapping larvae after passage through the digestive tract of sheep was tested. The results indicated that the ion beam implantation induced mutation of the nematode-trapping fungi is a positive mutation. The mutant spores through the digestive tract of sheep can kill livestock parasite nematode larvae in vitro. These results indicate the potential of the A. oligospora N mutant as a biological control agent for sheep nematodes.This study showed that such biotechnology could be explored for improving the effectiveness of the use of fungal infections to control livestock parasitic nematodes. This work represents the first application of nematode-trapping fungi in eukaryotic microorganisms.     

Egg hatch as an in vitro assay of thiabendazole resistance in nematodes

A technique is described in which thiabendazole resistance is assayed by hatching nematode eggs in solutions of thiabendazole. Eggs of Haemonchus contortus and Ostertagia circumcincta were hatched in a series of concentrations of thiabendazole in a 0.1% NaCl solution. Thiabendazole resistant strains hatched in higher concentrations of thiabendazole than non-resistant strains. The dose response thus obtained for a strain of worms can be tested statistically against strains which are known to be either resistant or non-resistant.