Fasciola hepatica: comparative effects of host resistance and parasite intra-specific interactions on size and reproductive histology in flukes from rats infected with isolates differing in triclabendazole sensitivity

The efficacies of putative fasciolicides and vaccines against Fasciola hepatica are frequently monitored in clinical and field trials by determination of fluke egg output in host faeces and by worm counts in the host liver at autopsy. Less often used are parameters based on fluke size and histology, yet these can provide important indications of specific effects on the development of particular germ-line or somatic tissues, especially in relation to the timing and profligacy of egg production. In this study, F. hepatica metacercariae of two distinct isolates, the triclabendazole (TCBZ)-sensitive Cullompton isolate and the TCBZ-resistant Oberon isolate, were administered to rats as single-isolate or mixed-isolate infections. At autopsy 16 weeks later individual adult flukes were counted, measured and the reproductive organs were examined histologically. The degree of development of the testis tubules in each fluke was represented by a numerical score, based on the proportion of the histological section profiles occupied by testis tissue. The level of anti-F. hepatica antibody in the serum of each rat was determined by ELISA. It was found that Cullompton flukes were significantly larger than Oberon flukes, and that significantly more Cullompton metacercariae developed to adults than Oberon metacercariae. The Cullompton flukes showed histological evidence of aspermy and spermatogenic arrest, which was reflected in quantitatively reduced testicular development, as compared with the Oberon isolate. In Cullompton flukes, parthenogenetic egg development is implied. The size of Cullompton and Oberon flukes was significantly related to the number of adult flukes recovered, to the number of metacercariae administered, and to the percentage success of infection. The testis development score in both isolates was significantly related to the number of adult flukes recovered but not to the number of metacercariae administered, or to the percentage success of infection. Fluke size was positively related to testis score for both isolates, and a significant negative relationship was found between percentage success of infection and metacercarial dose. The results are interpreted in terms of differing interactions between various numbers of young flukes and host immunity during invasion of and migration in the hepatic parenchyma, and of fluke intra-specific (possibly pheromonal) stimulatory effects in the final stages of development, within the host bile ducts. No significant relationships were found between host antibody levels and fluke size or testis score. False positive serological reactions were found in some rats that had been infected, but found to harbour no flukes at autopsy. Clearly the act of eliminating the flukes involved generation of an immune response.     

The effect of a parenteral ivermectin/closantel injection on the growth and reproductive development of early immature Fasciola hepatica in cattle

Sixteen calves approximately 6 months old were each infected with 500 metacercariae of Fasciola hepatica. Thirty-two days later they were weighed and divided into two groups, and on day 35 all calves in one of the groups were injected subcutaneously with an ivermectin/closantel combination. Both groups were sacrificed between days 70 and 72 to enable counting and examination of the flukes recovered from the bile ducts. Eggs released by the flukes were collected for incubation, hatching and estimation of egg viability. Flukes were counted, flat-fixed in 70% ethanol, stained with catechol and carmine and measured. The reproductive organs, namely testis, vitelline glands, ovary and uterus, were examined and scored on a 0-3 scale according to their state of development. This was visually assessed on the basis of size, distribution and staining density of their constituent tissues and the abundance of eggs in the uterus. A representative sample of flukes from each animal was fixed in formalin for histological sectioning to enable more detailed examination of the reproductive structures. Treatment of the immature flukes reduced the population in cattle by 42.6% as compared with the controls and as a result of the stunting effect due to the presence of closantel during early development the size of treated flukes was reduced by 43.9%. A bimodal pattern of size and reproductive score was also observed in flukes from treated cattle, suggesting that the stunting effect on individual flukes differed depending on whether or not they had gained access to the bile ducts or were still migrating in the hepatic parenchymal tissue at the time of drug exposure with the effect being greater once the fluke had gained access to the bile ducts. The mean reproductive score for untreated flukes was 8.76 and for treated flukes 5.64, a 35.6% reduction. This difference was highly significant (p<0.001). Egg shedding from treated flukes was significantly less than that from controls (p<0.05), but there were no differences in hatchability, suggesting that whilst drug treatment reduced the energy supply available for gametogenesis/oogenesis, it did not induce functional defects in the gonads or accessory reproductive organs. Histological examination confirmed that there was a reduction in development of testes, ovaries and vitellaria in treated flukes, with a consequent reduction in egg production. In the treated flukes, early spermatogonia and oogonia were the predominant cell types in the testes and ovary, whilst undifferentiated stem cells were abundant in the vitelline follicles. In untreated flukes, cells representing more advanced stages in gametogenesis and vitellogenesis predominated in the respective organs. It is likely that this inhibition of gametogenesis and vitellogenesis was caused by the effects of closantel treatment on intermediary metabolism in the flukes. Clearly these effects were evident even at a relatively early stage of fluke growth, and because of the impact on egg output may have epidemiological importance in addition to the reduction in fluke numbers.     

Molecular characterization of parthenogenic Fasciola sp. in Korea on the basis of DNA sequences of ribosomal ITS1 and mitochondrial NDI gene

Nucleotide sequences of ribosomal internal transcribed spacer (ITS1) and mitochondrial NADH dehydrogenase I (NDI) gene were analyzed to genetically characterize aspermic Fasciola forms in Korea. From the difference in ITS1 sequences, Korean flukes were divided into 3 haplotypes represented by Kor1, Kor2 and Kor1/2, which had nucleotides identical to F. hepatica, F. gigantica and those overlapped between the two species, respectively. NDI sequences also showed that Korean flukes could be classified into 3 distinct haplotypes (Kor1: F. hepatica-type, Kor2a and Kor2b: F. gigantica-type). The sequences of Kor1 and Kor2a were 100% identical to those of the haplotypes Fsp1and Fsp2, respectively, which are major Fasciola forms in Japan. These findings strongly suggest that aspermic Fasciola forms in Korea and Japan originated from same ancestors and have recently spread throughout both countries.     

Fasciola hepatica: comparison of flukes from Korea and the United States by isoelectric focusing banding patterns of whole-body protein.

Comparisons were made between the flukes from Chonnam, Korea and Oregon, USA by isoelectric focusing (IEF) of whole-body protein. Adult Fasciola hepatica were recovered from bile ducts of Korean native cattle. Whole-body protein of the flukes was subjected to IEF, and the banding patterns of the fluke protein were compared with those of North American F. hepatica recovered from experimentally infected calves. The overall banding pattern of F. hepatica from Korea was essentially identical to that of F. hepatica from the United States. These results provide further support for the usefulness of this technique in differentiating Fasciola species in other geographical areas.     

Molecular analysis of aspermic Fasciola flukes from Korea on the basis of the nuclear ITS1 region and mitochondrial DNA markers and comparison with Japanese aspermic Fasciola flukes

It has been speculated that populations of aspermic Fasciola flukes in Korea and Japan have a close phylogenetic relationship. To evaluate this, we analyzed 33 Korean aspermic Fasciola flukes on the basis of nuclear ribosomal internal transcribed spacer 1 (ITS1) and mitochondrial NADH dehydrogenase subunit 1 (nad1) and cytochrome c oxidase 1 (cox1) sequences. Fh, Fg, and Fh/Fg types were detected in the ITS1 region and displayed the fragment patterns of F. hepatica, F. gigantica, and both species, respectively by a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Additionally, three concatenated haplotypes of nad1 and cox1(nad1/cox1) were detected, and 2 of these, Kor1/Kor1 (Fsp1/Fsp1) haplotype and Kor2a/Kor2 (Fsp2/Fsp2) haplotype, were shared by Korean and Japanese aspermic flukes. The Fst value (0.019), calculated using the concatenated sequences, indicated that Korean and Japanese aspermic Fasciola populations were genetically undifferentiated. Interestingly, a combination of the Fh/Fg type and Kor1/Kor1 haplotype was found at the highest frequency in Korean aspermic flukes, whereas the Fg type and Fsp2/Fsp2 haplotype combination was found at a conspicuously high frequency in Japanese aspermic flukes. This indicates that a founder effect caused by the introduction of infected hosts may have played a key role in the introduction of aspermic Fasciola flukes from Korea into Japan.     

Fasciola hepatica: Histology of the testis in egg-producing adults of several laboratory-maintained isolates of flukes grown to maturity in cattle and sheep and in flukes from naturally infected hosts

A total of 8 calves approximately 6 months old and 22 lambs of similar age were infected with metacercariae of Fasciola hepatica of various laboratory-maintained isolates including: Cullompton (sensitive to triclabendazole) and Sligo, Oberon and Leon (reported as resistant to triclabendazole). Ten to 16 weeks after infection, flukes were harvested from these experimental animals and the histology of the testis tissue was examined in a representative sample of flukes from each population. Adult wild-type flukes were also collected from 5 chronically infected cattle and 7 chronically infected sheep identified at post-mortem inspection. The testis tissue of these flukes was compared with that of the various laboratory-maintained isolates. Whilst the testes of the wild-type, Oberon and Leon flukes displayed all the usual cell types associated with spermatogenesis in Fasciola hepatica (spermatogonia, spermatocytes, spermatids and mature sperm), the Cullompton flukes from both cattle and sheep showed arrested spermatogenesis, with no stages later than primary spermatocytes represented in the testis profiles. The presence of numerous eosinophilic apoptotic bodies and nuclear fragments suggested that meiotic division was anomalous and incomplete. In contrast to the wild-type flukes, no mature spermatozoa were present in the testes or amongst the shelled eggs in the uterus. A high proportion of the eggs collected from these flukes hatched to release normal-appearing miracidia after an appropriate incubation period, as indeed was the case with all isolates examined and the wild-type flukes. It is concluded that the eggs of Cullompton flukes are capable of development without fertilization, i.e. are parthenogenetic. The implications of this for rapid evolution of resistant clones following an anthelmintic selection event are discussed. Amongst the Sligo flukes examined, two subtypes were recognised, namely, those flukes with all stages of spermatogenesis and mature spermatozoa present in the testes (type 1), and those flukes with all stages of spermatogenesis up to spermatids present, but no maturing spermatozoa in the testes (type 2). Each sheep infected with the Sligo isolate had both type 1 (approximately 60%) and type 2 (approximately 40%) flukes present in the population. Spermatozoa were found amongst the eggs in the uterus in 64% of flukes and this did not necessarily reflect the occurrence of spermatozoa in the testis profiles of particular flukes, suggesting that cross-fertilization had occurred. The apparent disruption of meiosis in the spermatocytes of the Cullompton flukes is consistent with reports that Cullompton flukes are triploid (3n=30), whereas the Sligo and wild-type flukes are diploid (2n=20). In the Sligo flukes the populations are apparently genetically heterogenous, with a proportion of the flukes unable to produce fully formed spermatozoa perhaps because of a failure in spermiogenesis involving elongation of the nucleus during morphogenesis.     

两种片形吸虫感染家兔的试验比较

用大片形吸虫和肝片形吸虫感染家兔以便选择大片形吸虫对动物的最佳感染量,及明确肝片形吸虫和大片形吸虫的生物学和对动物宿主的致病力的差别。结果显示肝片形吸虫虫体在兔体内发育成熟的时间早于大片形吸虫,感染成活率更高,对动物的病理损害明显比感染大片形吸虫兔的病变要轻。本试验证实这两种片形吸虫除了形态学的差异外,在对动物致病力、病理损害等方面确实存在差别。     

Identification of the Egyptian species of Fasciola.

Reports on the species of Fasciola present in the Nile Delta, Egypt, appear controversial. Some authors reported the presence of both Fasciola gigantica and Fasciola hepatica, others reported F. gigantica only and mentioned that F. hepatica was found only in imported animals.This study was an attempt to identify the species of Fasciola flukes collected from locally bred animals. Morphologic, morphoanatomic, morphometric, and chemotaxonomic criteria of the fluke isolates were studied. Speciation based on morphologic and morphometric data was not decisive due to overlap in the values of most measurements. Morphoanatomic data proved the presence of both the species, and isoelectric focusing (IEF) of fluke soluble protein confirmed the presence of both F. gigantica and F. hepatica in Egypt.     

Fasciola hepatica: A comparative survey of adult fluke resistance to triclabendazole,nitroxynil and closantel on selected upland and lowland sheep farms in Northern Ireland using faecal egg counting,coproantigen ELISA testing and fluke histology

In order to investigate the incidence and distribution of adult fluke resistance to the fasciolicide tricalbendazole (TCBZ) amongst populations of Fasciola hepatica in sheep flocks in Northern Ireland (NI), individual rectal faeces samples were collected from 3 groups of 20 sheep, before (pre-dose), and 21 days after (post-dose) treatment of the animals with TCBZ, nitroxynil or closantel, on each of 13 well-managed sheep farms distributed across the province. The efficacy of each flukicide was determined for each farm, using faecal egg count reduction (FECRT) and F. hepatica coproantigen ELISA testing. In certain flocks, 2 sheep with high pre-dose faecal egg counts (FEC) were killed 3 days and 21 days respectively after TCBZ treatment, and the histology of the fluke reproductive organs was compared with that of flukes from untreated sheep, and from sheep treated with nitroxynil or closantel 2 days prior to death, using haematoxylin and eosin (H&E) staining and an in situ hybridisation method (TdT-mediated dUDP nick end labelling [TUNEL]) to demonstrate apoptosis. Results from FECRT revealed that in all flocks with a high fluke burden, TCBZ was ineffective in treating chronic fasciolosis, and this finding was generally supported by the results of the coproantigen reduction test (CRT). The histology of reproductive organs of flukes from TCBZ-treated sheep in these flocks was normal, when compared with untreated flukes, and this, together with the FECRT and CRT findings, indicated a likely diagnosis of TCBZ resistance in all the flocks with a high fluke burden. In contrast, nitroxynil and closantel were found to be fully effective against TCBZ-resistant flukes in each of the flocks bearing a high chronic fluke burden. All of the flocks with a high fluke burden and TCBZ resistance were managed on lowland in the South and East of NI. Upland flocks, in the North and West, had low fluke burdens, or were clear of infection; and FECs were too low to allow valid resistance testing. The study highlights the high level of penetration of TCBZ resistance throughout F. hepatica populations in areas of intensively managed sheep production with a high level of fluke challenge. Further, it emphasises the importance of pre-emptive chemotherapeutic action against chronic fasciolosis, using flukicides effective against the egg-producing adult flukes to minimise pasture contamination for the next season's lamb crop. This study also exemplifies the use of several complementary methods (FECRT; CRT; fluke histology; comparative anthelmintic efficacy testing) for confirmation of a diagnosis of fluke drug resistance.     

Innate and adaptive resistance of Indonesian Thin Tail sheep to liver fluke: a comparative analysis of Fasciola gigantica and Fasciola hepatica infection

In the current study, three independent trials directly compared Fasciola gigantica and Fasciola hepatica infection of ITT sheep. In all trials, F. hepatica infection resulted in higher worm burden recoveries and greater physiological damage to ITT sheep. Developmental differences of the two Fasciola species were also observed during the first twelve weeks of a primary infection, where the migration and growth of F. hepatica was more rapid than F. gigantica. Various immunological blood parameters were measured and indicated similar kinetics in the humoral and cellular responses during the time course of infection with each Fasciola species. In contrast to F. hepatica infection, we demonstrate an innate and adaptive comparative ability of ITT sheep to resist the early stages of infection with F. gigantica infection. Unraveling the mechanisms leading to this differential resistance may potentially lead to new methods for the control of fasciolosis and other human liver flukes.     

The effect of Taenia hydatigena infection on existing and concurrent infections of Fasciola hepatica in sheep

Sheep given a primary infection of Fasciola hepatica were challenged 18 weeks later with Taenia hydatigena or F hepatica, or both parasites together, or were not challenged. At the same time, control sheep were infected with T hydatigena and/or F hepatica separately or concurrently. All sheep were killed seven weeks after challenge and the number of cysts and flukes counted. Challenge infection with T hydatigena did not affect the numbers of flukes recovered from either primary or challenge F hepatica infections. On the other hand, the numbers of cysticerci were reduced in sheep previously infected with F hepatica but not in those given T hydatigena and F hepatica concurrently.     

Penetration in vitro of newly excysted juvenile flukes of Japanese Fasciola sp. through ligated intestines of rabbits, mice, rats and chickens.

Ligated intestines of rabbits, mice, rats and chickens were used to examine the penetration of newly excysted juvenile flukes of Japanese Fasciola sp. in vitro. In rabbit intestines, the penetration rate was relatively high in the rectum and duodenum. Penetration rates in the jejunum, ileum, cecum and colon were comparable to those in the rectum and duodenum, although it was lower in the appendix. In the case of mouse, juvenile flukes penetrated the duodenum, jejunum, cecum, and rectum at considerably high rates. In rat intestine, penetration by flukes was less in the duodenum and rectum, although flukes were detected in the jejunum. In chicken intestine, flukes barely penetrated the duodenum, jejunum and rectum. Consequently, newly excysted flukes of Fasciola sp. seem to penetrate any region of the intestine in rabbits and mice. In rats, the middle small intestine may be the site suitable for flukes to penetrate. In chickens, the difficulty in penetration of the intestinal wall may be one of the reasons why chickens are scarcely infected with Fasciola sp.     

Serum antibody isotype responses of Fasciola-infected sheep and cattle to excretory and secretory products of Fasciola species

This study investigated the immunoglobulin isotype responses of sheep and cattle chronically infected with Fasciola hepatica and Fasciola gigantica to adult F. hepatica excretory/secretory products (Fh-ES) or F. gigantica excretory/secretory products (Fg-ES), respectively. An antibody enzyme-linked immunosorbent assay (Ab-ELISA) was used to determine serum antibody (total Ig, IgG(1), IgM, IgG(2) and IgA) responses. At necropsy, the mean number of flukes recovered was lower in cattle than in sheep. All F. hepatica and F. gigantica infected sheep and cattle showed an increased total Ig levels from 3 to 4 weeks post-infection (wpi). Among isotypes IgG(1) was most dominant while IgM was the earliest (2 wpi) to be detected in both sheep and cattle infected with both F. hepatica and F. gigantica animals. IgG(2) response was early (2 wpi) in sheep infected by F. hepatica but there was no response in sheep infected with F. gigantica. There was a late and strong IgG(2) response in cattle infected with both flukes. The IgA isotype showed an early and a clear biphasic response in sheep with F. hepatica but was less pronounced in F. gigantica infected sheep. While IgA response to Fh-ES was noticed 5 wpi in F. hepatica infected cattle, it appeared much later (21 wpi) in those infected with F. gigantica. The dominance of IgG(1) isotype in infected sheep and cattle suggest an associated Th2 response. This early response to adult Fasciola spp. ES antigen suggests an early exposure to the antigen presumably through the cross-reacting ES products of juvenile flukes. There is clearly difference in IgG(2) isotype response in cattle (resistant) compared to sheep (susceptible). The late IgG(2) response in cattle may suggest late Th1 involvement in bovine cellular responses to adult Fh-ES/Fg-ES.     

The effect of newer anthelmintics on Fasciola hepatica in experimentally infected rats]

The reports deals with the results of testing seven new antihelminthics for Fasciola hepatica in the experimentally invaded Wistar rat. The greatest influence on juvenile flukes (2 and 4 weeks of age) was exerted by diamphenetid (Coriban) applied in a single dose of 100 mg kg-1. Hexachlorophene applied in the dose of 50 mg kg-1 showed the highest effect on sexually mature flukes. All the tested antihelminthics of the halogenated salicylanilide group were ineffective on juvenile stages and only slightly effective on mature F. hepatica flukes. It follows from the results that the effectiveness of some antifasciolics on laboratory animals need not always be in correlation with their effect in ruminants - hence it is necessary to verify the results obtained in laboratory animals and to check them on natural F. hepatica hosts.     

Fasciola hepatica: Histological demonstration of apoptosis in the reproductive organs of flukes of triclabendazole-sensitive and triclabendazole-resistant isolates,and in field-derived flukes from triclabendazole-treated hosts,using in situ hybridisation to visualise endonuclease-generated DNA strand breaks

Investigation of the triclabendazole (TCBZ) resistance status of populations of Fasciola hepatica in field cases of fasciolosis, where treatment failure has been reported, can be supported by histological examination of flukes collected from recently treated hosts. In TCBZ-sensitive flukes (TCBZ-S) exposed to TCBZ metabolites for 1–4 days in vivo, but not in TCBZ-resistant flukes (TCBZ-R), morphological changes suggestive of apoptosis occur in cells undergoing meiosis or mitosis in the testis, ovary and vitelline follicles. In order to verify or refute the contention that efficacy of TCBZ treatment is associated with apoptosis in the reproductive organs of flukes, histological sections of TCBZ-S (Cullompton isolate) flukes and TCBZ-R (Sligo isolate) flukes were subjected to the TdT-mediated dUDP nick end labelling (TUNEL) in situ hybridisation method, a commercially available test specifically designed to label endonuclease-induced DNA strand breaks associated with apoptosis. Additionally, sections of in vivo-treated and untreated flukes originating from field outbreaks of suspected TCBZ-S and TCBZ-R fasciolosis were labelled by the TUNEL method. It was found that in treated TCBZ-S flukes, strong positive labelling indicating apoptosis was associated with morphologically abnormal cells undergoing mitosis or meiosis in the testis, ovary and vitelline follicles. Background labelling in the positive testis sections was attributed to heterophagy of cell debris by the sustentacular tissue. The triggering of apoptosis was probably related to failure of spindle formation at cell division, supporting the contention that TCBZ inhibits microtubule formation. In treated TCBZ-R (Sligo Type 1) flukes, and in treated flukes from field outbreaks of suspected TCBZ-R fasciolosis, no significant labelling was observed, while sections of fluke derived from a field case of fasciolosis where TCBZ resistance was not suspected were heavily labelled. Light labelling was associated with the testis of untreated Cullompton (TCBZ-S) and Sligo Type 2 (TCBZ-R) flukes, which exhibit abnormal spermatogenesis and spermiogenesis, respectively. This was attributed to apoptosis and to heterophagy of effete germ line cells by the sustentacular tissue. It is concluded that demonstration of apoptosis by in situ hybridisation using the TUNEL method on sections of 1–4 days in vivo TCBZ-treated F. hepatica can contribute to the diagnosis of TCBZ resistance in field outbreaks of fasciolosis.     

Cellular and humoral responses in liver of cattle and buffaloes infected with a single dose of Fasciola gigantica

The cellular components of the hepatic inflammatory infiltrate in cattle and buffaloes infected with a single dose of 1000 Fasciola gigantica were analysed by immunohistochemistry and histology. T and B lymphocytes, plasma cells, eosinophils and mast cells were present in the hepatic lesions. It is proposed that both cellular and humoral immune responses were induced in the liver of cattle and buffaloes during infection with F. gigantica probably by antigens released by the developing flukes and by damage caused by the flukes during their migration in the liver. The local T cell response differed between these animals, with the response decreasing after 3 weeks post-infection in cattle in contrast to a gradually increasing response in buffaloes. Difference in the T cell response between cattle and buffaloes may be related to their differences in resistance and resilience to infection with F. gigantica.     

Epizootiology of fascioliasis in Montana.

During 1989-1990, the United States Department of Agriculture (USDA) meat inspection records were used to determine the distribution and incidence of liver flukes (Fasciola hepatica and Fascioloides magna) in Montana cattle. Of the cows and bulls slaughtered in USDA-inspected packing plants during a 12 month time period, 17.24% had livers that were condemned because of liver flukes. This was a 12% increase over USDA liver condemnations reported for 1973. Infected animals have been reported from 26 counties in Montana, mostly located in the south-central and western half of the state. Forty-nine percent of the 2.4 million cattle in Montana are raised in these counties. Lymnaeid snail species that may serve as intermediate hosts for Fasciola hepatica were found in most of the counties where liver flukes were reported. The principal vectors believed to be responsible for the transmission of Fasciola hepatica in Montana are species of the genus Fossaria. Stagnicola montanensis and Lymnaea stagnalis, which may serve as intermediate hosts for this parasite have also been collected. A known intermediate host for Fascioloides magna, Stagnicola caperata, was also found in several locations.     

The sensitivity and specificity of two methods for detecting Fasciola infections in cattle.

Counts of Fasciola spp. eggs in faeces and measurements of antibody concentration to the excretory/secretory antigens of Fasciola spp. by ELISA were related to the numbers of flukes in the livers of 92 cattle killed in the abattoirs of Hanoi City, Vietnam. In this population, about 22% of the cattle had no flukes, another 22% had between 1 and 10 flukes, 44% between 11 and 100 flukes and 12% had more than 100 flukes in their livers. Of the 14 animals less than 2 years of age, only three were infected. At 2 years of age the mean number of flukes per liver was 10 whereas at 3 years and older, the mean varied between 60 and 80 flukes. Prevalence of infection was 78.3%. No eggs of Fasciola spp. were detected in the faeces of one third of infected cattle and 60% of the counts were less than 100 eggs per gram. The sensitivity of the egg counting method was 66.7% and specificity 100%, overall accuracy was 73.9%. Corresponding values for the ELISA method were 86.1, 70 and 82.6%, respectively. The positive and negative predictive values for the egg counting method were 100 and 45.5% and for the ELISA method were 91.2 and 58.3%, respectively.     

The shedding of the outer glycocalyx of juvenile Fasciola hepatica

Freshly excysted Fasciola hepatica possess an outer glycocalyx which on incubation at 37 degrees C is rapidly shed. Using an ELISA technique the release of this parasite antigen was shown to be temperature-dependent and to occur in both normal bovine serum as well as in serum free conditions. The ELISA failed to detect the antibody--antigen complexes that occurred when flukes were incubated in immune serum. Release of specific parasite antigen fell slowly with in vitro cultured flukes, but increased with in vivo cultured flukes. Using a fluorescence inhibition assay, antigens with high ELISA titres inhibited surface fluorescence of the parasite suggesting that the ELISA was detecting surface antigens as well as other parasite metabolic products. Several metabolic blocking agents and commercial antihelminthics were titrated against juvenile F. hepatica to screen for inhibition of the surface--tegument shedding: there was little selective inhibition of surface shedding without a significant loss of motility.     

The susceptibility of the goat to Fasciola hepatica infections

Mixed breed goats were infected with metacercariae of Fasciola hepatica and the resulting worm burdens were quantitated after primary and secondary exposure of the goats to the parasite. Mean length and width of the parasite recovered after all primary exposures were 1.91 +/- 0.2 cm and 0.91 +/- 0.2 cm, respectively. A mean of 71.8 +/- 5.9% of the flukes were recovered from all of the primary infections. In the secondary infections, the mean length and width of the flukes from the physically smaller population was 0.88 +/- 0.27 cm and 0.53 +/- 0.19 cm, respectively. A mean of 67 +/- 6.7% of the flukes were recovered from this secondary infection. It appears that the goat is susceptible to challenge infections with F. hepatica and that its response to this infection is much like that of sheep.