Serological responses of cattle after treatment and during natural re-infection with Fasciola hepatica, as measured with a dot-ELISA system

The dot-ELISA reaction was used to study the dynamics of IgG titers in cattle naturally infected with Fasciola hepatica after anthelmintic treatment and during reinfection. Excretion/secretion products (ES) of the parasite were used as antigens for the dot-ELISA. IgG antibodies were no longer detectable by dot-ELISA, 4-6 months after nine animals received the first of three weekly doses of triclabendazole (15 mg kg(-1)) and were then maintained on a pasture free of F. hepatica metacercariae. Six fluke-free cattle began shedding F. hepatica eggs 3-6 months after grazing a pasture contaminated with metacercariae of the parasite. A detectable increase in dot-ELISA IgG antibody levels was observed 2-4 weeks after natural reinfection by grazing a similar pasture contaminated with F. hepatica metacercariae. The usefulness of the dot-ELISA system to diagnose chronic infection by serology is complicated by previous treatment against the parasite. It is concluded that the ES antigens can be useful to detect early infection of cattle with F. hepatica in a dot-ELISA system     

Myocastor coypus as a reservoir host of Fasciola hepatica in France

To clarify the role of the nutria Myocastor coypus in the epidemiology of domestic fasciolosis in Loire-Atlantique (department of western France), 438 nutrias were trapped in 9 humid areas of the department and 304 nutrias were trapped in 3 farms where Fasciola hepatica was present; all animals were necropsied. Liver flukes were found in 160 nutrias: 38 nutrias randomly taken in the department (8.7%) and 122 trapped in fasciolosis areas (40.1%). The average parasitic burden was 5.7 flukes per nutria. Sixty-five percent of the liver flukes measured more than 18 mm (size of sexual maturity). The coproscopic examinations carried out on 144 infected nutrias showed that 90% of the infected nutrias shed fluke eggs. The hatching rate was 39.6%. Two groups of 100 Lymnaea truncatula snails, originating from 2 different populations, were exposed to F. hepatica miracidiae hatched from eggs collected from infected nutrias. The prevalence of the infection was 74% and 58.6% in the 2 groups of snails. The average redial burden was 6.2 rediae per snail. The total number of metacercariae was 72.4 metacercariae per snail producing cercariae. Two groups of 5 sheep were orally infected by 150 metacercariae of nutria or sheep origin, respectively. The installation rates of F. hepatica in sheep were respectively 31.6% and 29.6% for the two groups. Specific antibody kinetics of sheep were similar whether the metacercariae were of nutria or sheep origin. M. coypus allows the complete development of F. hepatica and releases parasitic elements that are infective for domestic ruminants. Because of its eco-ethologic characteristics, the nutria could be a potential wild reservoir of F. hepatica in France.     

The influence of different food sources on cercarial production in Lymnaea truncatula experimentally infected with Digenea

Experimental infections of Lymnaea truncatula with a digenean species (Fasciola gigantica, F. hepatica, or Paramphistomum daubneyi) were performed under laboratory conditions to study the effect of four sources of food (microalgae, romaine lettuce, wheat germs, or modified Boray diet) on cercarial production. The mean number of F. hepatica, F. gigantica, or P. daubneyi cercariae was significantly greater when snail food was microalgae or Boray diet. The lowest cost prices for 100 metacercariae of F. hepatica (9.7-10.1 euros) were noted when algae of Boray diet were used as food for snails. In contrast, the highest prices (18.0-18.5 euros) were found in lettuce-reared snails. The choice of a source of food for breeding the intermediate hosts of F. hepatica and collecting metacercariae must take into account the importance of the commercial demand for metacercariae and will depend on the strategy the producer wishes to develop.     

Fasciola hepatica: an assessment on the vectorial capacity of Radix labiata and R. balthica commonly found in Belgium

A previous study conducted in Belgium revealed that genetic material of Fasciola sp. was present in snail species belonging to the genus Radix. Here, these snails were collected and identified by DNA-based techniques as Radix labiata and Radix balthica. These two species and Galba truncatula (the major intermediate host in Europe) were experimentally infected with Fasciola hepatica. The resulting metacercariae were fed to rats and the infection was monitored using several techniques. Microscopy revealed the presence of larval stages in 78.3, 45, and 6.25% of G. truncatula, R. labiata, and R. balthica snails, respectively. These results were confirmed by a PCR that amplifies a Fasciola sp. specific sequence. Furthermore, this PCR was found to be more sensitive than microscopic examination. R. labiata shed fewer metacercariae than G. truncatula but these were as infective to rats as those shed by G. truncatula. This study demonstrates that R. labiata may act as an incidental intermediate host for F. hepatica in Belgium.     

Effect of parasite burden on the detection of Fasciola hepatica antigens in sera and feces of experimentally infected sheep

The effect of Fasciola hepatica parasite burden on the detection of excretory/secretory (E/S) antigens in sera and feces of experimentally infected sheep was evaluated using a double antibody-based capture enzyme-linked immunosorbent assay (ELISA). Four groups of five sheep each were used. The first three groups were infected with 50, 100 and 200 metacercariae of F. hepatica, and the fourth group remained as non-infected control. On the day of infection and weekly thereafter, serum and fecal samples were taken. ELISA detected F. hepatica E/S antigen levels in serum from the first week post-infection (wpi) and in fecal supernatant from the fourth wpi, which were significantly (p<0.05) higher than controls. F. hepatica eggs were not detected until after the eighth wpi. The correlation between absorbance of E/S antigens in serum with the fluke burden was 0.77 (p<0.0001) and in feces 0.76 (p<0.0001) at 12th wpi. The sensitivity of the assay to detect E/S antigens in serum was 86.6% and in feces 93.3%. It is concluded that the ELISA technique used in this study offers a diagnostic alternative for detecting early infections of F. hepatica in sheep.     

comparison of an indirect haemagglutination assay and an Elisa for diagnosing Fasciola hepatica in experimentally and naturally infected sheep

Summary

An enzyme‐linked immunosorbent assay (ELISA) with somatic (S) or excretory‐secretory antigens (ES) was compared with an indirect haemagglutination assay (IHA) for ability to detect antibodies against Fasciola hepatica in sheep. The specificity of both assays was determined by testing sera collected from sheep experimentally or naturally mono‐infected with Fasciola hepatica, Haemonchus contortus, Ostertagia circumcincta, Cooperia curticei, Taenia ovis, Eimeria spp., Trichostrongylus vitrinus, Trichostrongylus colubriformis or Nematodirus battus respectively. With S or ES antigens the specificity of the ELISA was 98% or 95% respectively, whereas the specificity of the IHA was 86%. Antibodies directed against Fasciola hepatica were detected by the ELISA with S or ES antigens from 2 weeks after infection until the end of the experiment, whereas the IHA detected antibodies from week 3. We conclude that the ELISA with S antigens compares favourably with the IHA and can be used for the serodiagnosis of ovine fasciolosis in the Netherlands.     

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.     

Herd evaluation of Fasciola hepatica infection levels in Louisiana cattle by an enzyme-linked immunosorbent assay

An enzyme-linked immunosorbent assay (ELISA) was evaluated as a method of determining relative immunoconversion rates in calves and how immunoconversion rates and strength of optical density values correlate with prevalence of Fasciola hepatica fecal egg shedding. Ten to 55 calves and cows were examined from each of 10 separate beef cattle herds in central and southern Louisiana. Infection prevalence rates for calves averaged 8% higher when ELISA optical density values were used than those when fecal egg count data were used. Of 55 calves in 8 herds that were ELISA positive, 39 were shedding F hepatica eggs; of 53 calves that were shedding eggs, 14 were ELISA negative. Significant correlation of calf fecal and ELISA prevalence was observed for 8 herds by linear regression analysis. A chi 2 analysis showed that calf ELISA and fecal egg shedding data were not independent. Results indicate that positive ELISA reactions for as few as 10 to 15 calves from the fall calf crop of a given herd are sufficiently accurate to be used to assess F hepatica herd infection rates, the likelihood of liver condemnations at feedlot destinations, and variation between individual farms in fascioliasis infection risk. The test was less valuable as a diagnostic test when used in adult animals previously exposed to F hepatica or on an individual animal basis.     

THE EFFECT OF PRIOR INFECTION WITH FASCIOLA HEPATICA ON THE RESISTANCE OF SHEEP TO THE SAME PARASITE

The development of flukes that resulted from a challenge dose of 175 F. hepatica metacercariae was compared in 4 groups of sheep that were maintained under grazing conditions on fluke-free pasture. One group had been previously uninfected and all subgroups of each of the other 3 groups had been exposed to one of a range of previous doses of metacercariae. The preliminary infections of 2 of the 3 groups had been terminated respectively after 7 and 14 weeks. The third previously infected group had received its preliminary infection as 2 doses of metacercariae, 7 weeks apart. The latter infection had been terminated after 14 weeks. No appreciable differences in the mean numbers, length, prepatent period or fecundity of flukes, established as a result of the challenge dose of metacercariae, were detected between the control group and the 3 previously infected groups. It was concluded that under the conditions of the present experiment no evidence was detected that would suggest that previous infection with F. hepatica conferred any significant resistance to a future challenge.     

大片形吸虫幼虫的培育及其感染小鼠的研究

本试验旨在培育大片形吸虫囊蚴,着重研究大片形吸虫在中间宿主体内的发育过程和临床病理变化.人工培养和孵化大片形吸虫虫卵,用孵化出来的大片形吸虫毛蚴感染中间宿主小土蜗螺,收集大片形吸虫囊蚴,再用囊蚴感染试验小鼠.结果显示,囊蚴经口感染试验小鼠后1周即可在肝脏找到虫体,幼虫可在小鼠体内发育生存7～8周,随着时间的推移和感染囊蚴数量不同,可给小鼠肝脏造成不同程度的病变,甚至造成死亡.剖检小鼠可见肝脏质地脆,颜色发黄,脾脏肿大等严重病理变化.因此,可利用小鼠作为大片形吸虫幼虫感染的试验动物,进行片形吸虫病的早期诊断、免疫预防和治疗等方面的研究.     

Infection of Lymnaea truncatula and Lymnaea glabra by Fasciola hepatica and Paramphistomum daubneyi in farms of central France

Natural infection of Lymnaea glabra and Lymnaea truncatula by Fasciola hepatica and/or Paramphistomum daubneyi was studied at two periods of risk (June-July and September-October) in 11 French farms known for their high prevalences of F. hepatica infection in ruminants. A total of 1,778 L. truncatula and 2,396 L. glabra measuring 6 mm or more in height were collected to determine the prevalence of natural infection with F. hepatica and P. daubneyi. The role of four factors, i.e. season, snail co-infection (F. hepatica-P. daubneyi), ruminant host and the existence of single or mixed intermediate host communities, was investigated. There were no differences in prevalences between the two risk periods. Co-infections in L. glabra were more frequent than expected. The prevalences of infection with F. hepatica or P. daubneyi in L. glabra were higher in farms rearing sheep than in farms rearing cattle. A similar finding was also noted for L. truncatula infected with P. daubneyi only. L. glabra was a much better intermediate host for F. hepatica and P. daubneyi when it was the only available snail, possibly indicating an adaptation of parasites to their less usual host in local conditions. The two trematodes preferably developed in L. truncatula rather than in L. glabra when both host species lived in the same places.     

Experimentally induced Faciola hepatica infection in white-tailed deer. I. Clinicopathological and parasitological features.

Six white-tailed deer (Odocoileus virginianus) and six sheep were inoculated with metacercariae of Fasciola hepatica. Two animals of each species were given 100, 500 or 2500 metacercariae. Clinicopathological features of these infections were determined by analyses of blood samples collected each week from inoculated deer and sheep as well as from two noninoculated animals of each species. One animal in each inoculated group was killed and examined at six weeks postinoculation and the remainder at 15 weeks postinoculation. Compared with the values obtained from noninoculated controls, eosinophilia, hyperproteinemia and hyperglobulinemia occured in inoculated deer. There were no other significant changes in hematological values or in serum aspartate aminotransferase levels. Marked leukocytosis and eosinophilia, with hyperproteinemia, hyperglobulinemia, hypoalbuminemia, elevated serum aspartate aminotransferase levels and mild macrocytic normochromic anemia characterized the infection in lambs. Although approximately 29% of the inoculum was recovered from the hepatic parenchyma of the sheep, F. hepatica was found in only one of six inoculated deer. A patent infection was established in this deer and constitutes the second report of mature F. hepatica in this host.     

Temporal studies on Fasciola hepatica in Galba truncatula in the west of Ireland

The population dynamics and prevalence of Fasciola hepatica in Galba truncatula were investigated on the Teagasc hill sheep farm in Country Mayo, Ireland from August 2006 until March 2008. Galba truncatula (n=974) were collected at fortnightly intervals in 4 snail habitats on the farm, their relative density and mean monthly rainfall and temperature were recorded. Snail abundance was associated with rainfall and temperature. G. truncatula abundance was generally low during winter 2006/2007 (December 2006-February 2007), mid spring and summer 2007 (April-July) and late autumn and winter 2007/2008 (October 2007-February 2008). Overall, two peaks in G. truncatula abundance were evident during the study period, an early spring (March) and late summer/autumn (late August-October) peak. Seasonal differences were observed in the prevalence of F. hepatica in G. truncatula, the greatest prevalence was evident in 2007 during the summer (25%) and autumn (16%). Two seasonal transmission peaks, as evident by the presence of mature infections in snails, one in summer/autumn and the other in late winter/early spring. Not unsurprising, cercariae (28%) and rediae (29%) were the predominant developmental stages observed in mature and pre-adult snails respectively. Worthy of note in this study, cercariae were also evident in pre-adult snails. The present study highlights the influence of milder temperatures and wetter conditions on both snail and fluke development. If the trend in mild and wet weather conditions continues, there is the potential for an increase in the number of G. truncatula habitats and ultimately the prevalence of fasciolosis in grazing livestock in Ireland.     

Emergence of fasciolosis in cattle in East Anglia

Liver fluke (Fasciola hepatica) infection caused weight loss, diarrhoea, decreased milk yield and occasionally death in cattle in East Anglia during the winters of 2001 to 2003. The condition had previously been limited mainly to stock imported into this part of Britain from endemically infected areas. In composite faecal samples collected by 16 farm animal veterinary practices in Norfolk, Suffolk and Essex, fluke eggs were found in 15 (28.8 per cent) of 52 previously unaffected suckler herds and 10 (16.7 per cent) of 60 dairy herds. Antibodies to F hepatica were detected by elisa in 32 (53.3 per cent) of the bulk milk samples from these 60 dairy herds, including the 10 in which fluke eggs were found. The emergence of fasciolosis in East Anglia was attributed to recent higher summer rainfall, which favoured the intermediate snail host Lymnaea truncatula and the free-living stages of F hepatica, the increased influx of sheep from endemic fluke areas for seasonal grazing, and the wetter grazing conditions associated with the Environmentally Sensitive Area scheme.     

The effect of temperature and humidity on longevity of metacercariae of Fasciola gigantica

Studies were undertaken on the viability of the metacercariae of Fasciola gigantica when stored in water at 13^˚C for periods up to 23 weeks, exposed to the sunlight for up to 8 h or stored at a range of temperatures and humidities for up to 10 weeks. Excysted metacercariae were catergorized microscopically as viable (motile and undamaged), dubious (not motile and undamaged) or dead (visible necrosis). The infectivity of viable and dubious metacercariae and unselected reference metacercariae held in water at 7^˚C for 20 days or longer was assessed by comparing numbers of flukes recovered from infected Merino sheep. Mean recovery rates were 54.6%, 7.2% and 37.2%, respectively, for viable, dubious and unselected metacercariae. Metacercariae immersed in water remained viable longer than those allowed to desiccate. Viability was promoted by decreasing temperature and increasing humidity. Exposure to direct sunlight killed metacercariae within 8 h. Results indicated that in lowland Indonesian irrigated rice paddies, metacercariae immersed in water are likely to survive for less than 5 weeks while those that become desiccated will survive less than 2 weeks. This information, together with the option of exposing fresh rice stalks to direct sunlight before feeding them to livestock, can assist farmers in reducing infection with F. gigantica.     

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.     

Seasonal transmission of Fasciola hepatica to cattle in northwestern United States

Sentinel steers were placed with 3 beef herds on irrigated pastures in southern Idaho for 1-month periods from May until November 1982 to determine the transmission pattern of Fasciola hepatica. Transmission was found to increase through the pasture season, reaching a peak during November. Overwintering of metacercariae or snail-borne stages was not found to contribute to infections in the year under study. A variety of species of Lymnaea were found to be available in southern Idaho as potential intermediate hosts. The enzyme-linked immunosorbent assay was found to be a good serologic indicator of light infections with F hepatica. The serum gamma-glutamyltransferase activity was not diagnostically significant when the degree of fluke infection was low.     

Heterologous resistance to Fasciola hepatica conferred upon rats by passive transfer of serum from different breeds of sheep

Two experiments compared the protection against oral challenge with 20 metacercariae of Fasciola hepatica conferred on rats by intraperitoneal injection of serum from three breeds of sheep infected with F. hepatica (Barbados Blackbelly, St. Croix, Florida Native). Experiment 1 used serum from sheep 5-6 months of age following two infections of 250 metacercariae each, while Experiment 2 utilized serum collected from the same sheep at 10-11 months of age following either a primary (first exposure) or challenge (after two previous exposures of 250 metacercariae each) infection with 500 metacercariae. Similar numbers of flukes were recovered from rats given either immune or nonimmune (control) serum from each breed of sheep in Experiment 1. In Experiment 2, rats given serum from infected St. Croix sheep had significantly fewer flukes than rats given either control or immune serum from Barbados Blackbelly or Florida Native sheep. There was no significant correlation of fluke counts between individual serum donors (sheep) and serum recipients (rats).     

Host responses during experimental infection with Fasciola gigantica or Fasciola hepatica in Merino sheep I. Comparative immunological and plasma biochemical changes during early infection

This study reports the early biochemical changes in plasma, comparative host-immune responses and parasite recovery data in Merino sheep during the first 10 weeks of infection with Fasciola gigantica and Fasciola hepatica. One group of sheep were uninfected, four groups of sheep received incremental challenge doses of F. gigantica metacercariae (50, 125, 225 and 400, respectively) and the sixth group was challenged with 250 F. hepatica metacercariae. At 10 weeks post infection (wpi), sheep challenged with F. hepatica showed the greatest fluke recovery (mean 119, range 84-166); a significantly higher biomass of parasites recovered (2.5-fold greater than the highest dose of F. gigantica); and a greater mean % parasite recovery (39.3%, range 27-55%) than any group challenged with F. gigantica. Within the groups dosed with F. gigantica a strong dose-dependent response was observed in both fluke recovery and fluke biomass with increasing dose of metacercariae. The mean % parasite recovery of F. gigantica infected groups 1-5 were 26, 23, 26 and 25%, respectively, suggesting a uniform viability of parasite establishment independent of infection dose. At 6 wpi, elevated levels of plasma GLDH were observed in the F. gigantica infected groups compared to the uninfected sheep (p<0.005) whereas the F. hepatica challenged group had four-fold higher levels of GLDH compared to the F. gigantica infected group (p<0.001). Elevated levels of GGT as an indicator of epithelial damage in the bile duct was only seen in the group challenged with F. hepatica at 10 wpi when it rose from below 100 IU/l to approximately 250 IU/l (p<0.0001) whereas no detectable increase in GGT was observed in any of the groups challenged with F. gigantica. The white blood cell response to F. hepatica infection was biphasic with the initial peak at 4 wpi and a second peak at 9 wpi, corresponding to the period of migration of juvenile fluke in the liver and the time when adult flukes are migrating into the bile duct, respectively. This biphasic response was also evident in the changes in the eosinophil counts and serum haemoglobin levels. There was a trend toward higher parasite-specific IgG2 titres in sheep infected with lower worm burdens, suggesting that higher F. gigantica or F. hepatica burdens suppress IgG2 responses. The findings of this study suggest that, in early infection in a permissive host, F. hepatica appears to be more pathogenic than F. gigantica because of its rapid increase in size and the speed of its progression through the migratory phases of its life cycle.     

Fasciola in horses in the Republic of South Africa: a single natural case of Fasciola hepatica and the failure to infest ten horses either with F. hepatica or Fasciola gigantica

The faeces of 11 horses were examined for Fasciola spp. eggs. One of them was positive for Fasciola hepatica, a finding which was confirmed post-mortem. The 10 negative horses were subsequently infested with either F. hepatica or Fasciola gigantica, each animal receiving orally from 500-9,500 metacercariae. No clinical signs were observed and no fluke eggs were detected in the faeces, and neither immature nor adult Fasciola worms were recovered from the horses slaughtered 16-26 weeks post-infestation with F. hepatica and 28-34 weeks post-infestation with F. gigantica, respectively. The results indicate that horses have a high level of resistance to both South African Fasciola spp. Factors which can probably explain the conflicting reports for the prevalences of fascioliasis in horses throughout the world and the varying results obtained in experimental infestations are briefly considered.