Prevalence of Fasciola hepatica in the intermediate host Lymnaea truncatula detected by real time TaqMan PCR in populations from 70 Swiss farms with cattle husbandry

Bovine fasciolosis is an economically important parasitic disease. Quantitative real time PCR was utilized to determine the prevalence of Fasciola hepatica in the snail intermediate host Lymnaea truncatula from 70 selected, infected Swiss cattle farms, and to gain information on the infection risk to the definitive host. Snails from 130 habitats (36 streams, 21 wells, 24 drainage ditches, 33 spring swamps, 14 reeds, 1 drainage shaft and 1 pond) originating from 71 dairy cow pastures, 39 pastures for young stock, 14 hay fields and 6 dry cow pastures were collected. Of these, 51 populations were found to be infected with F. hepatica. A total of 4733 snails were examined of which 331 were infected (7.0%). The numbers of snails collected from different sites ranged from 1 to 159 snails. Clustering of infection in snails was found on the farm of origin with a mixed logistic model with random effects. The risk of infection of L. truncatula with F. hepatica was significantly higher in populations originating from spring swamps, wells and reeds compared to populations from streams. In addition the risk of snail infection was significantly lower in populations collected in young stock and dry cow pastures compared to dairy cow pastures. The greater the population size collected from a habitat also increased the risk of an individual snail being infected.     

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.     

An interactive map to assess the potential spread of Lymnaea truncatula and the free-living stages of Fasciola hepatica in Switzerland

The intermediate host of Fasciola hepatica is Lymnaea truncatula in Switzerland. The snail and the free-living stages of the parasite require a moderate climate and moisture for survival, reproduction, and transmission. In Switzerland, these conditions are present in many regions, resulting in a mean prevalence of bovine fasciolosis from 8.4 to 21.4%. An interactive map was created in order to demonstrate the relative risk of transmission by modelling the environmental conditions that promote the survival and reproduction of the larval stages of the parasite and the parasite's intermediate host. The map is based on temperature and rainfall data, soil conditions including ground water and forest cover in Switzerland. Extensive information on the free-living stages of F. hepatica and the intermediate host L. truncatula and how the development of these are affected by these environmental factors was used to create the interactive risk map.     

PCR diagnosis of Fasciola hepatica in field-collected Lymnaea columella and Lymnaea viatrix snails

Fasciolosis, caused by the trematode Fasciola hepatica, is a zoonosis of economic importance in livestock that is emerging as a chronic disease in humans. The intermediate hosts are lymnaeid snails, in which diagnosis of infection is traditionally based on cercarial shedding, tissue sectioning and crushing. We developed a PCR assay for the sensitive and specific detection of F. hepatica in field-collected Lymnaea sp. snails. A primer pair was designed to amplify a 405 bp fragment of the cytochrome c oxidase subunit 1 gene of F. hepatica. The PCR assay showed a limit of detection of 10 pg of genomic F. hepatica DNA. No cross-reactions were observed with samples from other related trematode species or from the snail hosts Lymnaea columella and Lymnaea viatrix. DNA sequencing of the amplicon showed 100% homology with F. hepatica, and 75-89% homology with other trematodes on regions that did not include the entire set of primers. Two samples from Argentina were analysed. For snails in sample 1 (n = 240), identified as L. columella, the infection rate was 17.5 and 51.3% by direct examination and PCR, respectively. For snails in sample 2 (n = 34), identified as L. viatrix, the infection rate was 2.9 and 61.8% by direct examination and PCR, respectively. Differences in infection rates between these diagnosis methods were significant for both samples. Our PCR technique showed to be effective for detecting specific F. hepatica infections of low intensity in the intermediate host, and hence it could be used to study the epidemiological situation in a given area, as well as to assess host suitability for the parasite.     

Survival of snails and characteristic lesions of Fasciola hepatica infection in four European species of Lymnaea.

The survival of snails and characteristic organ lesions were investigated in four different age categories (new-born, 1 and 2 mm shell height, adults) and in four species of Lymnaea (L. glabra, L. palustris, L. peregra ovata, L. truncatula) infected by Fasciola hepatica. Mortalities were higher in infected groups on Days 30 and 45 post-infection (+42% and +45%, respectively) than in controls. Lesions depended on the species and age of the Lymnaea. Epithelial necrosis of the kidney was recorded in L. glabra, L. palustris and L. p. ovata, whereas intralamellar deposits were found in the same organ of L. truncatula. All categories of L. truncatula had evolutive infections (snails with living parthenitae); the young snails of other species became infected, but the adults did not, as estimated at Day 45 post-infection. The lesions were related to the intensity of evolutive infections: necrosis of the digestive gland was found in moderate to low infections and, conversely, gonadal atrophy with epithelial necrosis was found in highly infected snails.     

Fasciola hepatica and Paramphistomum daubneyi: changes in prevalences of natural infections in cattle and in Lymnaea truncatula from central France over the past 12 years

A retrospective study was carried out over a 10- to 12-year period to analyse the changes in prevalences of natural fasciolosis and paramphistomosis among cattle and snails in central France, and to determine the causes which had induced these changes. The prevalences of natural fasciolosis in cattle increased from 1990 to 1993 (13.6% to 25.2%) and diminished afterwards up to 1999 (at 12.6%). Those of natural paramphistomosis showed a progressive increase between 1990 and 1999 (from 5.2 to 44.7%). The prevalences of natural infections and the numbers of free rediae counted in the snails (Lymnaea truncatula) infected with F. hepatica did not show any significant variations over time. By contrast, the prevalences of natural paramphistomosis in snails significantly increased from 1989 to 1996 and remained afterwards in the same range of values (3.7-5.3%), while the number of free rediae significantly increased up to 2000 (from a mean of 6.5 to 13.8 rediae per infected snail, respectively). Three hypotheses may explain the increase of paramphistomosis in cattle and snails: a better quality of diagnosis for the detection of P. daubneyi eggs in veterinary analysis laboratories, the use of specific molecules in the treatment of cattle fasciolosis since 1993, and the lack of an effective treatment up to now against cattle paramphistomosis. Since the objective of most farmers in central France is to obtain the highest antiparasitic efficiency with a single treatment of cattle per year, it is reasonable to assume that the prevalence of bovine paramphistomosis will continue to increase in the future.     

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 occurrence of the snail Lymnaea columella on grazing areas in New South Wales and studies on its susceptibility to Fasciola hepatica

Field surveys were carried out in the Lismore and Casino area for the presence of fresh water snails potentially responsible for the transmission of trematodes in ruminants. Although the North American snail, Lymnaea columella has previously only been reported from metropolitan areas, large populations of the snail were found east of Lismore. Natural infection with F. hepatica was detected in some of the snails and laboratory studies showed that this snail species was highly susceptible to infection with miracidia of F. hepatica and produced viable metacercariae. Three species of planorbid snails, Helicorbis australiensis, Pygmanisus pelorius and Gyraulus gilberti were also found. The first 2 planorbid species were infected with paramphistomid cercariae.     

Suitability of six lymnaeid snails for infection with Fasciola hepatica

The suitability of Fossaria (Bakerilymnaea) cubensis and Pseudosuccinea columella from Louisiana as intermediate hosts for Fasciola hepatica was compared to P. columella and Stagnicola elodes from Ann Arbor, MI, S. attenuata from Hidalgo, Mexico, Lymnaea gedrosiana from Iran and L. natalensis from Senegal. P. columella from LA was shown to be a more suitable host (51.3% became infected) than F. (B.) cubensis (15.2% and 26.4% of two populations became infected). The infection rate for P. columella from MI, was 50% and for L. gedrosiana was 32.5%, whereas L. natalensis, S. elodes and S. attenuata were refractory. F. (B.) cubensis and P. columella have some degree of suitability as intermediate hosts for F. hepatica under laboratory conditions, but field observations of their habitat in southern LA and characteristic management of cattle indicate that the former snail is more important as an intermediate host in this enzootic area. Tissue sections of suitable snails had few histopathological effects but physical damage caused by rediae was pronounced, mainly in the digestive gland and in the mantle.     

Prevalence of Paramphistomum daubneyi infection in cattle in central France

Investigations at the slaughterhouse of Limoges (Central France) were performed in 1994-1996 to determine the prevalence of Paramphistomum daubneyi infection in cattle. In 1994/1995, higher prevalences of P. daubneyi were recorded in May, October, and January. In 1996, smaller variations were recorded, and prevalence in April and May was higher. The prevalence of P. daubneyi in summer was lower. Breed and age of cattle were not significantly associated to P. daubneyi infection, but females were significantly more infected than males. P daubneyi infection was related to that of Fasciola hepatica, but not to that of Dicrocoelium lanceolatum.     

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.     

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.     

Distribution and habitats of the snail Lymnaea truncatula, intermediate host of the liver fluke Fasciola hepatica, in South Africa

This paper focuses on the geographical distribution and habitats of Lymnaea truncatula, the intermediate, snail host of the liver fluke, Fasciola hepatica, as reflected by the 723 samples in the database of the National Freshwater Snail Collection, Potchefstroom, South Africa. The 221 different loci (1/16-degree squares) on record reflect an extensive but discontinuous distribution, except in Lesotho and in parts of the Mpumalanga, Gauteng and North West provinces of South Africa. Although recorded from 12 different types of waterbody, it was mostly (42.0%) recovered from swamps. Most samples (45.8%) were collected in habitats with slow-flowing water. A muddy substratum was recorded for 62.5% of the samples. Most samples (86.3%) were collected in habitats with a mean annual air temperature of 10-20 degrees C, and more than 69% came from localities with a mean annual rainfall of 600-900 mm. An integrated decision tree constructed from the data indicated that temperature and types of waterbody play a decisive role in determining the presence of L. truncatula in a given area. A temperature index calculated for all mollusc species ranked L. truncatula second in a total of 53 species according to its association with low temperatures. It remains to be established whether its distribution is indeed discontinuous, and whether its preference for a particular habitat, amphibious habits and ability to aestivate could have resulted in some populations having been overlooked during surveys.     

Bovine fascioliasis in the Manawatu: epidemiology and farmer awareness

The prevalence of Fasciola hepatica was found to be 5.1% in slaughtered cattle originating from the Palmerston North Veterinary District. Approximately 4% of beef breeding, 7% of dairy and 21% of drystock herds showed evidence of infection. Of 34 dairy herd and 14 beef breeding herd owners, 0 and 21% respectively were aware of the presence of fluke on their properties. Distribution of affected dairy farms conformed with the ecological requirements of the intermediate host Lymnaea columella. The high prevalence of infection in drystock herds is consistent with the extensive movements known to occur into these herds from liver fluke-affected parts of the country. In view of the expanding geographical range of Fasciola hepatica and its intermediate host Lymnaea columella, farmers should be alerted to the potential risks that result from the introduction of infected cattle to properties which contain suitable habitats for the snail.     

Soil type and the distribution of Lymnaea truncatula

Habitat distribution of the snail Lymnea truncatula, the intermediate host of the liver fluke Fasciola hepatica, was related to a soil map in a part of Dyfed in South Wales. There was a close association with wet soils in general but, within these areas, habitats were concentrated in very wet localities of restricted size, such as ditches and seepages. Vegetation characteristic of these habitats is described. Soil maps could be a useful tool for the veterinary officer, agricultural adviser and research worker in identifying fluke-prone land.     

The uptake and distribution of radioselenium in the larvae of Fasciola hepatica and its snail host Lymnaea columella

Radiolabeled metacercariae of Fasciola hepatica were obtained in vivo by incubating infected Lymnaea columella snails with 20 muCi radioselenomethione (75Se-M) per snail in 5 ml of water for 5 h, or in vitro by incubating a batch of unlabeled F. hepatica metacercariae with 75Se-M for 24 h. Radioassay showed that only 5% of the 75Se-M was incorporated into maritas (juvenile flukes) from the in vivo labeled metacercariae. The inner cyst wall of in vivo labeled metacercariae contained 46% of the total activity, of which 21% was dissolved in the excysting medium. The outer, tan-colored cyst wall contained 49% of the radioactivity. Through diffusion/attachment, maritas from in vitro labeled metacercariae could occasionally be labeled with 0.4% of the total radioactivity. However, the activity was lost after inoculation into the body of mice. The outer and inner cyst walls of in vitro labeled metacercariae contained 92% and 7.6%, respectively, of the total activity. Microautoradiography demonstrated that 75Se-M was evenly distributed in the body of marita and the cyst wall of inner and outer layers from an in vivo labeled metacercaria. A 9 X 4 micron rectangularly-shaped aggregate of Ag degree grains was present on the outer periphery of the inner cyst wall. Microautoradiography of in vitro labeled metacercariae demonstrated a significant concentration of Ag degree grains on the cyst walls. The ventral plug contained fewer Ag degree grains per unit area compared to the other portion of the inner cyst wall. Uptake and distribution of 75Se-M in the snail host were also studied. It appeared that rediae and cercariae tended to concentrate the label in the foot, the mantle and the digestive gland. Little or no radioactivity was present in the areas where F. hepatica larvae were not found.     

An epidemiological study of Fasciola hepatica in the Netherlands

Summary

Transmission of F.hepatica under natural conditions was analysed in a three year programme. The variables used were the indirect haemagglutination (IHA) technique, worm establishment in tracer lambs and the population dynamics, infection rate and shedding pattern of Lymnaea truncatula.

It is concluded that fluke eggs, infected snails and metacercariae on herbage can survive the winter in the Netherlands. Metacercarial availability was positively correlated to the amount of rainfall in the grazing period. The role of developed eggs that survive the winter is important, because this results in earlier infections in the herd.

The use of the serological diagnosis method IHA is important to detect F. hepatica infection in an early stage. Use of cellophane paper on floats is a useful method for determining the shedding pattern of cercariae from L. truncatula. It is concluded that collection of metacercariae on cellophane floats, inventarization of L. truncatula and its infection level are useful tools for the prediction of liverfluke infections.     

The distribution of Fasciola hepatica in Queensland, Australia, and the potential impact of introduced snail intermediate hosts

A survey was conducted to establish the distribution of the liver fluke, Fasciola hepatica, in the state of Queensland, Australia, and to evaluate the impact of the introduced snail intermediate hosts, Pseudosuccinia columella and Austropeplea viridis. Serum samples from a total of 5103 homebred cattle in 142 beef herds distributed throughout the state and 523 pooled milk samples from dairy herds from the state's major dairying regions were tested for antibodies to F. hepatica by ELISA. Snails were collected on infected properties around the limits of the F. hepatica distribution. F. hepatica infection was detected in 44 dairy herds and two beef herds. The distribution of infected herds indicates that F. hepatica is established only in southeast Queensland. The distribution there was patchy but the parasite was more widespread than suggested by an earlier survey. The predominant intermediate host species found along the northern limit of the distribution was P. columella. We conclude that the introduction of P. columella and A. viridis has not yet had a major impact on the distribution of F. hepatica in Queensland. However, the presence of P. columella, which is much more adaptable to tropical habitats than the native intermediate host, Austropeplea tomentosa, at the northern limit of the F. hepatica distribution suggests that there is potential for the parasite to expand its range.     

The effect of cyclic temperatures on the growth of Fasciola hepatica and Lymnaea viatrix

The experiment aimed to measure the effect of constant and variable temperatures on the growth of Lymnaea viatrix snails, on the development of a Peruvian isolate of Fasciola hepatica eggs and on the development of F. hepatica in the snails. This was carried out by cultivating infected and uninfected snails and fluke eggs in artificial, temperature controlled chambers. L. viatrix snails were found to develop at a rate dependent on environmental temperature, but developed at least as well under conditions of varying temperature as at the same mean constant temperature. F. hepatica eggs held at constant or varying temperatures, developed at a rate comparable to other reports. However, eggs developing at varying temperatures appeared to have reduced hatchability. Parasite development within the snails was slow, though within the limits calculated from the literature, and varying temperature did not appear to reduce development compared to constant temperatures.     

Geographical distribution and habitat preferences of the invader freshwater snail species Lymnaea columella (Mollusca: Gastropoda) in South Africa

The present geographical distribution of Lymnaea columella, as recorded in the National Freshwater Snail Collection, is described and discussed. It appears that L. columella is the most successful colonist of all the freshwater snail species in South Africa, and, together with Bulinus tropicus and Lymnaea natalensis, it forms the most widely distributed freshwater snail species in the region. Data regarding the 2,341 discovery sites of L. columella recorded by collectors during surveys are analysed. L. columella cluster mainly in rivers or streams with perennial, slow flowing, or stagnant freshwater with many plants and a mud substrate. L. columella is regarded as a more effective intermediate host for the transmission of fascioliasis than the indigenous snail species. The economical implications of this are discussed briefly in the light of its success as an invader species.