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Eurytrematosis is a disease caused by flukes of the genus

Pseudosuccinea columella is considered invasive and has become an important intermediate host of both Fasciola species in many regions of the world. This systematic review assessed the geographical distribution of P. columella, and its implications in the transmission of Fasciola hepatica and Fasciola gigantica, globally. A literature search was conducted on Google Scholar, JSTOR and PubMed databases using Boolean operators in combination with predetermined search terms for thematic analysis. Results show that P. columella has been documented in 22 countries from Europe (3), Africa (8), Oceania (2), North America (3) and South America (6). Furthermore, this snail species has shown to adapt to and inhabit a vast array of freshwater bodies including thermal lakes and ditches with acidic soils. Studies showed that P. columella transmits F. hepatica, with natural and experimental infections documented in sub-Saharan Africa, Europe, South America and North America. Experimental infection studies in Cuba showed the presence of P. columella populations resistant to F. hepatica infection. Furthermore, some populations of this invasive snail collected from F. hepatica endemic locations in Brazil, Venezuela, Australia, South Africa, Colombia and Argentina were found without Fasciola infection. As a result, the role played by this snail in the transmission of Fasciola spp. in these endemic areas is still uncertain. Therefore, further studies to detect natural infections are needed in regions/countries where the snail is deemed invasive to better understand the veterinary and public health importance of this snail species in Fasciola-endemic areas and determine the global dispersion of resistant populations of P. columella.

Fascioliasis is a zoonotic disease caused by liver flukes transmitted by freshwater lymnaeid snails. Donkey and horse reservoir roles have been highlighted in human endemic areas. Liver fluke infection in mules has received very limited research. Their role in disease transmission, epidemiological importance and Fasciola hepatica pathogenicity are studied for the first time. Prevalence was 39.5% in 81 mules from Aconcagua, and 24.4% in 127 from Uspallata, in high-altitude areas of Mendoza province, Argentina. A mean amount of 101,242 eggs/mule/day is estimated. Lymnaeids from Uspallata proved to belong to ribosomal DNA internal transcribed spacer (ITS) markers ITS-1 and ITS-2 combined haplotype 3C of Galba truncatula. These lymnaeids were experimentally susceptible to infection by egg miracidia from mules. Infectivity, number of cercariae/snail and shedding period fit the enhanced F. hepatica/G. truncatula transmission pattern at very high altitude. This indicates that the mule is able to maintain the F. hepatica cycle independently. Individual burdens of 20 and 97 flukes were found. Mule infection susceptibility is intermediate between donkey and horse, although closer to the latter. Anatomo-pathology and histopathology indicate that massive infection may cause mule death. Haematological value decreases of red blood cells, haemoglobin, leucocytes and lymphocytes indicate anaemia and strong immunosuppression. Strongly increased biochemical marker values indicate liver function alterations. The mule probably played a role in the past exchanges with Chile and Bolivia through Mendoza province. Evidence suggests that mules could contribute to the spread of both F. hepatica and G. truncatula to human fascioliasis-endemic areas in these countries.

The enzyme-linked immunosorbent assay (ELISA) technique can play an important role in the early detection of fascioliasis. However, they have some diagnostic limitations, including cross-reaction with other helminths. It seems that the combination of recombinant parasite proteins as antigen can reduce these problems. Hence, the present study was aimed to design and confirm the antigenic recombinant multi-epitope (rMEP) construct of three protein epitopes (linear and conformational B-cell epitopes) of the parasite using immunoinformatic tools. For this purpose, the tertiary structures of Fasciola hepatica cathepsin-L1, saposin-like protein 2 and 16.5-kDa tegument-associated protein were predicted using the I-TASSER server. Validation of the modelled structures was performed by Ramachandran plots. The antigenic epitopes of the proteins were achieved by analysing the features of the IEDB server. The synthesized gene was cloned into the pET-22b (+) expression vector and transformed into the Escherichia coli BL21. Sodium dodecyl sulfate polyacrylamide gel electrophoresis was used to verify and analyse the expression of the rMEP protein. Western blotting was utilized to confirm rMEP protein immunogenicity in two forms, one using an anti-His tag antibody and the other with human pooled sera samples (fascioliasis, non-fascioliasis and negative control sera). Our results demonstrated that the rMEP designed for the three proteins of F. hepatica was highly antigenic, and immune-detection techniques confirmed the antigen specificity. In conclusion, the presented antigenic multi-epitope may be very helpful to develop serodiagnostic kits such as indirect ELISA to evaluate the proper diagnosis of fascioliasis in humans and ruminants.

A cross-sectional socio-economic study of fascioliasis in livestock was carried out at the Livestock Development Cooperation (SODEPA) slaughterhouse in Etoudi, Yaoundé during the periods covering the beginning and late dry season in the northern and eastern regions of Cameroon. A total of 768 cattle and 267 sheep were inspected for the presence of Fasciola species. The overall infection rate in cattle and sheep was 18% (n = 767) and 27% (n = 267), respectively. For the animals that were Fasciola positive, a total of 267.86 kg of liver in cattle was condemned, resulting in a loss of US$1124, while a total of 57 kg of liver was condemned from the sheep population, amounting to a loss of US$114. A total of US$76,097 was determined as losses incurred from condemnation of liver for both cattle and sheep per annum based on the total number slaughtered each year. The findings indicate that fascioliasis is present in cattle and sheep slaughtered in Cameroon and that it causes great economic losses due to condemnation of liver. The local climatic conditions, husbandry systems and the presence of snails (intermediate hosts) are probably the main factors influencing the incidence of the disease and may account for the epidemiological significance found in this study.

An abattoir study was carried out between May and October 2016 to determine the parasitic causes of organ condemnation during meat inspection and to evaluate the attendant financial losses in Fako abattoirs, in the South-West region of Cameroon. Organs (liver, lungs, heart, tongue, kidney, spleen and intestine) were examined at meat inspection following standard procedures and the financial loss was estimated by considering the total weight of condemned organs and the price per kilogram of marketable organs, obtained from the local market. The organs of 1472 cattle were examined, of which 357 (24.38%) were condemned. The organs condemned because of parasitic infestations were the liver (333) and small intestine (24), and the infections were caused by flukes of Fasciola sp. and proglottids of Moniezia sp., respectively. Hydatid cysts and cysticerci were absent. The prevalence of fasciolosis and monieziosis was 22.62% and 1.63%, respectively. Condemnation deprived the region of 665.457 kg of meat, with an associated financial loss of CFA 1,330,902 (USD 2505), during the study period. Parasitic diseases worsen the food insecurity situation as they result in the withdrawal of a considerable amount of meat from the food chain. Fasciolosis, the leading parasitic cause of meat condemnation in Fako, is also zoonotic. It is therefore important that effective control measures are implemented countrywide against this parasitosis.

Trematode infections cause serious economic losses to livestock worldwide. Global production losses due to fasciolosis alone exceed US$3 billion annually. Many trematode infections are also zoonotic and thus a public health concern. The World Health Organization has estimated that about 56 million people worldwide are infected by at least one zoonotic trematode species, and up to 750 million people are at risk of infection. Fasciolosis caused by the flukeFasciola giganticais endemic in Nigeria and is one of the most common causes of liver condemnation in abattoirs. Total cattle losses fromFasciolainfection in Nigeria have been estimated to cost £32.5 million. Other trematode infections of cattle, including paramphistomosis, dicrocoeliasis and schistosomiasis, have all been reported in various parts of Nigeria, with varying prevalence. Most publications on trematode infections are limited to Nigerian local and national journals, with very few international reports. This paper therefore summarized the current data on distribution, control and zoonotic trematode infections in Nigeria and other African countries. We also identified research gaps and made recommendations for future research and areas for funding for policy/planning.

An in vivo study in the laboratory rat model has been carried out to monitor changes to the tegument and gut of adult Fasciola hepatica following treatment with myrrh (‘Mirazid’). Rats infected with the triclabendazole-resistant Dutch isolate were dosed orally with Mirazid at a concentration of 250 mg/kg and flukes recovered 2, 3 and 7 days post-treatment (pt). The flukes were processed for examination by scanning and transmission electron microscopy. A variety of changes to the external surface were observed, culminating in the sloughing of the tegumental syncytium. Internal changes to the syncytium and tegumental cell bodies were more severe and were evident from 2 days pt onwards. Swelling of the basal infolds (leading to flooding of the surface layer) and a decline in secretory body production were the major changes seen. The gastrodermal cells were less severely affected than the tegument, pointing to a trans-tegumental route of uptake for Mirazid by the fluke. Some loss of muscle fibres in the main somatic muscle layers was observed, which may be correlated with the decline in movement of flukes seen at recovery.

Fasciola hepatica is a helminth parasite that causes huge economic losses to the livestock industry worldwide. Fasciolosis is an emerging foodborne zoonotic disease that affects both humans and grazing animals. This study investigated the associations between climatic/environmental factors (derived from satellite data) and management factors affecting the spatial distribution of this liver fluke in cattle herds across different climate zones in three Mexican states. A bulk-tank milk (BTM) IgG1 enzyme-linked immunosorbent assay (ELISA) test was used to detect F. hepatica infection levels of 717 cattle herds between January and April 2015. Management data were collected from the farms by questionnaire. The parasite's overall herd prevalence and mean optical density ratio (ODR) were 62.76% and 0.67, respectively. The presence of clustered F. hepatica infections was studied using the spatial scan statistic. Three marked clusters in the spatial distribution of the parasite were observed. Logistic regression was used to test three models of potential statistical association from the ELISA results using climatic, environmental and management variables. The final model based on climatic/environmental and management variables included the following factors: rainfall, elevation, proportion of grazed grass in the diet, contact with other herds, herd size, parasite control use and education level as significant predictors. Geostatistical kriging was applied to generate a risk map for the presence of parasites in dairy herds in Mexico. In conclusion, the spatial distribution of F. hepatica in Mexican cattle herds is influenced by multifactorial effects and should be considered in developing regionally adapted control measures.

Fascioliasis is an important disease affecting livestock, with great costs to producers worldwide. It has also become a serious issue for human populations in some endemic areas as an emerging zoonotic infection. There are two Fasciola species of liver fluke responsible for this disease, which occur worldwide, Fasciola hepatica and Fasciola gigantica. Identifying these two species on the basis of adult or egg morphology requires specialist knowledge due to the similarity of characters, and may misidentify putative intermediate or hybrid forms. In this study we sequenced the internal transcribed spacer 2 (ITS-2) rDNA of liver flukes collected from multiple species of hosts from seven localities in the Punjab and Baluchistan provinces of Pakistan, to determine the distribution of these two species. All 46 flukes processed in this study, collected from seven sites, showed the rDNA ITS-2 genotype corresponding to F. gigantica, contradicting previous reports, based on adult and egg morphology, that both species are present in Pakistan, with F. hepatica being the more common.

Experimental infections of three EgyptianPseudosuccinea columellapopulations with sympatric miracidia ofFasciolasp., coming from cattle- or sheep-collected eggs, were carried out to determine the capacity of this lymnaeid to support larval development of the parasite. Using microsatellite markers, the isolates of Egyptian miracidia were identified asFasciola hepatica. Apart from being independent of snail origin, prevalences ranging from 60.4 to 75.5% in snails infected with five miracidia ofF. hepaticawere significantly higher than values of 30.4 to 42.2% in snails with bi-miracidial infections. The number of metacercariae ranged from 243 to 472 per cercarial-shedding snail and was independent of snail origin, parasite origin and miracidial dose used for infection. IfP. columellawas subjected to two successive bi-miracidial infections withF. hepatica, prevalence of infection was 63.3%, with a mean of 311 metacercariae per snail. These values were clearly greater than those already reported forRadix natalensisinfected with the same parasite and the same protocol. Successful experimental infection ofP. columellawithF. hepaticasuggests that this lymnaeid snail is an important intermediate host for the transmission of fascioliasis in Egypt.

Although the efficacy of triclabendazole treatment against Fasciola gigantica has been shown before in cattle and buffaloes, there appears to be no published report on the efficacy of triclabendazole in impala or other antelope species. As part of a health monitoring programme at Mulungushi International Conference Centre, Lusaka, Zambia, a coprological examination was undertaken to investigate the helminthological status of captive impala (Aepyceros melampus). Of 39 fresh coprological samples, 46% contained F. gigantica. The source of infection was identified to be a fountain within the grounds of the conference centre. Lymnaea natalensis, collected from the study site, were induced to shed cercariae, and were thus confirmed as the snail intermediate host. In managing this disease, triclabendazole at 6 mg/kg was administered together with the feed bait. Water from the pond was drained; vegetation within it cleared and the pond allowed to dry for 1 week before water was replaced. Three weeks post-treatment, faecal examination revealed that the Fasciola had been cleared. To the best of our knowledge, this is a first case of Fasciola spp. infection reported in captive wild impala in Zambia and provides evidence that triclabendazole may be delivered to free-ranging antelope using medicated bait.

An in vivo study in the laboratory rat model has been carried out to monitor changes to the spermatogenic cells in the testis tubules of adult Fasciola hepatica following treatment with the artemisinins, artemether and artesunate. Rats infected with the triclabendazole (TCBZ)-resistant Sligo isolate were dosed orally with artemether at a concentration of 200 mg/kg and flukes recovered at 24, 48 and 72 h post treatment (pt). Rats infected with the TCBZ-resistant Oberon isolate were dosed orally with artesunate at a concentration of 200 mg/kg and flukes recovered 24, 48, 72 and 96 h pt. The flukes were processed for histological and transmission electron microscope (TEM) examination. Changes to the spermatogenic cells were evident at 24 h pt with artemether. The spermatogonial and spermatocyte cells contained abnormal mitochondria, there were fewer spermatids and spermatozoa in the tubules than normal, and a number of cells showed signs of apoptosis. There was a further decline in cell numbers at 48 h pt and the organization of the spermatocyte and spermatid rosettes was atypical. Sperm formation had become abnormal and those spermatozoa present possessed only a single axoneme. By 72 h pt, the testis tubules were vacuolated and filled with abnormal cells and cell debris. Only spermatogonial cells could be identified and there was widespread evidence of apoptosis in the cells. Distinct cellular changes following artesunate treatment did not become apparent until 48 h pt. The changes seen were similar to those described for artemether, but were generally less severe at matching time-periods. The fine structural changes occurring in the spermatogenic cells were compared to those observed in other cell types and fluke tissues and the overall information was collated to identify the cellular targets for artemisinin action and to establish the time-line for drug action.

Vietnam is recognized to be endemic for fasciolosis. However, most of the available publications have not been published in international journals. This review is based on national and international Vietnamese publications and highlights the current status of fasciolosis in Vietnam. It also provides some information available for neighbouring countries. Updated data on responsible species, distribution, transmission and control aspects are summarized. The central region of Vietnam is reported as being highly endemic for fasciolosis, with a high number of human patients (more than 20,000 in 2011). Fasciola gigantica is reported to be the main species in Vietnam. However, hybrids between F. gigantica and F. hepatica were identified. Both humans and animals are infected by the ingestion of raw vegetables and possibly contaminated drinking water. Three lymnaeid snail species (Austropeplea viridis, Radix auricularia and Radix rubiginosa) may act as intermediate hosts of Fasciola spp. However, due to the likely misidentification of snail species and cercariae during the past decade the critical analysis of published data is difficult. A better understanding of transmission aspects of fasciolosis would allow the implementation of preventive measures of this important neglected zoonotic disease.

The ability of total homogenate (TH) ofFasciola hepaticaconjugated with aluminium hydroxide (alum) or Freund's complete adjuvant (FCA) to protect cattle against experimental fasciolosis was evaluated. Compared with the infected group, the immunized animals with alum-TH and FCA-TH presented a significant reduction in fluke burden (85.9% and 96.8%, respectively), a higher percentage of short-sized worms, a marked reduction in the released eggs in faeces (89% and 57%, respectively), as well as an increased production of specific antibodies before infection. The alum-TH immunized group also showed a significant increase in the antigen-specific proliferation of peripheral blood mononuclear cells (PBMC) as early as 4 weeks before infection. Although both immunized groups (alum-TH and FCA-TH) were able to develop an efficient protective immune response to metacercarial challenge, an earlier PBMC response, lower hepatic damage and less effect on weight gain were found in alum-immunized animals. Therefore, alum is a good candidate for future immunization against bovine fasciolosis.

In vitro trials investigating the effects of albendazole and triclabendazole anthelmintics on the growth profiles of the egg-parasitic fungi Paecilomyces lilacinus and Verticillium chlamydosporium were undertaken. In addition, in vivo trials were conducted in goats fed on millet grain cultures of each fungus and administered albendazole and triclabendazole anthelmintics. In vitro growth revealed V. chlamydosporium to be more sensitive to albendazole compared to P. lilacinus. In contrast, triclabendazole had the least inhibitory effect on in vitro growth of both P. lilacinus and V. chlamydosporium. Similar to albendazole, growth of P. lilacinus was more vigorous at 0.5 ppm concentration of triclabendazole. Efforts to re-isolate these egg-parasitic fungi from faeces of goats fed on fungal millet grain cultures before and following single intraruminal administration of albendazole and triclabendazole showed that P. lilacinus was not able to be re-isolated from the faeces at any sampling period. In contrast, V. chlamydosporium was able to be re-isolated from the faeces at all of the sampling periods except for the samples taken at 8–18 h and 18–24 h after administration of albendazole and triclabendazole, respectively. Lack of fungal activity at these times coincided with peak plasma availability of anthelmintics and suggests faecal levels of drugs were also high at these times and impacted negatively on fungal viability.

The prevalence of Fasciola hepatica infection, in endemic countries, in patients with established cirrhosis is unknown. We hypothesized that, in endemic countries, the presence of fascioliasis may be detected in a serum pool of cirrhotic patients. Forty-four previously stored serum samples of patients with established liver cirrhosis, in the Hospital Nacional Cayetano Heredia in Lima, Peru, were collected from 1998 to 2003 and assessed for hepatitis B, C and fascioliasis antibodies (Fas2 ELISA). Hepatitis B surface antigen (HBsAg) was positive in 8.8% (n = 34), hepatitis B core antibody (anti-HBc) in 32.5% (n = 34), hepatitis C antibodies (anti-HCV) in 9.1% (n = 33), and 9.1% (n = 44) were Fas2 ELISA positive. This disease is an example of an emerging tropical infection which can be present in chronic liver diseases, requiring greater clinician awareness especially in endemic rural areas. Further clinical studies are warranted.

Every month during the year 2006–2007, the 24, 48, 72 and 96 h LC50values of a molluscicide, cypermethrin, were determined for a snailLymnaea acuminata, with concomitant estimation of levels of temperature, pH, dissolved oxygen and carbon dioxide and electrical conductivity, both in control and test water. On the basis of a 24 h toxicity assay, it was noted that LC50values of 10.39, 10.90 and 11.19 mg l− 1during the months of May, June and July, respectively, were most effective in killing the snails, while the molluscicide was least effective in the month of January, when its 24 h LC50was 65.84 mg l− 1.There was a significant positive correlation between LC50of cypermethrin and levels of dissolved O2/pH of water in corresponding months. On the contrary, a negative correlation was observed between LC50and dissolved CO2/temperature of test water in the same months. In order to ascertain that such a relationship between toxicity and abiotic factors is not coincidental, the nervous tissue of the snail was assayed for the activity of acetylcholinesterase (AChE), acid phosphatase (ACP) and alkaline phosphatase (ALP) to sublethal concentrations (40% and 80%) of 24 h LC50during each of the 12 months of the same year. The findings confirmed that abiotic factors indeed influence toxicity of cypermethrin in the snail. A significant positive rank correlation between AChE, ACP and ALP activity did exist following exposure to the corresponding sublethal concentrations. Moreover, there was a maximum inhibition of 61.29 and 76.16% of AChE and ACP, respectively, in snails exposed to 80% of the 24 h LC50in the month of May. A similar treatment caused a maximum inhibition of 70.53% of ALP activity in the month of June. This work shows conclusively that the best time to control the snail population with cypermethrin is during the months of May and June.

This study focuses on the development of fibrosis of the liver of cattle with Fasciola hepatica infection, correlating with the intensity of infection. Animals with an established diagnosis of chronic F. hepatica infection were identified in a slaughterhouse in Lima, Perú. The study included 24 fresh cattle livers from infected animals and two uninfected controls. Tissues were stored at 4°C for approximately 8 h after which they were brought to a necropsy room and examined. Between 9 and 12 biopsies were randomly obtained from each liver. Histological staining of formalin-fixed liver sections with haematoxylin and eosin (H & E) and Masson's trichrome were performed. Liver samples were examined using a pathology protocol that included 30 items. Histopathologically, 16 out of 30 liver specimens (67.6%) showed diffuse fibrotic lesions (cirrhosis) with a mean number of Fasciola of 116 ± 30 (range 4–435). Pathological data were matched to number of adult parasites and presence of cirrhosis after being reviewed by two independent pathologists. There was concordance between the two pathologists (K = 0.72). The group with cirrhosis showed an average of 116 ± 30 adult parasites whereas the group not showing cirrhosis contained 56 ± 28 flukes (P = 0.2). To measure how number of flukes and diagnosis of cirrhosis are related we used Kendall's tau-b coefficient; the correlation was +0.296 (P = 0.04). Receiver Operating Characteristic (ROC) curve results showed that the best point was 38 parasite adults, which had 93.8% sensitivity and 75% specificity. We conclude that as the number of F. hepatica adult forms increases, the likelihood of developing liver fibrosis will also increase in cattle.

A serological and coprological survey of fasciolosis was conducted in bovine hosts from the Sargodha district, Pakistan using excretory–secretory (ES) antigens of Fasciola gigantica from cattle and buffaloes. Livers, faecal and blood samples of 146 cattle and 184 buffaloes were collected from slaughterhouses and examined for the presence of any Fasciola in bile ducts and ova in faeces. Serum was separated. ES antigens were prepared by incubating adult Fasciola in phosphate-buffered saline for 6–8 h and then filtering using a 0.22-μm syringe filter. Checkerboard titration was performed and optimum concentrations of antigen and serum were determined. Sero-prevalence was found to be 50.00 and 38.35% in buffalo and cattle, respectively. Using liver examination as the gold standard, enzyme-linked immunosorbent assay (ELISA) sensitivity was found to be 100% in both buffalo and cattle as compared with that of coprological examination in buffalo (61.79%) and cattle (54.54%). This indigenous ELISA was also highly specific, with values of 96.84 and 98.90% in buffalo and cattle, respectively. Positive predictive values were calculated as 96.74 and 98.21% in buffalo and cattle, respectively, while negative predictive values were 100%. For the validation of indigenous ELISA in field surveys, faecal and blood samples were collected from six sub-districts (tehsils) in the district of Sargodha. Sera were screened for the presence of anti-fasciola antibodies using both the indigenous and commercial ELISA kits. While both kits were equally sensitive, the indigenous ELISA was found to be more specific. The highest prevalence of fasciolosis was found in December, as ascertained using both serological and coprological examination. Significant differences were found in prevalences of fasciolosis in different sub-districts and age groups, together with feeding and watering systems.

We determined the prevalence and seasonality of infections by Fasciola of goats and bovine species (cattle and water buffalo) in Hubei and Anhui provinces of China. Faecal samples were collected at 2- to 3-month intervals from 200 goats in Hubei province and from 152 bovine species in Anhui province. All faecal samples were examined for the presence of parasites. We determined the nucleotide sequences of the first and second internal transcribed spacers (ITS-1 and ITS-2) of the nuclear ribosomal DNA (rDNA) of 39 Fasciola worms from Anhui province. The prevalence of Fasciola infection in goats ranged between 3.5 and 37.0%, with mean eggs per gram (EPG) ranging between 29.0 and 166.0. Prevalence and EPG exhibited downward trends over time with significant differences. The prevalence of Fasciola infection in cattle ranged between 13.3 and 46.2% (mean EPG, 36.4–100.0), and that of water buffalo ranged between 10.3 and 35.4% (mean EPG, 25.0–89.6), with a higher prevalence of infection and EPG from June to October compared with December to March. Analysis of ITS-1 and ITS-2 sequences revealed that F. hepatica and F. gigantica were present in all bovine species of Anhui province and that F. gigantica mainly infected water buffalo. This is the first demonstration of Fasciola infection in Hubei province and detection of F. hepatica and F. gigantica in Anhui province. The present study of Hubei province shows that mass treatment of livestock with closantel sodium injections in April and August/September controlled Fasciola infection effectively.

Over the past decades, various free-living animals (hosts) and their parasites have invaded recipient areas in which they had not previously occurred, thus gaining the status of aliens or exotics. In general this happened to a low extent for hundreds of years. With variable frequency, invasions have been followed by the dispersal and establishment of non-indigenous species, whether host or parasite. In the literature thus far, colonizations by both hosts and parasites have not been treated and reviewed together, although both are usually interwoven in various ways. As to those factors permitting invasive success and colonization strength, various hypotheses have been put forward depending on the scientific background of respective authors and on the conspicuousness of certain invasions. Researchers who have tried to analyse characteristic developmental patterns, the speed of dispersal or the degree of genetic divergence in populations of alien species have come to different conclusions. Among parasitologists, the applied aspects of parasite invasions, such as the negative effects on economically important hosts, have long been at the centre of interest. In this contribution, invasions by hosts as well as parasites are considered comparatively, revealing many similarities and a few differences. Two helminths, the liver fluke,Fasciola hepatica,of cattle and sheep and the swimbladder nematode,Anguillicola crassus,of eels are shown to be useful as model parasites for the study of animal invasions and environmental global change. Introductions ofF. hepaticahave been associated with imports of cattle or other grazing animals. In various target areas, susceptible lymnaeid snails serving as intermediate hosts were either naturally present and/or were introduced from the donor continent of the parasite (Europe) and/or from other regions which were not within the original range of the parasite, partly reflecting progressive stages of a global biota change. In several introduced areas,F. hepaticaco-occurs with native or exotic populations of the congenericF. gigantica, with thus far unknown implications. Over the fluke's extended range, in addition to domestic stock animals, wild native or naturalized mammals can also serve as final hosts. Indigenous and displaced populations ofF. hepatica, however, have not yet been studied comparatively from an evolutionary perspective.A. crassus, from the Far East, has invaded three continents, without the previous naturalization of its natural hostAnguilla japonica, by switching to the respective indigenous eel species. Local entomostrac crustaceans serve as susceptible intermediate hosts. The novel final hosts turned out to be naive in respect to the introduced nematode with far reaching consequences for the parasite's morphology (size), abundance and pathogenicity. Comparative infection experiments with Japanese and European eels yielded many differences in the hosts' immune defence, mirroring coevolution versus an abrupt host switch associated with the introduction of the helminth. In other associations of native hosts and invasive parasites, the elevated pathogenicity of the parasite seems to result from other deficiencies such as a lack of anti-parasitic behaviour of the naïve host compared to the donor host which displays distinct behavioural patterns, keeping the abundance of the parasite low. From the small amount of available literature, it can be concluded that the adaptation of certain populations of the novel host to the alien parasite takes several decades to a century or more. Summarizing all we know about hosts and parasites as aliens, tentative patterns and principles can be figured out, but individual case studies teach us that generalizations should be avoided.

Leucine aminopeptidase (LAP) and cathepsin L1 (CL1) are important enzymes for the pathogenesis and physiology ofFasciola hepatica. These enzymes were analysedin silicoto design a chimeric protein containing the most antigenic sequences of LAP (GenBank; AAV59016.1; amino acids 192–281) and CL1 (GenBank CAC12806.1; amino acids 173–309). The cloned 681-bp chimeric fragment (rFhLAP-CL1) contains 270 bp from LAP and 411 bp from CL1, comprising three epitopes, DGRVVHLKY (amino acids 54–62) from LAP, VTGYYTVHSGSEVELKNLV (amino acids 119–137) and YQSQTCLPF (amino acids 161–169) from CL1. The ~25 kDa rFhLAP-CL1 chimeric protein was expressed from the pET15b plasmid in the Rosetta (DE3)Escherichia colistrain. The chimeric protein rFhLAP-CL1, which showed antigenic and immunogenic properties, was recognized in Western blot assays usingF. hepatica-positive bovine sera, and induced strong, specific antibody responses following immunization in rabbits. The newly generated chimeric protein may be used as a diagnostic tool for detection of antibodies againstF. hepaticain bovine sera and as an immunogen to induce protection against bovine fasciolosis.

Knowledge of the morphological phenotypes of the liver fluke Fasciola hepatica (Trematoda: Digenea) is analysed. The influence of parasite age on its dimensions, the adult fluke growth model, variation in a biometric variable versus time, and variation in a biometric variable versus another biometric variable (allometric model) are revised. The most useful allometric model appears to be (y2m]#x2212;y2)/y2=c [(y1m−y1)/y1]b, where y1=body area or body length, y2=one of the measurements analysed, y1m, y2m=maximum values towards which y1 and y2, respectively, tend, and c, b=constants. A method based on material standardization, the measurement proposal and allometric analysis is detailed. A computer image analysis system (CIAS), which includes a colour video-camera connected to a stereomicroscope (for adult studies) and a microscope (for egg studies), facilitates the processing of digital imaging. Examples of its application for the analysis of the influence of different factors on the liver fluke phenotype are shown using material from the Northern Bolivian Altiplano, where human and domestic animal fascioliasis is caused by F. hepatica only. Comparisons between the development of livestock fluke populations from highlands and lowlands are discussed and the relationships between host species and liver fluke morphometric patterns is analysed.

Fascioliasis is an important trematode infection of herbivores worldwide with increasing evidence of prevalence as a disease of humans. Vaccination studies with purified native and recombinantFasciolaantigens suggest that this approach to diminished morbidity and mortality and reduced transmission is a realistic goal. Among the major potential vaccine candidates are fatty acid binding protein (FABP), cysteine (cathepsin) proteases, haemoglobulin, leucine aminopeptidase, and a saposin-like protein. In the case ofFasciola hepaticaFABP, cross-reaction and cross-protection againstSchistosoma mansoniis an important feature. In addition to protective effects with significant worm burden reductions, some vaccine candidates also have anti-fecundity (smaller flukes), anti-pathology (less liver lesions), and anti-embryonation effects. Optimism is tempered by the fact that fascioliasis in humans is an orphan disease and in need of governmental and foundation support.

Triclabendazole was introduced in the early 1980s for the treatment ofFasciola hepaticainfections in livestock. Due to its high activity against immature flukes, it has become established as the principal anti-fluke drug on the market. More recently, triclabendazole has been used to treat human cases of fascioliasis and is now the drug of choice for this infection, too. Resistance to triclabendazole was first reported in 1995 in a field population in Australia and, since that time, resistant populations have been identified in several countries in Europe. Parallel to the spread of resistance has been a sharp increase in the prevalence of fascioliasis, which has been attributed largely to climate changes. Consequently, farmers are faced with an alarming scenario, as none of the other fasciolicides on the market possess such high activity against the damaging immature stages of fluke. The main aim of this review is to assess current understanding of the mechanism of action of triclabendazole against the fluke and the mechanism by which the fluke has become resistant to it. The use of triclabendazole against animal and human infections is summarized and suggestions are given on ways to deal with resistance. Gaps in the knowledge of various aspects of its use are highlighted and this may serve to open up future research areas.

Considered a secondary zoonotic disease until the mid-1990s, human fascioliasis is at present emerging or re-emerging in many countries, including increases of prevalence and intensity and geographical expansion. Research in recent years has justified the inclusion of fascioliasis in the list of important human parasitic diseases. At present, fascioliasis is a vector-borne disease presenting the widest known latitudinal, longitudinal and altitudinal distribution.Fasciola hepaticahas succeeded in expanding from its European original geographical area to colonize five continents, despite theoretical restrictions related to its biology and in turn dependent upon environmental and human activities. Among the different epidemiological situations, human hypo- to hyperendemic areas, including epidemics, are noteworthy. A global analysis of the distribution of human cases shows that the expected correlation between animal and human fascioliasis only appears at a basic level. Areas presenting very high human prevalences and intensities, especially in children and females, have been recently described. In hypo- to hyperendemic areas of Central and South America, Europe, Africa and Asia, human fascioliasis presents a range of epidemiological characteristics related to a wide diversity of environments. Thus far well-known epidemiological patterns of fascioliasis may not always explain the transmission characteristics in any given area and control measures should consider the results of ecoepidemiological studies undertaken in the zones concerned.

Experimental infections of 1-mm high snails using three populations of Lymnaea (L. glabra, L. ovata and L. truncatula) and a cattle strain of Fasciola hepatica miracidia were carried out under laboratory conditions to determine if the snail species had an effect on the number of free rediae, their growth, and cercarial productivity in relation to each redial category (R1a, R1b, R2a, or R2b/R3a). The total number of rediae ranged from 6.4 to 7.5 per snail. The mean body length of rediae varied from 1–1.2 mm (R1a) to 0.3–0.4 mm (R2b/R3a). The width of the intrapharyngeal lumen also varied from 26.0–38.8 μm to 3.0–4.2 μm, respectively. The redial category had a significant effect on both measurements, whereas snail species only had a significant influence on body length. The mean number of cercariae produced by all living rediae at day 49 post-exposure ranged from 63.0 in L. glabra to 87.2 in L. truncatula. In L. ovata and L. truncatula, 55.8% and 58.6% of cercariae, respectively, were produced by R2a rediae, whereas 53.9% of cercariae in L. glabra were formed by the R1b rediae. When young snails were infected with F. hepatica, the species of snail had an effect on the number of living rediae, their length and their cercarial productivity.

Experimental infections of Lymnaea truncatula with Fasciola hepatica were performed to study the consequences of the presence of predators (sciomyzid larvae or zonitid snails) on the characteristics of larval F. hepatica development in surviving snails. Controls consisted of infected snails that were not subjected to predators. Compared to controls, the survival rate at day 30 post-exposure, the duration of cercarial shedding, and the number of cercariae shed by surviving snails were significantly lower when predators were present in snail breeding boxes, whatever the type of predator used. In contrast, the prevalences of Fasciola infections in snails, and the length of time between exposure and the onset of cercarial shedding showed no significant variation. The progressive development of a stress reaction in surviving snails against predators during the first 30 days of experimental exposure to F. hepatica would influence snail survival during the cercarial shedding period and, consequently, the number of cercariae shed by the snails.

The length and width of 1297 Fasciola hepatica eggs shed in cattle hosts, 337 in sheep and 199 in nutria, were measured from several parts of France. The data were compared with those obtained from other studies in Spain, France (where rats were also investigated), Germany and the Netherlands. One way analysis of variance and discriminant analysis were used to assess differences between host origins. The distribution of length and width of eggs were analysed using skewness and kurtosis Fisher coefficients. The eggs recovered from sheep, cattle, rodents and lagomorphs were different in size: the eggs found in rodents (length L × width W in μm: 8592) and lagomorphs (L × W in μm: 9100) were smaller than those found in sheep and cattle (L×W in μm: 10,000). These morphological differences in F. hepatica eggs were host-induced in rats (L×W in μm: 9709 in cattle to 8949 in rats) and rabbits (L×W in μm: 9709 in cattle to 8432 in rabbits). These differences in size of eggs might correspond to their being less able to develop into miracidia in less frequent hosts such as rodents and rabbits.

IgG and IgM antibody responses to fluke cysteine proteinases in Paragonimus ohirai- and Fasciola sp.-infected rats were followed by means of cystatin capture ELISA using fluke excretory-secretory products for 10 weeks after infection. The specific IgG antibodies were detectable at week 2 postinfection in all P. ohirai-infected and some Fasciola-infected rats. Levels of specific IgG antibodies increased rapidly between week 2 and 6, and slightly thereafter, in both infected groups. From week 3, specific IgG antibody levels were higher in Fasciola-infected than P. ohirai-infected rats. Sera from infected rats did not react with heterologous cysteine proteinases throughout the infection periods. In both infected groups, the kinetic patterns of specific IgM antibody responses were similar to those of specific IgG antibody responses although the ELISA levels of the IgM antibody responses were much lower. In abnormal infections with P. ohirai metacercariae x-irradiated at 2 krad, the specific IgG antibodies were detectable at week 2 postinfection with similar ELISA values to normal P. ohirai infection, but thereafter increased little. In infections with P. westermani, for which the rat is not a suitable host, even stunted worms induced a comparable specific IgG antibody response, although the response was lower than in normal infections with P. ohirai. These results indicate that cystatin capture ELISA can distinguish clearly between Paragonimus and Fasciola infections which show immunodiagnostic cross-reactivity and is useful even in the early stages of the infection and in the infection of unsuitable hosts.

Fasciola hepatica secretes proteolytic enzymes to aid it to penetrate and migrate through the host tissues. Two of these proteinases have been purified and shown to be cathepsin L-like, and are termed, CL1 (27.5 kD) and CL2 (29 kD). The immunogenicity of these proteinases was investigated over the course of an experimental infection and following drug treatment. Four groups of rabbits were studied: group 1: orally infected with 50 metacercariae; group 2: infected and treated 8 weeks after infection; group 3: infected, treated at week 8 and reinfected at week 13 and group 4: non-infected control group. Sera were collected weekly from each group until week 20 postinfection. CL1 and CL2 were incubated with the different sera and then analysed by gelatin substrate polyacrylamide gel electrophoresis (GS-PAGE). Analysis of groups 1, 2 and 3 showed that CL1 and CL2 neutralizing antibodies appear at week 5 post-infection. In group 1, these remained throughout the 20 weeks of infection. In group 2, neutralizing antibodies disappeared at week 13, that is, 5 weeks after anti-Fasciola treatment. In group 3, CL1 and CL2 neutralizing antibodies disappeared at week 13 but reappeared by week 15, that is 2 weeks after reinfection. Pooled sera from group 4, showed no inhibitory capacity. ELISA results using CL1 and CL2 as antigen, correlate very well with the inhibitiory time course observed by GS-PAGE. These results suggest that purified cathepsin Ls are antigenic molecules recognized early in the infective process and capable of inducing a specific humoral response, strong enough to neutralize, at least partially, their enzymatic activity.

Fascioliasis is an important food-borne parasitic disease caused by the two trematode species, Fasciola hepatica and Fasciola gigantica. The phenotypic features of fasciolid adults and eggs infecting buffaloes inhabiting the Central Punjab area, Pakistan, have been studied to characterize fasciolid populations involved. Morphometric analyses were made with a computer image analysis system (CIAS) applied on the basis of standardized measurements. Since it is the first study of this kind undertaken in Pakistan, the results are compared to pure fasciolid populations: (a) F. hepatica from the European Mediterranean area; and (b) F. gigantica from Burkina Faso; i.e. geographical areas where both species do not co-exist. Only parasites obtained from bovines were used. The multivariate analysis showed that the characteristics, including egg morphometrics, of fasciolids from Central Punjab, Pakistan, are between F. hepatica and F. gigantica standard populations. Similarly, the morphometric measurements of fasciolid eggs from Central Punjab are also between F. hepatica and F. gigantica standard populations. These results demonstrate the existence of fasciolid intermediate forms in endemic areas in Pakistan.

Laboratory investigations on three aquatic populations of Galba truncatula, originating from the Peruvian Altiplano and French Massif Central, were carried out during three successive snail generations to determine if these populations might be successfully used for the metacercarial production of Fasciola hepatica under experimental conditions. High numbers of surviving snails at day 30 post-exposure (>70%), high prevalences of F. hepatica infections (>60%), and prolonged productions of cercariae for a mean period of 35 to 47 days were observed in the three populations, whatever the snail generation. In the Peruvian population, metacercariae of F. hepatica significantly decreased in numbers from a mean of 251 in the parent snails to 124 per snail in the F2 generation, whereas no significant variation was observed in the two French populations. As these aquatic snails rarely emerged out of water, the use of these populations for the commercial production of F. hepatica metacercariae was of great interest, because the daily time spent watching the breeding boxes of snails was clearly shorter, thereby reducing the cost of producing metacercariae compared with using amphibious snails reared with romaine lettuce.

The 14.5 kDa fatty acid binding protein (FABP) was isolated from the crude extract of adultFasciola giganticaworms. Polyclonal anti-FABP IgG was generated in rabbits immunized with prepared FABP antigen. Sandwich enzyme-linked immunosorbent assay (ELISA) was applied to detect coproantigen in stools and circulatingFasciolaantigen (CA) in sera of 126 water buffaloes by using purified and horseradish peroxidase (HRP)-conjugated anti-FABP IgG. Sandwich ELISA sensitivity was 96.97% and 94.95%; while specificity was 94.12% and 82.35% for coproantigen and CA detection, respectively. However, sensitivity and specificity of the Kato–Katz technique was 73.74% and 100%, respectively. The diagnostic efficacy of sandwich ELISA was 96.55% and 93.1% for coproantigen and CA detection, respectively. In contrast, the diagnostic efficacy of the Kato–Katz technique was 77.59%. In conclusion, these results demonstrate that the purified 14.5 kDa FABP provides a more suitable antigen for immunodiagnosis of early and current bubaline fascioliasis by using sandwich ELISA.

Quantities of trace elements including copper, zinc, cobalt, manganese and iron were investigated in the liver tissue of rats at the acute or chronic stages of fascioliasis following treatment with zinc-copper hydroxochloride mixed crystals. Oral dosing (with food) of zinc–copper mixed crystals to healthy rats increased zinc and copper levels in the liver and decreased the iron content compared with controls. Manganese and cobalt levels did not change significantly. Significant reductions in all trace elements except manganese occurred in the liver of rats with acute or chronic fascioliasis. Manganese levels were slightly increased in rats at the acute stage and slightly decreased in rats at the chronic stage of fascioliasis. The application of mixed zinc–copper crystals at the acute or chronic stages of fascioliasis lead to a restoration of zinc and copper levels and a slight reduction in the iron levels in liver tissue of rats. The beneficial effects of applied salts were more apparent in rats chronically infected withFasciola hepatica.