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Background: The potential molluscicidal extracts, obtained from indigenous plants Cannabis sativa , Acacia nilotica , and Tinospora cordifolia , were tested for toxicity against freshwater pulmonate snail Lymnaea acuminata , an intermediate host of Fasciola hepatica . The organic extracts had a significant effect on young snails. Materials and Methods: All organic extracts and column-purified fractions gave median lethal concentrations (19-100.05 mg/L; 24 h) that fell well within the threshold level of 100 mg/L, set for a potential molluscicide by the World Health Organization. Results: The toxicity of T. cordifolia stem acetone extract (96 h LC 50 : 16.08 mg/L) was more pronounced compared with C. sativa leaf ethanol extract (96 h LC 50 : 16.32 mg/L) and A. nilotica leaf ethanol extract (96 h LC 50 : 24.78 mg/L). β-caryophyllene, gallic acid, and berberine were characterized and identified as active molluscicidal components. Co-migration of β-caryophyllene (retardation factor [R f ] 0.95), gallic acid ( Rf 0.30), and berberine (R f 0.23) with column-purified parts of Cannabis sativa , Acacia nilotica , and Tinospora cordifolia on thin-layer chromatography demonstrates same R f value, that is, 0.95, 0.30, and 0.23, respectively. Conclusion: This study indicates that these extracts thus represent potential plant-derived molluscicides that are worthy of further investigations.

The use of natural products for disease control is a promising approach to solving the problem of drug resistance. The aim of the research reported here was to evaluate the fasciolicidal and anti-Clostridium novyi type B activities of propolis administered orally to sheep infected with Fasciola gigantica and C. novyi type B. Sheep infected with both pathogens were divided into two groups: an infected treated group and an infected non-treated group. The treatment was oral administration of 50 mg propolis extract/kg daily for 15 days. The body weight of the sheep, fecal egg counts of F. gigantica, serum levels of F. gigantica IgG, concentrations of cytokines (IL-2, IL-10, and IL-17), and bacterial counts of C. novyi were evaluated. Following treatment, the sheep had increased body weight and a significant decrease in the egg count, which was reduced by 54.54% at 15 days post treatment. The level of anti- Fasciola IgG increased, whereas levels of IL-2, IL-10, and IL-17 decreased in propolistreated sheep. Treatment of sheep with propolis produced a significant reduction in fecal count of C. novyi, from 8 × 10 9 to 3 × 10 3 colony units per gram at 15 days post treatment. This research highlights the therapeutic potential of Egyptian propolis extract as a treatment against F. gigantica and C. novyi type B infections, and investigated its mode of action through its effect on some cellular and humoral responses in sheep with both infections.

Fasciolosis is a neglected tropical disease caused by the liver fluke Fasciola gigantica. The absence of successful vaccine and emerging resistance in flukes against the drug of choice, triclabendazole, has necessitated the search for alternatives including phyto-therapeutic approaches. Curcumin and thymoquinone, the active ingredients of Curcuma longa and Nigella sativa plants respectively, were first screened for their binding affinity with Glutathione-S-transferase (GST) molecule through in silico molecular docking followed by in vitro treatment of worms with varying concentrations of the test compounds. The in silico molecular docking of curcumin and thymoquinone with sigma GST revealed strong hydrogen bonding as well as hydrophobic interactions with high fitness scores but showing inter-specific differences. The in vitro treatment of F. gigantica worms with both curcumin and thymoquinone resulted in a significant increase in the generation of reactive oxygen species (ROS) whereas the level of reduced glutathione, a primary redox regulator, was found to be significantly decreased (p < 0.05). The two compounds not only inhibited the GST activity, which is an important detoxification enzyme and also a key drug/vaccine target for the control of fasciolosis but also significantly inhibited the activity of antioxidant enzymes glutathione peroxidase and glutathione reductase that are vital in maintenance of redox homeostasis. The immunohistochemistry performed using anti sigma GST polyclonal antibodies revealed that both the compounds used in the present study significantly reduced immunofluorescence in the vitellaria, developing eggs present in the ovary and the intestinal caecae indicating inhibition of GST enzyme in these regions of the worms. Further, following treatment with curcumin and thymoquinone, chromatin condensation and DNA fragmentation was also observed in F. gigantica worms. In conclusion, both curcumin and thymoquinone generated oxidative stress in the worms by production of ROS and significantly inhibiting their antioxidant and detoxification ability. The oxidative stress along with induction of apoptotic like events would compromise the survival ability of worms within the host. However, further studies are required to establish their anthelmintic potential alone and in combination with the commonly used anthelmintic drugs under in vivo conditions., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Fascioliasis is a neglected zoonosis with major public health implications in humans. Although triclabendazole (TCBZ) is the drug of choice, there are records of TCBZ failure worldwide. TCBZ-resistant fascioliasis is treated with alternative approved drugs including nitazoxanide (NTZ), with varying levels of efficacy. Data on NTZ efficacy after TCBZ failure in Egypt is scarce. This study evaluated the efficacy of NTZ in cases of TCBZ failure during an outbreak of fascioliasis in Assiut governorate of Upper Egypt., Methodology/principal Findings: This prospective study included 67 patients from the outpatient clinic in Manfalout locality of Assiut governorate with clinical manifestations of acute fascioliasis. These included high eosinophilia (> 6% eosinophils in peripheral blood), positive anti-Fasciola antibodies, and hepatic focal lesions (HFL) or ascites on abdominal ultrasound or computed tomography. All patients initially received TCBZ at recommended doses. Patients were followed up after 1 month to assess response. According to the responses, patients were categorized as non-responders and responders. The non-responders received a trial of NTZ and were re-assessed for response based on clinical manifestations, eosinophil count, and abdominal ultrasound. Patients not responding to NTZ received additional doses of TCBZ. One month after initial TCBZ treatment, 37 patients responded well to TCBZ, while 30 patients failed to respond with persistence of fever, abdominal pain, high eosinophilia, and HFL. Most non-responders were male (56.7%); females predominated among TCBZ responders (62.2%). The mean age of the non-responders was relatively lower, at 20.57 ± 14.47 years (p = 0.004). Following NTZ therapy, HFL disappeared in 9/30 (30%) patients and eosinophil counts normalized in only 2 (6.7%) patients, indicating an overall efficacy of 36.6%. The remaining cases received additional doses of TCBZ with complete clinical, biochemical, and radiological resolution., Conclusions/significance: Nitazoxanide was partially effective in TCBZ failure in acute human fascioliasis in Upper Egypt. Further studies with larger samples are highly encouraged and further research is urgently needed to find new therapeutic alternatives to TCBZ., Competing Interests: The authors have declared that no competing interests exist.

An in vivo study was carried out to investigate the ultrastructural effects of triclabendazole (TCBZ) on immature Fasciola gigantica in a goat model. Five goats were infected with an oral gavage of 150 metacercarial cysts of F. gigantica and anthelmintic treatment occurred at 4 weeks post infection with an oral dose of 10 mg/kg. They were euthanized at 0 (untreated), 24, 48, 72 and 96 h post treatment (h pt). Juvenile flukes were recovered from each of the goat's liver and processed for transmission electron microscopy (TEM). The untreated control flukes showed normal ultrastructure and no apparent changes were observed at 24 h pt. At 48 h pt, moderate levels of disruption were observed to the tegument and minor changes to the sub-tegument which included widespread blebbing and disruption of apical tegumental membrane, swollen mitochondria, reduced number of secretory bodies, swelling of basal infolds leading to severe vacuolation, and relatively mild disruption to the subtegumental muscle fibres, parenchyma and tegumental cells, whereas the gastrodermal cells appeared less affected. By 72 h pt, sloughing of the tegumental syncytium was evident leading to the exposure of the basal lamina and the disruption was severe in the subtegument too. At 96 h pt, the flukes were totally devoid of tegument and the disruption was extremely severe, distorting the ultrastructure of the entire fluke's body. The results of the present study revealed that the flukes showed time-dependent progressive disruption to the internal tissues which became increasingly severe over time pt. This is the first study to detail the time-scale and impacts on ultrastructural morphology of the in vivo TCBZ treatment of the immature tropical liver fluke, Fasciola gigantica., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Fasciolosis, caused by the liver fluke Fasciola gigantica, and paramphistomosis are widespread in cattle in Tanzania, and the use of trematocides is encouraged by the Government livestock extension officers. However, reduced efficacy of oxyclozanide against Fasciola gigantica and amphistomes (rumen flukes), and albendazole against F. gigantica, has been reported in some regions. This study was conducted to assess the efficacy of different trematocides against F. gigantica and amphistome infections in cattle at Iringa Rural and Arumeru Districts., Methods: Cattle found with concurrent infection of F. gigantica and amphistomes were randomly grouped into six experimental groups. One control group was left untreated while five treatment groups were treated with one of five trematocides that include: albendazole, nitroxynil, oxyclozanide, closantel and triclabendazole. Post-treatment faecal sample collection was done on the day of treatment and again at 7, 14 and 28 days, from each cattle. The samples were processed by Flukefinder® method to recover and identify eggs. Assessment of the efficacy of the trematocides against F. gigantica and amphistomes was conducted using faecal egg count reduction (FECR) tests., Results: The findings of the present study in both districts indicate that nitroxynil, oxyclozanide, closantel and triclabendazole are effective against patent F. gigantica infection, as the calculated FECR% for each trematocide was 100% by day 14 post-treatment. However, albendazole found to have reduced efficacy of against F. gigantica, as FECR% was 49% in Arumeru District and 89% in Iringa Rural District by day 14 post-treatment. Oxyclozanide was the only trematocide found to be effective against amphistomes with FECR of 99%., Conclusions: Albendazole had reduced efficacy against F. gigantica in cattle in Arumeru and Iringa Rural Districts, Tanzania. The reduced efficacy was prominent in Arumeru, where cattle are commonly treated with anthelmintics, than in Iringa Rural, where cattle are seldom treated.

Fascioliasis is caused by the helminth parasites of genus Fasciola. Thioredoxin glutathione reductase (TGR) is an important enzyme in parasitic helminths and plays an indispensable role in its redox biology. In the present study, we conducted a structure-based virtual screening of natural compounds against the Fasciola gigantica TGR (FgTGR). The compounds were docked against FgTGR in four sequential docking modes. The screened ligands were further assessed for Lipinski and ADMET prediction so as to evaluate drug proficiency and likeness property. After refinement, three potential inhibitors were identified that were subjected to 50 ns molecular dynamics simulation and free energy binding analyses to evaluate the dynamics of protein-ligand interaction and the stability of the complexes. Key residues involved in the interaction of the selected ligands were also determined. The results suggested that three top hits had a negative binding energy greater than GSSG (-91.479 KJ · mol -1 ), having -152.657, -141.219, and -92.931 kJ · mol -1 for compounds with IDs ZINC85878789, ZINC85879991, and ZINC36369921, respectively. Further analysis showed that the compound ZINC85878789 and ZINC85879991 displayed substantial pharmacological and structural properties to be a drug candidate. Thus, the present study might prove useful for the future design of new derivatives with higher potency and specificity., (© 2017 Wiley Periodicals, Inc.)

Background: Curcumin and nisin have been widely reported for their antibacterial and anticancer potency. However, their therapeutic applications are hampered by several factors, which necessitate their development into nanosize ranges for improved delivery and activities. Their incorporation into a single nanosynthesized form may suggest desirable efficacy on parasites. The aim of the study was to assess the ovicidal activity of the curcumin-nisin polylactic acid (PLA) entrapped nanoparticle on the Fasciola eggs and its reproductive toxicity., Methods: The nanoparticle was formulated by double emulsion method. The eggs of the adult Fasciola spp. were exposed to different concentrations (0.3125-5 mg/mL) of the nanoparticle to monitor hatchability. Mice were exposed to 0.5 mL of the formulated drug at varying concentrations (10-20 mg/kg) and then sacrificed for sperm morphology assay., Results: The mean particle size, polydispersity index, and drug entrapment efficiency of the formulated drug were 288.4±24.3 nm, 0.232, and 51.7%, respectively. The highest nanoparticulate concentration (5 mg/mL) showed the least percentage egg hatching (41.7%) compared with the other treatment groups and positive control (albendazole) (45.1%). The aberrations observed in sperm cells were not concentration-dependent and no significant differences were observed in the mean aberrations between the nanoparticulate drug-exposed groups and the negative control (p>0.05)., Conclusions: The results confirmed the ovicidal activity of the curcumin-nisin nanoparticulate drug against the Fasciola species. The formulation also showed no toxicity to sperm cells. More robust studies on anti-fascioliasis activity of the drug on adult Fasciola spp. and in vivo and in vitro toxicity studies are recommended.

Schistosomes are blood-dwelling trematodes with global impact on human and animal health. Because medical treatment is currently based on a single drug, praziquantel, there is urgent need for the development of alternative control strategies. The Schistosoma mansoni genome project provides a platform to study and connect the genetic repertoire of schistosomes to specific biological functions essential for successful parasitism. G protein-coupled receptors (GPCRs) form the largest superfamily of transmembrane receptors throughout the Eumetazoan phyla, including platyhelminths. Due to their involvement in diverse biological processes, their pharmacological importance, and proven druggability, GPCRs are promising targets for new anthelmintics. However, to identify candidate receptors, a more detailed understanding of the roles of GPCR signalling in schistosome biology is essential. An updated phylogenetic analysis of the S. mansoni GPCR genome (GPCRome) is presented, facilitated by updated genome data that allowed a more precise annotation of GPCRs. Additionally, we review the current knowledge on GPCR signalling in this parasite and provide new insights into the potential roles of GPCRs in schistosome reproduction based on the findings of a recent tissue-specific transcriptomic study in paired and unpaired S. mansoni. According to the current analysis, GPCRs contribute to gonad-specific functions but also to nongonad, pairing-dependent processes. The latter may regulate gonad-unrelated functions during the multifaceted male-female interaction. Finally, we compare the schistosome GPCRome to that of another parasitic trematode, Fasciola, and discuss the importance of GPCRs to basic and applied research. Phylogenetic analyses display GPCR diversity in free-living and parasitic platyhelminths and suggest diverse functions in schistosomes. Although their roles need to be substantiated by functional studies in the future, the data support the selection of GPCR candidates for basic and applied studies, invigorating the exploitation of this important receptor class for drug discovery against schistosomes but also other trematodes.

At present, there are no medicinal plant extracts currently available for treatment and control of fasciolosis. The present work could provide, for the first study, conclusions on the in vitro fasciolicidal properties of the ethanol extract of Terminalia catappa L. (TcCE) leaves against adult Fasciola gigantica after incubation with RPMI-1640 medium containing the TcCE at various concentrations and times when compared with triclabendazole (TCZ). The relative motility and survival index values of the TcCE-treated flukes decreased at a more rapid rate than the TCZ-treated flukes. The death of the parasites was observed after exposed to TcCE at 3 h incubation with 400, 800 and 1000 µg mL-1, and at 6 h incubation in 100 and 200 µg mL-1. Vacuolization, blebbings and partial disruption on the parasites' tegument were observed by light microscopy. When examined by scanning electron microscopy, TcCE caused similar tegumental alterations in the parasites as those observed in TCZ treatment but with larger damage at comparative incubation periods, consisting of swelling, blebbing, disrupted blebs, loss of spines, leading to the erosion, lesion and eventual disruption of the total tegument. Therefore, the TcCE may exert its fasciolicidal effect against F. gigantica by initially causing the tegumental alteration.

Helminth infections are the cause of morbidity in Cambodian cattle but other factors such as nutritional deficiencies and concurrent diseases may enhance the effects of parasites. The present study aimed to investigate the impact of anthelmintic treatment, feed supplementation, or both on gastrointestinal strongyle (GIS) and trematode infections as well as on morbidity parameters in Cambodian village cattle. At the beginning of the dry season, cattle populations in six villages were randomly assigned to a group: (A) receiving anthelmintic treatment (ivermectin+clorsulon) at week 0; (P) feed pellet supplementation during week 0-13 or both (AP). On five visits (week 0-29), faecal and blood samples were obtained for parasitological examination and haematocrit determination, respectively. Body condition (BCS), hind quarter fouling (HQFS), diarrhoea (DS), and conjunctiva colour (FAMACHA © ) were scored and heart girth circumference was determined. To investigate the impact of treatment over time (week 0-29), a mixed model was used with treatment, time, and their interaction as fixed effects, and animal and village as random factors. At baseline, the proportion of GIS positive animals was high (67.9%), whereas trematode infections were low (Paramphistomum: 8.8%; Fasciola: 2.6%). Very thin to emaciated cattle (BCS 1-2) were more prevalent (11.4%) and FAMACHA © scores of ≤3 or below (65.8%) less prevalent than in an earlier study in the region. A Time ⨯ Treatment interaction was present for faecal egg counts (FEC) of GIS, GIS prevalence (both p<0.0001), PCV (p=0.0034), DS (p=0.0086) and HQFS (p=0.0241). For GIS FEC, treatment groups differed at a specific time point, with levels of treatment group P being higher than in A at week 6 (p=0.0054). For Paramphistomum prevalence as well as FAMACHA © scoring, heart girth and BCS, the interaction between treatment and time was not significant, yet, time in itself had a significant impact on all (p<0.0001). The beneficial effects of protein supplementation were unclear from the current study., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Fasciolosis an economically important global disease of ruminants in the temperate and tropical regions, caused by Fasciola hepatica and F. gigantica, respectively, also poses a potential zoonotic threat. In India alone it causes huge losses to stakeholders. Anthelmintics including triclabendazole have been used to control this menace but the emerging resistance against the available compounds necessitates identification of novel and alternative therapeutic measures involving plant derived natural compounds for their anthelmintic potential. Thymoquinone (T) and curcumin (C), the active ingredients of Nigella sativa and Curcuma longa respectively have been used as antiparasitic agents but the information on their flukicidal effect is very limited. Adult flukes of F. gigantica were in vitro exposed to different concentrations of thymoquinone and curcumin separately for 3h at 37+ 1°C. A significant (p<0.05) reduction in the worm motility at 60 μM concentration of both T and C was observed though all the worms remained alive after 3h exposure, whereas the effect on egg shedding was statistically insignificant. Pronounced tegumental disruptions and erosion of spines in the posterior region and around the acetabulum was evident. A significant (p<0.05) decrease in glutathione-S-transferase and superoxide dismutase activity and reduced glutathione (GSH) level was observed, while protein carbonylation increased differentially. A significant inhibition of CathepsinL (CatL) gene expression in thymoquinone treated worms was also evident. Further, in silico molecular docking of T and C with CatL revealed a stronger interaction of curcumin with the involvement of higher number of amino acids as compared to thymoquinone that could be more effective in inhibiting the antioxidant enzymes of F. gigantica. It is concluded that both the compounds understudy will decrease the detoxification ability of F. gigantica, while inhibition of CatL will significantly affect their virulence potential. Thus, both thymoquinone and curcumin appeared to be promising anthelmintic compounds for further investigations., Competing Interests: The authors have declared that no competing interests exist.

Fasciolosis is an important cattle and human disease caused by Fasciola hepatica and Fasciola gigantica. One of the possible methods to control this problem is to interrupt the life cycle of Fasciola by killing its larva (redia and cercaria) in host snail. Molecular identification of cercaria larva of F. gigantica was done by comparing the nucleotide sequencing with adult F. gigantica. It was noted that nucleotide sequencing of cercaria larva and adult F. gigantica were 99% same. Every month during the year 2011-2012, in vivo treatment with 60% of 4 h LC50 of phyto cercaricides citral, ferulic acid, umbelliferone, azadirachtin and allicin caused significant inhibition of acetylcholinesterase (AChE) and cytochrome oxidase activity in the treated cercaria larva of F. gigantica. Whereas, activity of both enzymes were not significantly altered in the nervous tissues of vector snail Lymnaea acuminata exposed to same treatments. Maximum reduction in AChE (1.35% of control in month of June) and cytochrome oxidase (3.71% of control in the month of July) activity were noted in the cercaria exposed to 60% of 4 h LC50 of azadirachtin and allicin, respectively., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Reports of resistance to triclabendazole (TCBZ) among fluke populations have increased in recent years. Allied to this, there has been a rise in the prevalence of the disease, which has been linked to climate change. Results from questionnaire surveys conducted in Northern Ireland (NI) in 2005 (covering the years 1999-2004) and 2011 (covering the years 2008-2011) have provided an opportunity to examine the extent to which fluke control practices have changed over a prolonged time-frame, in light of these changes. A number of differences were highlighted. There was a significant shift away from the use of TCBZ over time, with it being replaced largely by closantel. The timing of treatments had moved earlier in the year, perhaps in response to climate change (and an altered pattern of disease). In relation to the frequency of drug treatments, there were no major changes in the overall pattern of drug treatments between the two survey points, although on both occasions approximately one-third of flock owners gave more than 3 treatments per year to ewes. In lowland areas in 2011, flock owners were rotating drug classes more often (each year and at each treatment) than in 2005, whereas in upland areas, flock owners were rotating less often and more were not rotating at all. Between 2005 and 2011, the percentage of flock owners giving quarantine treatments to bought-in stock had halved, to a very low level (approximately 10%). Using data from a complementary TCBZ resistance survey (Hanna et al., 2015), it has been shown that the way in which data are selected and which efficacy formula is applied can influence the calculation of drug efficiency and impact on diagnosis of resistance., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Fasciolosis is a food borne zoonosis, caused by the digenetic trematode Fasciola. Freshwater lymnaeid snails are the intermediate host of the trematodes. Chlorophyllin, a semi-synthetic derivative of chlorophyll and its formulations obtained from freeze dried cow urine (FCU) had their toxicity tested against redia and cercaria larvae of F. gigantica. The larvicidal activity of chlorophyllin and its formulations were found to depend on both, time and concentration used against the larvae. Toxicity of chlorophyllin + FCU (1:1 ratio) in sunlight against redia larva (8 h LC50: 0.03 mg/mL) was more pronounced than using just chlorophyllin (8 h LC50: 0.06 mg/mL). Toxicity of chlorophyllin + FCU in sunlight against redia (8 h LC50: 0.03 mg/mL) was higher than against cercaria (8 h LC50: 0.06 mg/mL). The larvicidal activity of chlorophyllin in sunlight (redia/cercaria larvae: 8 h LC50: 0.06 mg/mL) was more pronounced than under laboratory conditions (redia: 8 h LC50: 22.21 mg/mL/, cercaria 8 h LC50: 96.21 mg/mL). Toxicity of FCU against both larvae was lower than that of chlorophyllin and chlorophyllin + FCU. Chlorophyllin and its formulations + FCU were 357.4 to 1603.5 times more effective against redia/cercaria larvae in sunlight than under laboratory conditions. The present study has shown that chlorophyllin formulations may be used as potent larvicides against fasciolosis.

Aim of present study was to screen medicinal plants for flukicidal activity in vitro to develop alternative sources of treatment for trematodes infection. For this purpose, crude methanolic extracts (CME) of Cymbopogn jwarancusa and Conyza canadensis were prepared and live adult flukes viz; Fasciola gigantica, and Paramphistomum cervi isolated from liver and bile ducts of slaughtered buffalo were subjected to different drug concentrations as well as positive and negative control. Motility inhibition and paralysis leading to the death of parasites was considered as flukicidal activity of plants extracts. The results revealed that CME of C. jwarancusa and C. canadensis showed significant (P<0.05) flukicidal activity compared to positive control. Also there was a significant effect of different concentrations (P<0.05) and exposure of time on the flukes (P<0.05). Furthermore, ED50 for C. jwarancusa and C. canadensis against F. gigantica were 13.1 and 41.4 mg/ml, respectively. In the case of P. cervi, it was 10.8 and 29.0 mg/ml. It can be concluded that both tested plants showed greater flukicidal activity as compared to positive control with Albendazole till the 8(th) hr. These potent plants needs further studies invivo to elucidate their mode of action.

Toxicity of chlorophyllin against redia and cercaria larvae of Fasciola gigantica was studied under irradiation of visible light. Highest and lowest toxicity of chlorophyllin against both larvae was noted under red (redia - 8 h LC₅₀ 7.88 mg/10 mL and cercaria - 11.99 mg/10 mL) and green (redia - 8 h LC₅₀ 32.12 mg/10 mL and cercaria - 8 h LC₅₀ 43.80 mg/10 mL) light irradiation respectively. The highest toxicity of chlorophyllin under red light irradiation against redia (8h LC₅₀ 7.88 mg/10 mL)/cercaria (8h LC₅₀ 11.99 mg/10 mL) was followed by white (8 h LC₅₀ redia - 20.48 mg/10 mL, 8 h LC₅₀ cercaria - 18.0 3mg/10 mL), blue (8 h LC₅₀ redia - 33.10 mg/10 mL/ 8 h LC₅₀ cercaria - 19.98 mg/10 mL) and yellow (8 h LC₅₀ redia - 23.87 mg/10 mL/ 8 h LC₅₀ cercaria - 23.48 mg/10 mL). Chlorophyllin treatment in darkness (control I) and without treatment of chlorophyllin, while all other conditions were same as treatment group (control II) caused no mortality of redia/cercaria larva. Chlorophyllin might be a promising new safe strategy to replace synthetic larvicide in fasciolosis control programme., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Fasciolosis caused by Fasciola spp. is considered the most important helminth infection of ruminants in tropical countries. Anthelmintic resistance has become a global concern. This study compared the efficacy of the commonly used anthelmintics, determined the toxicity level and any indication of resistance. Thirty two water buffaloes naturally-infected with Fasciola spp. were used to determine the efficacy of triclabendazole (TBZ), albendazole (ABZ), and bromofenofos (BRO) using Fecal Egg Count Reduction Test (FECRT). To test the toxicity of the drugs given, serum glutamic-pyruvic transaminase (SGPT) was evaluated before and within one week after treatment. One dose administration of ABZ registered an efficacy of 79.17%, 73.33% for TBZ and 70.83% for BRO. Efficacy in two dose- treatment group was 83.33% for both BRO and ABZ, and 90.00% for TBZ. Two dose-treatment was effective for TBZ (90%), ineffective for BRO and ABZ. SGPT levels were not significantly different between pre-treatment and post- treatment across all treatments. Giving one or two doses of anthelmintics, at one month interval, does not increase the efficacy of the three drugs tested. The study also implies that anthelmintic resistance may have developed in the animals.

Triclabendazole (TCBZ), the anthelmintic drug active against both mature and immature liver flukes, was used to investigate the effect of in vivo treatment on the tegumental surface of juvenile Fasciola gigantica. Five goats were infected with 150 F. gigantica metacercariae each by oral gavage. Four of them were treated with single dose of TCBZ at 10mg/kg at four weeks post-infection. They were euthanized at 0 (untreated), 24, 48, 72 and 96h post treatment. Juvenile flukes were manually retrieved from the goat livers and processed for scanning electron microscopy. In control flukes, the anterior region was adorned with sharply pointed spines projecting away from the surface, while in the posterior region, spines become shorter and narrower, loosing serration and with the appearance of distinct furrows and papillae. The dorsal surface retained the same pattern of surface architecture similar to that of ventral surface. Flukes obtained from 24h post-treatment did not show any apparent change and were still very active. However, there were limited movements and some blebbing, swelling, deposition of tegumental secretions and some flattening displayed by the flukes of 48h post-treatment. All the worms were found dead 72h post-treatment and showed advanced level of tegumental disruptions, consisting of severe distortion of spines, sloughing off the tegument to expose the basal lamina, formation of pores and isolated patches of lesions. By 96h post-treatment, the disruption was extremely severe and the tegument was completely sheared off causing deeper lesions that exposed the underlying musculature. The disruption was more severe at posterior than anterior region and on ventral than dorsal surface. The present study further establishes the time-course of TCBZ action in vivo with 100% efficacy against the juvenile tropical liver fluke., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The effect of plumbagin (PB, 5-hydroxy-2-methyl-1,4-naphthoquinone) against newly excysted juveniles (NEJs) and 4-weeks-old immature parasites of Fasciola gigantica were compared with triclabendazole (TCZ). The anthelmintic efficacy of 1, 10 and 100μg/ml of PB or TCZ following incubation in vitro for 1-24h was compared using a combination of relative motility (RM), survival index (SI) and larval migration inhibition (LMI) assays for parasite viability. The RM and SI values of the PB-treated group decreased at a more rapid rate than the TCZ-treated group. For NEJs, the decreased RM values were first observed at 1h incubation with 1μg/ml PB, and 90% of flukes were killed at 24h. In contrast, in TCZ-treated groups a 10-fold higher concentration of TCZ (10μg/ml) resulted in only 9% dead parasites after 24h incubation. In 4-weeks-old juvenile parasites, PB reduced the RM value at 10μg/ml with 100% of flukes dead after 3h, while TCZ decreased RM values at the concentration of 100μg/ml but with only 5% of flukes killed at 24h. NEJs treated with PB exhibited 88%, 99% and 100% of LMIs at the concentrations of 1, 10 and 100μg/ml, respectively. NEJs incubated with TCZ have an LMI of only 32% at the highest concentration of 100μg/ml. Similarly PB had a significantly greater killing of immature 4weeks juvenile stages than TCZ at all concentrations; however, 4-weeks-old juvenile parasites were more resistant to killing by PB or TCZ at all concentrations when compared to NEJs. Further studies were carried out to investigate the alterations of the parasite tegument by scanning electron microscope (SEM). PB caused similar tegumental alterations in 4-weeks-old juveniles as those observed in TCZ treatment but with greater damage at comparative time points, comprising of swelling, blebbing and rupture of the tegument, loss of spines, and eventual erosion, lesion and desquamation of the total tegument. These data indicate that PB had a greater fasciolicidal effect against immature stages of F. gigantica parasites than TCZ and warrant further studies for use as a potential new anthelmintic against Fasciola infections., (Copyright © 2013 Elsevier Inc. All rights reserved.)

In eastern and southern part of India Dregea volubilis (Family Asclepediaceae) is widely used as anthelmintic in traditional system of medicine. The present study was conducted to evaluate the fasciocidal activity of the methanol extract of D. volubilis leaves (MEDV) and to observe the drug's effect on organisms through scanning electron microscopic (SEM) study. Live parasites (Trematode: Fasciola gigantica) were collected in 0.9% phosphate-buffered saline from the bile ducts of buffalo. Those were incubated in the said media at 37 ± 1 °C either as control, or with MEDV at 5, 10, 25, 50 and 100 mg/ml as test groups or with albendazole at 10mg/ml as standard group. The efficacy of the extract was determined on the basis of paralysis (temporary loss of spontaneous movement of the organisms) and/or death of the liver flukes. Death was confirmed when the organisms lost their motility permanently and their motility could not be revived even when vigorously shaken or dipped in warm water. MEDV at all concentration effectively paralyzed first and then killed the liver flukes (p < 0.001). Maximum fasciocidal activity was found with concentration of 100 mg/ml at 38.83 ± 3.41 min. Through SEM study, severe damages were observed in both the suckers as well as on the tegumental surfaces of the treated liver flukes. The study confirmed the fasciocidal activity of the MEDV., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Fascioliasis is a common parasitic disease in livestock in China. However, human fascioliasis is rarely reported in the country. Here we describe an outbreak of human fascioliasis in Yunnan province. We reviewed the complete clinical records of 29 patients and performed an epidemiological investigation on the general human population and animals in the outbreak locality. Our findings support an outbreak due to Fasciola gigantica with a peak in late November, 2011. The most common symptoms were remittent fever, epigastric tenderness, and hepatalgia. Eosinophilia and tunnel-like lesions in ultrasound imaging in the liver were also commonly seen. Significant improvement of patients' condition was achieved by administration of triclabendazole®. Fasciola spp. were discovered in local cattle (28.6%) and goats (26.0%). Molecular evidence showed a coexistence of F. gigantica and F. hepatica. However, all eggs seen in humans were confirmed to be F. gigantica. Herb (Houttuynia cordata) was most likely the source of infections. Our findings indicate that human fascioliasis is a neglected disease in China. The distribution of triclabendazole®, the only efficacious drug against human fascioliasis, should be promoted.

Human fascioliasis has the widest latitudinal, longitudinal and altitudinal distribution of any vector-borne disease, yet only 3 cases have been reported from South Africa, the last in 1964. We report 2 cases from the same geographic area associated with local consumption of watercress, suggesting an endemic focus.

Objective: To evaluated the histological changes within the tegument of adult Fasciola gigantica (F. gigantica) that led to the gross changes that were visible externally., Methods: The effects of oleoresin extract of myrrh (Mirazid(®)), myrrh volatile oil and triclabendazole sulphoxide (reference drug) on the tegumental structure of adult F. gigantica following treatment in vitro had been determined by light microscopy., Results: The internal changes in the tegument observed in this study were compatible with surface changes seen in the previous scanning electron microscopy study, using the same drugs. The swelling of tegumental syncytium was a particular feature of their action, but its level was much greater with myrrh volatile oil, in which vacuolization of the tegument and loss of spines were observed., Conclusions: The present study demonstrated the fasciocidal properties of Mirazid(®) oleoresin extract, and it might be possible to reinforce its fasciocidal activity by increasing its content of myrrh volatile oil.

Objective: To evaluate the effects of Mirazid(®) and myrrh volatile oil on adult Fasciola gigantica (F. gigantica ) under laboratory conditions., Methods: The effects of oleoresin extract of myrrh (Mirazid(®)) and myrrh volatile oil on the surface morphology of adult F. gigantica following treatment in vitro had been determined by scanning electron microscopy. The results were compared with those observed in the fluke tegument following incubation in triclabendazole sulphoxide (TCBZ-SO), active form, (Fasinex(®), Ciba-Geigy)., Results: Observations of the efficacy of Mirazid(®) oleoresin extract and myrrh volatile oil indicated that both products showed dose-dependent anthelmintic efficacy. The anterior half of the fluke was consistently more severely affected than the posterior half. The surface changes induced by Mirazid(®) oleoresin extract were less severe than those observed after exposure to either myrrh volatile oil or TCBZ-SO. Flukes showed swelling after these treatments, but its level and blebbing were much greater with myrrh volatile oil; in which patches of tegumental sloughing were observed in the apical cone and the posterior mid-body region of flukes. This was not observed after treatment with Mirazid(®) oleoresin extract., Conclusions: The comparatively more disruption, observed in myrrh volatile oil exposed specimens, compared to that exposed to Mirazid(®) oleoresin extract might suggest that the anthelmintic activity of Mirazid(®) oleo resin extract was attributed to its content of volatile oil. So, increasing the concentration of myrrh volatile oil in Mirazid(®) might possibly help to developing its anthelmintic activity.

The in vitro effect of artesunate (ATS) on the 3-week-old juveniles of Fasciola gigantica was compared with triclabendazole (TCZ) by incubating the parasites in M-199 medium containing the drugs at concentrations of 20, 40, and 80 μg/ml for 1, 3, 6, 12, and 24h. The anthelmintic activities of these drugs were evaluated based on the relative motility value (RM) and the alterations of the tegument as observed by scanning (SEM) and transmission (TEM) electron microscopy. The RM values of TCZ-treated flukes decreased significantly from 6 to 24h for all dosages. For ATS-treated flukes, RM value decreased markedly from 12 to 24h, but the rates of decline were less than TCZ at the same doses. When observed by SEM, the tegument showed similar sequence of morphological changes after treatments with both drugs, comprising of swelling of tegumental ridges, followed by blebbing and later rupturing of the blebs, leading to erosion and lesion, and disruption of the tegument. When examined by TEM, ultrastructural changes in the tegument and associated structures after treatments with TCZ and ATS were similar which comprised of swelling, blebbing of the tegument, dilation of basal infoldings, and depolymerization of the microtrabecular network. After a longer incubation time, the tegument was completely sloughed off and the tegument cell bodies became necrotic. Additionally, in ATS-treated flukes, mitochondria showed severe swelling, rupturing of outer membrane, and their interior filled with flocculent materials., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background: Fascioliasis is an emerging zoonotic disease of considerable veterinary and public health importance. Triclabendazole is the only available drug for treatment. Laboratory studies have documented promising fasciocidal properties of the artemisinins (e.g., artemether)., Methodology: We carried out two exploratory phase-2 trials to assess the efficacy and safety of oral artemether administered at (i) 6×80 mg over 3 consecutive days, and (ii) 3×200 mg within 24 h in 36 Fasciola-infected individuals in Egypt. Efficacy was determined by cure rate (CR) and egg reduction rate (ERR) based on multiple Kato-Katz thick smears before and after drug administration. Patients who remained Fasciola-positive following artemether dosing were treated with single 10 mg/kg oral triclabendazole. In case of treatment failure, triclabendazole was re-administered at 20 mg/kg in two divided doses., Principal Findings: CRs achieved with 6×80 mg and 3×200 mg artemether were 35% and 6%, respectively. The corresponding ERRs were 63% and nil, respectively. Artemether was well tolerated. A high efficacy was observed with triclabendazole administered at 10 mg/kg (16 patients; CR: 67%, ERR: 94%) and 20 mg/kg (4 patients; CR: 75%, ERR: 96%)., Conclusions/significance: Artemether, administered at malaria treatment regimens, shows no or only little effect against fascioliasis, and hence does not represent an alternative to triclabendazole. The role of artemether and other artemisinin derivatives as partner drug in combination chemotherapy remains to be elucidated.

Triclabendazole is the drug of choice against Fasciola infections in humans and animals. However, parasite resistance against triclabendazole is spreading in veterinary field, and there are no drugs of comparable activity currently available for the treatment and control of fascioliasis. The efficacy of a new rhodanine derivative Ro-354 against adult Fasciola gigantica in vitro was investigated. One hour post incubation, scanning and transmission electron microscopic examination revealed an evident disruption of the tegument of F. gigantica as blebbing, swelling and furrowing. Moreover, an increase in severity of tegumental damage as sloughing and absence of spines was observed. In conclusion, Ro-354 shows potent activity against F. gigantica in vitro, and, the authors recommend carrying out more studies to detect its efficacy in vivo.

A total of 60 patients with schistosomiasis (40), fascialiosis (15) and heterophyiasis (5) were selected Beni-Sweif and Mansoura Districts and subjected to history taking, clinical examination, Kato thick smear, sedimentation and hatching test (for schistosomiasis cases) at the beginning of the study, 2 & 3 months after treatment with Oleo-resin of Myrrh (Mirazid) in a dose of 10 mg/kg/day for 6 consecutive days an hour before breakfast. The results showed a significant improvement in symptoms with minimal negligible or no side effects. The cure rates, 2 & 3 months after treatment were 80.7%% & 11.8%% for schistosomiasis, 93.3% & 6.6% for fascioliasis, and 100% for heterophyiasis. The clinical picture of schistosomiasis before treatments were easy fatigability, anorexia, nausea, vomiting, epigastria pain, abdominal distention, right upper guardant pain, colicky abdominal pain, left upper and/or lower guardant pain, abdominal rumbling, dysentery, diarrhea, rectal bleeding, constipation, and alternating bowel habit. Those of fascioliasis were abdominal distention, dripping of saliva, right upper guardant, colicky abdominal pain, weight loss, easy fatigability, intermittent jaundice, anorexia, nausea, vomiting, epigastria, left upper and/or lower quadrant pain, right layer quadrant pain, loin pain, abdominal rumbling, diarrhea, constipation, and alternating bowel habit. The safety and efficacy of C. molmol extract in treating heterophyiasis (100%), fascioliasis (100%) and schistosomiasis (92.5%) were documented.

Objective: To evaluate the in vitro effects of different concentrations of ivermectin and/or artemether on Fasciolagigantica worms and to study the parasitological changes and tegumental alterations using scanning electron microscopy (SEM)., Methods: Fasciola gigantica worms were incubated in vitro for 24 and 48 h with three concentrations of either ivermectin or artemether (10, 20 and 50 microg/ml) or both in half concentration of either (5, 10 and 25 microg/ml)., Results: Exposure of Fasciola worms to 25+25 microg/ml of combined drug regimens or to 50 microg/ml of either ivermectin or artemether for 48 h led to 100%, 41.7% and 75% worm killing which were accompanied by a significant reduction in egg laying capacity and significant increase in dead eggs maximally recorded in combined drug regimens. SEM of the flukes incubated for 48 h with combined drug regimens showed maximal tegumental disruption with swelling of the worm body, roughness, blebbing, sloughing and complete loss of spines. Disruption to the tegument of the flukes induced by artemether was more than that of ivermectin., Conclusions: Artemether alone or combined with ivermectin in half doses had potent fasciocidal activities. Besides, half doses of combined drug regimens had higher ovicidal effects than each drug alone. In vivo studies are recommended to explore the efficacy of combined regimens against Fasciola infection., (Copyright 2009 Elsevier Inc. All rights reserved.)

Effects of essential oil of Allium sativum (garlic) and Piper longum (Indian long pepper) were evaluated on muscular activity of whole Fasciola gigantica and its strip preparation. The whole flukes and longitudinal strip preparations of the flukes were isometrically mounted to record the spontaneous muscular activity (SMA) and to evaluate effects of cumulative doses (0.1, 0.3, 1.0 and 3.0mg/ml) of the plant essential oils. Whole flukes and the strip preparations exhibited continuous SMA without any significant difference in its baseline tension, frequency and amplitude for 2h. Essential oil of A. sativum produced significant reduction in the frequency and the amplitude of the SMA of whole fluke at 1 and 3mg/ml concentrations. It caused complete paralysis of the fluke after 15 min of administration of 3mg/ml concentration. Similar to whole fluke, essential oil of A. sativum (3mg/ml) also produced flaccid paralysis in the strip preparations of the flukes. Essential oil of P. longum firstly induced marked excitatory effect and then there was flaccid paralysis of the whole fluke following 15 min exposure at 3mg/ml concentration. Complete flaccid paralysis of the strip preparation was also ensued after 15 min of administration of 3mg/ml concentration of P. longum. In both the essential oils, the whole fluke and strip preparations did not recover from paralysis following 2-3 washes. In conclusion, the observations demonstrated irreversible paralytic effect of essential oils of A. sativum and P. longum on F. giganticain vitro which might possibly help to developing herbal-based anthelmintic.

Myrrh is an herbal product that has been used since ancient ages for traditional medication and other purposes. The revolution of myrrh as an antiparasitic agent in Egypt began in the 1990s through scientific evidence-based research. The human trematode infections in Egypt were the main focus of research with stories of success and disagreement, at times. The present paper reviewed the antiparasitary activity of myrrh with stress on its possible mode of action, its safety, efficacy and effectiveness on trematode infections in experimental studies and clinical trials in Egypt as well as its molluscicidal effects on the intermediate hosts of trematodes.

The effect of the crude extract of Artocarpus lakoocha (70% composition is 2,4,3',5'- tetrahydroxystilbene -THS) on adult Fasciola gigantica was evaluated after incubating the parasites in M-199 medium containing 250, 500, 750 and 1000 microg/ml of the crude extract, or triclabendazole (TCZ) at the concentrations of 80 and 175 microg/ml as the positive control, for 3, 6, 12 and 24h, using relative motility (RM) assay and observation by scanning electron microscope (SEM). Decreased contraction and motility were first observed after 3h incubation with TCZ at the concentration 80 and 175 microg/ml. TCZ markedly reduced the parasite's motility at the concentration of 175 microg/ml at 6h, and killed the worms after 12h exposure. The crude extract of A. lakoocha at all concentrations reduced the parasite's motility similar to TCZ at 3h incubation. In 250 and 500 microg/ml of the crude extract, the values were decreased from 3 to 12h, then they were stable between 12 and 24h and reduced to the level approximately 30-40% of the control. At 750 and 1000 microg/ml concentrations the crude extract rapidly reduced the RM values from the start to 12h and killed the parasites between 12 and 24h incubation. The crude extract also inhibited the larval migration by 75% and 100% at the concentrations of 250-500 and 750-1000 microg/ml, respectively. TCZ and the crude extract caused sequentially changes in the tegument including swelling, followed by blebbings that later ruptured, leading to the erosion and desquamation of the tegument syncytium. As the result, lesion was formed which exposed the basal lamina. The damage appeared more severe on the dorsal than the ventral surface, and earlier on the anterior part and lateral margins when compared to the posterior part. The severity and rapidity of the damages were enhanced with increasing concentration of the crude extract. Hence, the crude extract of A. lakoocha, may exert its fasciolicidal effect against adult F. gigantica by initially causing the tegumental damage.

The anthelmintic activity of the ethanolic extracts of stems and leaves of Meryta denhamii Seem. against adult liver flukes "Fasciola gigantica" was studied in vitro. Although leaves extract was inactive, stems extract exhibited anthelmintic activity and recorded LC50 and LC90 values, 16 and 26 gm/l respectively. The mode of action of the ethanolic extract of the stems on the adult flukes was evaluated by scanning electron microscopy (SEM). Tegumental sloughing, loss of spines and deformity of suckers were observed. These damages are responsible for the vermicidal effect of the ethanolic extract of the stems. The saponin contents of the stems (4.25%) and leaves (2.45%) were determined using haemolytic index. The plant was identified based on the macro and micromorphological features of the stem and leaf.

Triclabendazole (TCBZ) is an efficient fasciolicide that affects both juvenile and adult worms. After oral administration it is rapidly metabolized to TCBZ sulphoxide and sulphone that were found responsible for its fasciolicidal activity. Parasite defense mechanisms include detoxifying and anti-oxidant enzymes that would suppress its oxidative killing. The present work aimed at evaluating these enzymes under TCBZ-SX. Thirty juvenile and 30 adult F. gigantica worms collected from the liver parenchyma and bile ducts formed the subject of the study. Levels of superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR), glutathione S-transferase (GST), and glutathione (GSH) were measured in juvenile and adult worms incubated, without and with 15 and 50 microg/ml TCBZ-SX for 18 hours at 37 degrees C. Results demonstrated a significant decrease in SOD activity and an increase in GST level in both juvenile and adult worms after incubation in the two concentrations. The remaining enzymes and GSH revealed variable levels.

Little is known about the response of Fasciola spp., inside its snail host to pollutant toxicity. The effect of long-term exposure of Lymnaea natalensis to 0.1 microg/l, 10 microg/l, or 100 microg/l cadmium (Cd) on the infection rates with F. gigantica, percentage of cercariae-shedding snails and cercariae shedding pattern was investigated. The snails were exposed to Cd for 7 days either pre-infection with single Fasciola miracidium or during the late pre-patent period of the infection. The possible interaction between metal exposure and acclimatization temperature was also studied in three ranges; 16-18 degrees C, 23-25 degrees C and 30-32 degrees C. Results clearly showed that host exposure to 100 microg/l Cd significantly affected the pattern of Fasciola development inside its snail host, and that the acclimatization temperature was a key factor affecting the role played by Cd. Pre-infection exposure to Cd caused a significant reduction in the infection rates where the effect was temperature-dependent. Post-infection exposure to Cd significantly increased the percentage of cercariae-shedding snails; the effect was temperature-independent. Interestingly, post-infection exposure significantly altered the differential cercarial output pattern, while no clear impact on the total was found. The alteration in the differential cercarial output was represented as a significantly higher proportion of floating metacercaria as compared to fixed ones and also higher proportion of dead cercariae which may directly reduce the transmission of Fasciola to the final host. The mechanisms of cadmium impact are briefly discussed.

Triclabendazole "Fasinex" is the drug of choice against fasciolosis because of its high efficacy against both mature and immature flukes, however parasite resistance against this drug is increasing. Hence, there is pressing need for new fasciolicidal drugs. In the present study, the in vitro effect of artemether on adult flukes was evaluated using scanning electron microscopy. After 24 h incubation with 10 microg/ml artemether, the tegument of the apical cone appeared to be slightly more swollen than normal. This swelling became so severe and the spines appeared sunken, with their tips protruding from a swollen and blebbed base, on increasing the concentration to 20 microg/ml. With the higher concentration of 30 microg/ml, extensive and severe tegumental swelling occurred in the apical cone region of the flukes. There were many blebs around ventral sucker, a number of which appeared to have burst causing lesion. The tegumental changes occurred following incubation in artemether were comparable with those observed with triclabendazole in its active sulphoxide metabolite form (TCBZ-SX).

The aim of the present work was to investigate the Schistosoma mansoni and Fasciola gigantica cross-reactivity between adult worms and egg homogenates of the parasites. Immunoprophylactic effects of crude Schistosoma mansoni worms and egg antigens mixed with or without saponins extracted from Atriplex nummularia were studied followed by challenge with 80 cercariae of Schistosoma mansoni. Our results showed that post 1st immunization with schistosome egg antigens (SEA) there was a significant change (P approximately 0.05) in the IgM levels against Fasciola egg homogenate (FgEH) without saponins. Post 2nd immunization with SEA mixed with saponins the levels of IgM increased significantly (P approximately 0.05) against Fasciola worm homogenate (FgWH) as compared with a non-immunized group. Post 2nd immunization the level of IgG was significantly elevated (P approximately 0.05) by SEA mixed with saponins against FgWH. Post 2nd immunizations with SEA mixed with saponins showed a significant change (P approximately 0.05) in IgG levels against FgEH. These results clearly demonstrated that there is a cross-reactivity between Schistosoma mansoni eggs and Fasciola gigantica worms and eggs. Saponins were found to be immunostimulatory adjuvants in our study.

The efficacy of treating encysted metacercariae (EMC) of Fasciola gigantica with different concentrations (conc.) of acetic acid, citric acid, cetrimide, potassium permanganate and sodium hydroxide, for 15 & 30 minutes was evaluated. The efficacy of these chemicals on the vitality and infectivity of the EMC was evaluated by the development of fascioliasis infection, and the histopathological changes in the livers of experimentally infected Albino rabbits. The results showed that 1% sodium hydroxide had a lethal effect on EMC, 10% to 40% potassium permanganate destroyed the infectivity power of EMC, and acetic acid gave an adverse effect on the EMC in conc. more than 2.5%. But, neither citric acid nor cetrimide affected the vitality or infectivity of EMC and all rabbits acquired fascioliasis.

This study evaluated the effect of temperature, molluscicides (Copper sulphate and Niclosamide), fertilizers (Superphosphate and Ammonium sulphate) on the hatchability of Fasciola gigantica eggs. The results showed that hatchability decreased to 4% when eggs were incubated in bile secretion at 37 degrees C for 5 days and to 1.4% for 10 days, but few eggs incubated in water at 37 degrees C hatched. Bile secretion at 37 degrees C was a poor medium for in-vitro egg preservation. But, hatching occurred only when eggs were transferred to water at 26 degrees C. Temperature fluctuation from 26-4 degrees C or from 32-4 degrees C had an inhibitory effect on embryos development (35.2% & 32.3%, respectively) as compared to controls (60% & 63.9%, respectively). The incubation period (19 & 17 days) was higher than controls (14 & 12 days, respectively). The LC50 & LC90 of Copper sulphate and Niclosamide against Biomphalaria alexandrina and Lymnaea natalenesis had no toxic effect on Fasciola eggs. The higher concentrations of Copper sulphate (30 ppm) and Niclosamide (1 ppm) slightly lower eggs hatchability rate than controls. The rate decreased by increasing the exposure time from 3 to 6 hours with both molluscicides. Ammonium sulphate had a lethal effect on eggs, but Superphosphate had some inhibitory effect on egg development, which increased by increasing Superphosphate concentration or with the prolongation of the exposure time.

Phenyl vinyl sulfone is a synthetic inhibitor of cysteine protease and has antihelminthic and antiprotozoal properties. Phenyl vinyl sulfone was assayed in vitro for antifasciola activity against adult Fasciola gigantica worms using a well-established culture medium. Worms were treated with phenyl vinyl sulfone for incubation periods ranging from 0 to 12h and its activity was assessed in terms of viability, motility and death of worms. Phenyl vinyl sulfone exhibited a minimum effective concentration of 50 ppm after 12h. Three hundred parts per million concentrations were most potent causing immediate death of adult flukes in vitro. Histopathological studies showed that there was tegumental flattening, rupture of vesicles, and spine loss. Marked reduction in size and number of ova and sperms in the convoluted tubules of the reproductive organs was observed in comparison to the untreated control group. In conclusion, phenyl vinyl sulfone shows potent activity against F. gigantica in vitro, and the authors recommend carrying out more studies to detect its efficacy in vivo.

Praziquantel is still very effective for the treatment of schistosomiasis, but there are rising concerns on the potential risk of developing resistances because of the extensive use of this drug. Triclabendazole, a systemic anthelmintic, is very effective against other trematodes such as Paragonimus spp. and Fasciola spp. It has been reported to be effective in vitro and in experimental animals against Schistosoma mansoni. However, its antischistosomal efficacy in humans has not yet been evaluated. The objective of the study was to evaluate the efficacy of triclabendazole at the dosage currently used for the treatment of human fascioliasis (10mg/kg body weight) in subjects co-infected with S. mansoni and Fasciola spp. The study was carried out in Behera, a highly endemic area for both parasites, by personnel of the Egyptian Ministry of Health and Population. Ten subjects (m = 4, f = 6; age, 8-58years), who were infected at the same time by Fasciola spp. and S. mansoni, were enrolled. Six weeks after therapy, seven subjects were still excreting ova of S. mansoni, whereas none was excreting Fasciola spp. ova. At the given dosage, triclabendazole appeared not to be sufficiently effective in the treatment of S. mansoni.

Objectives: To study the clinical efficacy of Triclabendazole (TCBZ) on Egyptian patients infected with Fasciola and understand the effect of Fascioliasis on the pharmacokinetics of TCBZ., Methods: The pharmacokinetics of TCBZ administered as a single oral dose (10 mg/kg) was investigated in both infected and parasite--free Egyptian subjects. After oral administration, TCBZ is metabolized to a sulphone and sulfoxide derivatives. The latter is responsible for the fasciolicidal activity of TCBZ, and it could be used as a marker of drug bioavailability. Blood samples were collected following the oral administration, and TCBZ sulfoxide plasma concentrations were determined by a sensitive and specific HPLC method., Results: Pharmacokinetic parameters (Cmax, AUC0-48, t1/2 and tmax) for TCBZ sulfoxide were calculated. In patients; the mean Cmax was 9.11 +/- 1.3 microg/ml, the mean AUC(0-48) was 91 +/- 10.5 microg h ml(-1), the mean t1/2 was 7.4 +/- 0.6 h, and the tmax was 3.0 +/- 0.4 h. In normal subjects, the mean Cmax was 8.48 +/- 0.92 microg/ml, the mean AUC(0-48) was 85 +/- 6.55 microg h ml(-1), the mean t1/2 was 6.2 +/- 0.357 h, and the tmax was 3 +/- 0.4 h. No significant difference could be detected in the patients as compared to normal subjects, which would suggest that Fascioliasis does not affect any of the studied parameters. No eggs in faeces could be detected following TCBZ treatment. Also, most of the clinical investigations showed significant decline back to the normal ranges post-treatment which indicates complete curing and high TCBZ efficacy., Conclusion: Fasioliasis as an infective condition widely spread in Egypt has no significant effect on the pharmacokinetic parameters of the orally administered TCBZ and at the same time it is very effective against the parasite which strongly and safely suggests the use of this medication for the treatment of this infection.

The effect of Siwa propolis on adult flukes was evaluated using scanning electron microcopy. It gave an overview of the surface architecture of the tegument of Fasciola gigantica apical cone. The base of the spines appeared to be "flaking off" and showed severe blebbing after 24h incubation with 10 micro/ml propolis. This swelling became so sever and the spines were barely visible, on increasing the concentration to 20 micro/ml. Besides, there were many large blebs on the apical cone, a number of which appeared to have burst, causing lesions and the tegument was marked by a number of pits caused by the loss of spines. With the higher concentration of 30 micro/ml, erosion of the surface had occurred to such extent that no tegument remained, only a mass of fibrous structures. The tegumental changes occurred following incubation in propolis were compared with those observed with triclabendazole (TCBZ) "Fasinex" because of its high efficacy against both mature and immature flukes.

Contamination by fasciolids takes place through ingestion of metacercariae attached to vegetables. Experimental studies were performed with plant-made foods suggesting a role in human contamination in Iran and on the usefulness of potassium permanganate as a preventive tool for killing metacercariae attached to vegetables used in salads in Egypt. In the foods assayed, although viability decreases with time, a high percentage of the metacercariae were still alive 2 and 4 weeks after preparation. Infection of laboratory animals proved that metacercariae kept their infectivity. The 5-minute tests of potassium permanganate effects showed that metacercarial viability was not affected even at the very high doses of 300, 600, and 1,200 mg/L. Careful, subsequent washing of leaves and vegetables with water is therefore needed after its application. A review on similar studies performed with metacercariae belonging to fasciolid and other trematode species affecting humans is included.

Batches of encysted metacercariae of F. gigantica, adhered to transparent polyethylene sheets, were treated with KMnO4, while others were treated with NaDCC at specific concentrations and exposure times. Assessment of the effects was carried out by the detached percentage and viable metacercariae and by scanning electron microscope (SEM) ultrastructure changes. In addition, their effects on leaves of green vegetables were reported. The results showed that all metacercariae were detached and were dead by exposure to KMnO4 (96%) and NaDCC (100%) were detached from the polyethylene sheets. SEM showed that the deformities in the metacercariae soaked in NaDCC were more severe than those dipped in KMnO4. However, neither KMnO4 nor NaDCC affected the consistency, color, taste or flavor of the vegetables' leaves. The two disinfectants particularly NaDCC, proved to be safe, effective against the encysted metacercariae with no side-effects.

The efficacy of strategic anthelmintic control of liver flukes (Fasciola) and gastrointestinal (GI) nematodes on the performance of ewes and lambs on pasture was assessed on a farm in the highlands of Kenya. In May 1999, 45 Corriedale ewes, aged between 2 and 3 years, were ear-tagged, weighed and allocated randomly to three equal treatment groups based on body weight. Faecal samples taken at this time revealed low levels of strongyle-type eggs per gram of faeces (EPG) and the presence of liver fluke eggs in only a few of the animals. All the animals were then set stocked for 12 months on separate paddocks in an area endemic for both Fasciola and GI nematodes. The ewes in group 1 were given a combined anthelmintic treatment against Fasciola and GI nematodes during the periods recommended for the control of Fasciola in the area (February, June and October). The ewes in group 2 were given the combined treatments 3 weeks after the onset of both the short and long rainy seasons (November and April, respectively). Those in group 3 were given separate treatments for Fasciola (February, June and October) and nematodes (3 weeks after the onset of the rainy seasons). The anthelmintic treatment against Fasciola consisted of triclabendazole at 10 mg per kg body weight, and that against nematodes was levamisole at 10 mg per kg body weight. The nematode EPG for the ewes in group 1 were higher than in groups 2 and 3 during both rainy seasons. The nematode EPG did not differ significantly between groups 2 and 3. The prevalence of Fasciola eggs (number of ewes shedding eggs in a group) in the ewes in groups I and 3 remained very low throughout the study period compared to those in group 2. The highest birth weights and the weight gains of lambs were recorded for the group of ewes given separate anthelmintic treatments for Fasciola and nematodes (group 3). The results of this trial indicated that, in an area like Nyandarua District, where liver flukes and GI nematodes are important constraints to sheep production, the best practice is to give separate treatments for the two groups of parasites at the recommended times.

Human fascioliasis cases in Japan have been reported infrequently and the most appropriate therapy for this disease remains to be determined. This case report describes a patient with the infection unsuccessfully treated with multiple high doses of praziquantel for over 1 year and eventually cured by the administration of triclabendazole in two single doses of 10 mg/kg and 12.5 mg/kg 5 months apart.

The responses of rabbits experimentally infected orally with 25 Fasciola gigantica metacercariae each, to treatment with triclabendazole (TCBZ) in a single oral dose of 10 mg/Kg body weight or with the newly synthesized drug (BT6) in repeated oral doses of 500 mg/Kg for 3 consecutive days, were monitored by faecal egg and fluke counts and the enzyme-linked immunosorbent assay (ELISA), both treatments at 4-, 8- and 12- weeks of infection. In all infected rabbits, eggs appeared in the faeces between 10 and 16 weeks post-infection (P1) and Were egg-negative in the TCBZ-treated animals. Compared with the infected untreated control (group B), the reduction in the fluke burdens were 93.75,100 and 100% in TCBZ-treated rabbits (groups C, D and E), respectively and 33.33, 27.08 and 27.08% in the BT6 treated groups F, G and H, respectively. Total red blood corpuscle (RBCs) counts, haemoglobin (Hb) concentration and packed-cell volume (PCV) values in rabbits (group B) indicated the presence of moderate anaemia from the start of infection with a significant decrease from 10-12 weeks onwards. RBCs count, Hb and PCV values from groups C, D and E were significantly higher (p < 0.001) than in those from group B on week 10, 8 and 10 PI, respectively. The increasing was continued till week 16 PI. By two weeks PI there was significant eosinophilia in infected rabbits, maintained at 4 and 6 weeks PI then dropped up to 8 weeks but still statistically higher than the control values. The ELISA detected antibodies against F. gigantica as early as week 2 PI, rising to high levels 6, 10 and 12 weeks later in groups C, D and E, respectively, whilst peaked at 10 weeks in BT6 treated animals, then tended to drop but remained positive through 16 weeks. We conclude TCBZ is a potent fasciolicidic drug. Eosinophilia and anaemia might be indicators for fascioliasis. The ELISA with adult fluke excretory - secretory (ES) antigens could be a feasible method for the diagnosis of experimental fascioliasis in rabbits and post-treatment monitoring.

The molluscicidal properties of Solanum nigrum L. were tested against three Egyptian snail species (Biomphalaria alexandrina, Bulinus truncatus and Lymnaea natalensis), each an intermediate host of parasites causing human schistosomiasis or fascioliasis. The plant was collected in two regions within Egypt: Fayium and Giza. Snails were exposed for 24 and 48 h, to the dry powdered fruits and leaves or to crude water extracts of the powders, and mortality was recorded. The water extract of the leaves collected in Fayium (FLWE) had the highest molluscicidal activity, with median lethal concentrations (LC50) of 18.6 mg/litre for Bi. alexandrina, 14.5 mg/litre for Bu. truncatus and 17.7 mg/litre for L. natalensis. When Bi. alexandrina infected with Schistosoma mansoni were exposed to FLWE (20 or 25 mg/litre), they shed significantly fewer cercariae than unexposed snails (P < 0.02). The cercaricidal properties of FLWE were directly tested against S. haematobium, S. mansoni and Fasciola gigantica cercariae and a time-concentration relationship was observed; the concentrations needed to kill all cercariae (LC100) within 30 min of exposure were 30 mg/litre for both S. haematobium and S. mansoni and 40 mg/litre for F. gigantica.