In vivo efficacy of combination therapy with albendazole and atovaquone against primary hydatid cysts in mice.

Alveolar echinococcosis (AE) is caused by the larval stage of Echinococcus multilocularis. Chemotherapy for AE involves albendazole (ABZ), which has shown insufficient efficacy. More effective chemotherapy for AE is needed. Previously, we have demonstrated that atovaquone (ATV), an antimalarial, inhibits mitochondrial complex III of E. multilocularis and restricts the development of larval cysts in in vivo experiments. Therefore, in this study, we evaluated the efficacy of ABZ and ATV combination therapy on E. multilocularis in culture and in vivo experiments. Protoscoleces were treated with 50 µM ABZ and/or ATV in the medium; the duration of parasite elimination was determined under aerobic and anaerobic culture. In the in vivo experiment, the effects of ABZ and ATV combination treatment in BALB/c mice infected orally with eggs from the feces of an adult-stage E. multilocularis-infected dog were compared with those of standard oral ABZ therapy. In the culture assay, the duration of elimination associated with ABZ and ATV combination treatment was shorter than that associated with ATV alone under aerobic conditions. Protoscolex viability progressively reduced owing to the combination treatment under anaerobic conditions; however, either drug used singly did not exhibit antiparasitic effects under hypoxia. Furthermore, compared with ABZ alone, the combination treatment significantly reduced the growth of the primary cyst in the liver of mice infected orally with parasite eggs (P = .011). ATV enhances the effect of ABZ in the treatment of AE in mice.

Digest the Sugar, Kill the Parasite: A New Experimental Concept in Treating Alveolar Echinococcosis.

INTRODUCTION: The E. multilocularis laminated layer (LL) is a heavily glycosylated parasitic structure that plays an important role in protecting the larval stage (metacestode) of this parasite from physiological and immunological host reactions. We elaborated an experimental design with the idea to modify the (glycan) surface of the LL by a targeted digestion. This should allow the host defense to more easily recognize and attack (or kill) the parasite by immune-mediated effects. METHODS: Experimentally, E. multilocularis (clone H95) metacestodes were cultured in vitro with or without addition of α1-3,4,6-galactosidase or ß1-3-galactosidase in the medium. Morphological changes were subsequently measured by microscopy at different time points. Parasites were then recovered at day 5 and reinjected into mice for assessing their viability and infectious status. For finally recovered parasites, the respective load was assessed ex vivo by wet weight measurement, and host-related PD1 and IL-10 levels were determined as the key immunoregulators by using flow cytometry. RESULTS: Our experiments demonstrated that the parasite vesicular structure can be directly destroyed by adding galactosidases into the in vitro culture system, resulting in the fact that the parasite metacestode vesicles could not anymore infect and develop in mice after this glycan digestion. Moreover, when compared to the mice inoculated with E. multilocularis metacestode without galactosidases, PD1 expression was upregulated in CD4+ Teffs from mice inoculated with E. multilocularis metacestode pretreated with ß1-3-galactosidase, with a lower IL-10 secretion from CD4+ Teffs; there was no difference of PD1 and IL-10 expression levels regarding CD4+ Teff from mice inoculated with E. multilocularis metacestode pretreated with α1-3,4,6-galac-tosidase. DISCUSSION: We raised our hypothesis that this "aborting" effect may be linked to an altered PD1 and IL-10 response fine-tuning between immunopathology and immune protection. These findings justify a continuation of these experiments upon therapeutical in vivo administration of the enzymes.

In vitro effects of lonidamine and 6-aminonicotinamide against Echinococcus granulosussensu stricto and Echinococcus multilocularis.

Echinococcosis is a zoonotic disease caused by cestode species of the genus Echinococcus, which demonstrates considerable medical and veterinary concerns. The development of novel drugs for echinococcosis treatment is urgently needed. In this study, we demonstrated that lonidamine (LND) and 6-aminonicotinamide (6-AN) exhibited considerable in vitro effects against both larval- and adult-stage of E. granulosussensu stricto (s. s.) and E. multilocularis. The combination of LND and 6-AN exhibited a significantly higher activity than the single drug treatment. These results highlight the therapeutic potential of LND, 6-AN and the combination of LND and 6-AN for the treatment of echinococcosis.

Application of target repositioning and in silico screening to exploit fatty acid binding proteins (FABPs) from Echinococcus multilocularis as possible drug targets.

Fatty acid binding proteins (FABPs) are small intracellular proteins that reversibly bind fatty acids and other hydrophobic ligands. In cestodes, due to their inability to synthesise fatty acids and cholesterol de novo, FABPs, together with other lipid binding proteins, have been proposed as essential, involved in the trafficking and delivery of such lipophilic metabolites. Pharmacological agents that modify specific parasite FABP function may provide control of lipid signalling pathways, inflammatory responses and metabolic regulation that could be of crucial importance for the parasite development and survival. Echinococcus multilocularis and Echinococcus granulosus are, respectively, the causative agents of alveolar and cystic echinococcosis (or hydatidosis). These diseases are included in the World Health Organization's list of priority neglected tropical diseases. Here, we explore the potential of FABPs from cestodes as drug targets. To this end, we have applied a target repurposing approach to identify novel inhibitors of Echinococcus spp. FABPs. An ensemble of computational models was developed and applied in a virtual screening campaign of DrugBank library. 21 hits belonging to the applicability domain of the ensemble models were identified, and 3 of the hits were assayed against purified E. multilocularis FABP, experimentally confirming the model's predictions. Noteworthy, this is to our best knowledge the first report on isolation and purification of such four FABP, for which initial structural and functional characterization is reported here.

The importance of uninterrupted albendazole treatment in patients with unresectable alveolar echinococcosis undergoing liver transplantation.

BACKGROUND: This study focused on the monitoring of patients who had undergone liver transplantation (LT) because of unresectable alveolar echinococcosis (AE). The role of long-term administration of albendazole (ABZ) in patients with residual/recurrent AE lesions and without AE lesions was evaluated. METHODS: Albendazole therapy was prescribed to patients diagnosed with AE 4-6 weeks after LT on the background of continuous suppressive therapy while following the protocol for managing patients after LT. Clinical data (general condition, blood counts, and level of hepatic transaminases), ultrasound scans (USs), magnetic resonance imaging (MRI) or multispiral computed tomography (MSCT), and serological data were collected from four patients with residual/recurrent AE lesions and without AE lesions. The results of the USs, MRI, and MSCT examinations at diagnosis and at the end of follow-up were retrospectively reviewed for all patients. RESULTS: Observation of patients over a long period (up to 10 years) showed that the continuous (without interruption) use of ABZ restrained the development of metacestodes. Interruptions in taking the drug, associated with the manifestation of hepatotoxicity in some patients, led to the development of lesions in other organs in which the parasite had not previously been detected. No new foci were found in the transplanted livers of the patients. CONCLUSION: Liver transplantation, together with continuous anti-relapse chemotherapy, prolongs the patient's life, both in the absence and in the presence of metastases in other organs.

Albendazole solid dispersions against alveolar echinococcosis: a pharmacotechnical strategy to improve the efficacy of the drug.

Alveolar echinococcosis is a neglected parasitic zoonosis caused by Echinococcus multilocularis. The pharmacological treatment is based on albendazole (ABZ). However, the low water solubility of the drug produces a limited dissolution rate, with the consequent failure in the treatment of the disease. Solid dispersions are a successful pharmacotechnical strategy to improve the dissolution profile of poorly water-soluble drugs. The aim of this work was to determine the in vivo efficacy of ABZ solid dispersions using poloxamer 407 as a carrier (ABZ:P407 solid dispersions (SDs)) in the murine intraperitoneal infection model for secondary alveolar echinococcosis. In the chemoprophylactic efficacy study, the ABZ suspension, the ABZ:P407 SDs and the physical mixture of ABZ and poloxamer 407 showed a tendency to decrease the development of murine cysts, causing damage to the germinal layer. In the clinical efficacy study, the ABZ:P407 SDs produced a significant decrease in the weight of murine cysts. In addition, the SDs produced extensive damage to the germinal layer. The increase in the efficacy of ABZ could be due to the improvement of water solubility and wettability of the drug due to the surfactant nature of poloxamer 407. In conclusion, this study is the basis for further research. This pharmacotechnical strategy might in the future offer novel treatment alternatives for human alveolar echinococcosis.

Short communication: Efficacy of albendazole in Echinococcus multilocularis-infected mice depends on the functional immunity of the host.

Alveolar echinococcosis (AE) is a deadly parasitic disease that requires lifelong treatment with albendazole. Development of host immunity is pivotal with regard to the clinical outcome of AE, but its influence on conventional albendazole treatment is unknown. Using T-cell deficient athymic nude mice, we demonstrated that functional immunity is required for albendazole to be efficacious against murine AE. These results call for attention given the increasing number of immunocompromised patients with AE.

Successful surgical management of hepatic alveolar echinococcosis by inductive therapy with albendazole - a case report.

We report the case of a 65-year-old female patient with hepatic alveolar echinococcosis (AE) caused by Echinococcus multilocularis. This infrequent zoonosis has a considerable morbidity and mortality. The malignant appearing hepatic mass was initially misdiagnosed as cholangiocarcinoma of the right hepatic lobe (segments VII, VIII, and IVa, sized 10.9 cm × 7.6 cm) involving the right and middle hepatic vein and extending close to the left hepatic vein. During exploratory laparotomy, the frozen-section biopsy was indicative of AE (World Health Organization [WHO] classification: stage P3N0M0). Due to the high operative risk, it was decided to pretreat the patient with albendazole as inductive therapy in order to remove the AE secondarily in accordance with the patient's request. After year-long treatment with albendazole (under strict control of the maximum blood levels), a right hemihepatectomy was successfully performed. Postoperative treatment with albendazole had to be stopped prematurely after 11 months due to considerable subjective intolerance and a more-than-tenfold elevation of transaminases despite normal therapeutic albendazole blood levels. A 18F-FDG-PET/CT scan revealed no evidence of AE residues. Conducting follow-up examinations by 18F-FDG-PET/CT scans every 2 years is planned in order to recognize possible recurrence at an early stage.

Enhanced Oral Bioavailability and Anti-Echinococcosis Efficacy of Albendazole Achieved by Optimizing the "Spring" and "Parachute".

Maximizing the pharmacological efficacy of albendazole (ABZ), an anti-echinococcosis drug, is essential in the long-term treatment of patients with echinococcosis. As a weakly alkaline drug, ABZ has a pH-dependent solubility that decreases dramatically from gastric fluid (pH 1.4) to intestinal fluid (pH 6.5), where it is absorbed. In this study, we endeavored to develop an optimized tablet formulation of ABZ to improve its dissolution and oral bioavailability from two aspects: a faster initial dissolution in the gastric pH condition (i.e., the "spring") and a more prolonged drug supersaturation in the intestinal pH condition (i.e., the "parachute"). To achieve this goal, ABZ-HCl salt was selected first, which demonstrated a higher intrinsic dissolution rate under pH 1.4 compared with the ABZ free base that is used in the commercial product Albenda. Second, by comparing the ABZ supersaturation kinetics under pH 6.5 in the presence of various polymers including poly(vinylpyrrolidone) (PVP), PVP/VA, hydroxypropyl methylcellulose (HPMC), and HPMC acetate succinate (HPMC-AS), HPMC-AS was found to be the most effective crystallization inhibitor for ABZ, likely due to the hydrophobic interaction between ABZ and HPMC-AS in an aqueous environment. The newly designed tablet formulation containing ABZ-HCl and HPMC-AS showed ∼3 times higher oral bioavailability compared with that of Albenda in Beagle dogs. More significantly, the anti-echinococcosis efficacy of the improved formulation was 2.4 times higher than that of Albenda in a secondary hepatic alveolar echinococcosis Sprague-Dawley rat model. The strategy of simultaneously improving the spring and parachute of an oral formulation of ABZ, by using a highly soluble salt and an effective polymeric crystallization inhibitor, was once again proven to be a viable and readily translatable approach to optimize the unsatisfactory oral medicines due to solubility and bioavailability limitations.

In vitro and in vivo effects of 3-bromopyruvate against Echinococcus metacestodes.

While searching for novel anti-echinococcosis drugs, we have been focusing on glycolysis which is relied on by Echinococcus for energy production and intermediates for other metabolic processes. The aim of this study was to investigate the potential therapeutic implication of glycolytic inhibitors on Echinococcus. Our results demonstrate that at an initial concentration of 40 µM, all inhibitors of glycolysis used in the current experiment [3-bromopyruvate (3-BrPA), ornidazole, clorsulon (CLS), sodium oxamate and 2,6-dihydroxynaphthalene (NA-P2)] show considerable in vitro effects against Echinococcus granulosus protoscoleces and Echinococcus multilocularis metacestodes. Among them, 3-BrPA exhibited the highest activity which was similar to that of nitazoxanide (NTZ) and more efficacious than albendazole (ABZ). The activity of 3-BrPA was dose dependent and resulted in severe ultrastructural destructions, as visualized by electron microscopy. An additional in vivo study in mice infected with E. multilocularis metacestodes indicates a reduction in parasite weight after the twice-weekly treatment of 25 mg/kg 3-BrPA for 6 weeks, compared to that of the untreated control. In particular, in contrast to ABZ, the administration of 25 mg/kg 3-BrPA did not cause toxicity to the liver and kidney in mice. Similarly, at the effective dose against Echinococcus larvae, 3-BrPA showed no significant toxicity to human hepatocytes. Taken together, the results suggest that interfering with the glycolysis of the parasite may be a novel chemotherapeutical option and 3-BrPA, which exhibited a remarkable activity against Echinococcus, may be a promising potential drug against cystic echinococcosis (CE) and alveolar echinococcosis (AE).

Effect of anacardic acid against echinococcosis through inhibition of VEGF-induced angiogenesis.

Echinococcosis is a zoonotic infection caused by cestode species of the genus Echinococcus, with limited treatment options. It is urgent to develop new anti-hydatid agent. In this paper, we reported anacardic acid (AA), a natural product isolated from the Brazilian cashew-nut shell liquid, which presented a high activity against metacestodes of Echinococcus multilocularis (E. multilocularis) and Echinococcus granulosus sensu stricto (E. granulosus s.s.) in vitro and in vivo. AA exerted a better efficacy on E. granulosus s.s. protoscoleces and E. multilocularis metacestodes than that of albendazole (ABZ) and dihydroartemisinin (DHA) in vitro, and an inhibition on the growth of Echinococcus metacestode as effective as ABZ in vivo. Moreover, we also found that one of the mechanisms of AA against Echinococcus could be the suppression of angiogenesis on/in the metacestode mass through inhibiting vascular endothelial growth factor (VEGF)-induced signalling pathways. This work finds that AA is a new promising potential candidate drug for echinococcosis treatment.

Drug repurposing for the treatment of alveolar echinococcosis: in vitro and in vivo effects of silica nanoparticles modified with dichlorophen.

Alveolar echinococcosis is a neglected parasitic zoonosis caused by the metacestode Echinococcus multilocularis, which grows as a malignant tumour-like infection in the liver of humans. Albendazole (ABZ) is the antiparasitic drug of choice for the treatment of the disease. However, its effectiveness is low, due to its poor absorption from the gastro-intestinal tract. It is also parasitostatic and in some cases produces side-effects. Therefore, an alternative to the treatment of this severe human disease is necessary. In this context, the repositioning of drugs combined with nanotechnology to improve the bioavailability of drugs emerges as a useful, fast and inexpensive tool for the treatment of neglected diseases. The in vitro and in vivo efficacy of dichlorophen (DCP), an antiparasitic agent for intestinal parasites, and silica nanoparticles modified with DCP (NP-DCP) was evaluated against E. multilocularis larval stage. Both formulations showed a time and dose-dependent in vitro effect against protoscoleces. The NP-DCP had a greater in vitro efficacy than the drug alone or ABZ. In vivo studies demonstrated that the NP-DCP (4 mg kg-1) had similar efficacy to ABZ (25 mg kg-1) and greater activity than the free DCP. Therefore, the repurposing of DCP combined with silica nanoparticles could be an alternative for the treatment of echinococcosis.

In vitro efficacies of solubility-improved mebendazole derivatives against Echinococcus multilocularis.

Recently, we introduced an epoxy group to mebendazole by a reaction with epichlorohydrin and obtained two isoforms, mebendazole C1 (M-C1) and mebendazole C2 (M-C2). The in vitro effects of mebendazole derivatives at different concentrations on Echinococcus multilocularis protoscoleces and metacestodes as well as cytotoxicity in rat hepatoma (RH) cells were examined. The results demonstrated that the solubility of the two derivatives was greatly improved compared to mebendazole. The mortality of protoscoleces in vitro reached to 70-80% after 7 days of exposure to mebendazole or M-C2, and M-C2 showed higher parasiticidal effects than mebendazole (P > 0.05). The parasiticidal effect of M-C1 was low, even at a concentration of 30 µm. The percentage of damaged metacestodes that were treated with mebendazole and M-C2 in vitro at different concentrations were similar, and M-C1 exhibited insignificant effects on metacestodes. Significant morphological changes on protoscoleces and metacestodes were observed after treatment with mebendazole and M-C2. In addition, the introduction of an epoxy group to mebendazole also reduced its cytotoxicity in RH cells. Our results demonstrate that the introduction of an epoxy group not only improved the solubility of mebendazole, but also increased its parasiticidal effects on E. multilocularis and reduced its cytotoxicity in RH cells.

Activity of Thymus capitatus essential oil components against in vitro cultured Echinococcus multilocularis metacestodes and germinal layer cells.

The essential oil (EO) of Thymus capitatus, seven fractions (F1-F7) obtained from silica gel chromatography, and several pure EO components were evaluated with respect to in vitro activities against Echinococcus multilocularis metacestodes and germinal layer (GL) cells. Attempts to evaluate physical damage in metacestodes by phosphoglucose isomerase (PGI) assay failed because EO and F1-F7 interfered with the PGI-activity measurements. A metacestode viability assay based on Alamar Blue, as well as transmission electron microscopy, demonstrated that exposure to EO, F2 and F4 impaired metacestode viability. F2 and F4 exhibited higher toxicity against metacestodes than against mammalian cells, whereas EO was as toxic to mammalian cells as to the parasite. However, none of these fractions exhibited notable activity against isolated E. multilocularis GL cells. Analysis by gas chromatography-mass spectrometry showed that carvacrol was the major component of the EO (82.4%), as well as of the fractions F3 (94.4%), F4 (98.1%) and F5 (90.7%). Other major components of EO were ß-caryophyllene, limonene, thymol and eugenol. However, exposure of metacestodes to these components was ineffective. Thus, fractions F2 and F4 of T. capitatus EO contain potent anti-echinococcal compounds, but the activities of these two fractions are most likely based on synergistic effects between several major and minor constituents.

The genomes of four tapeworm species reveal adaptations to parasitism.

Tapeworms (Cestoda) cause neglected diseases that can be fatal and are difficult to treat, owing to inefficient drugs. Here we present an analysis of tapeworm genome sequences using the human-infective species Echinococcus multilocularis, E. granulosus, Taenia solium and the laboratory model Hymenolepis microstoma as examples. The 115- to 141-megabase genomes offer insights into the evolution of parasitism. Synteny is maintained with distantly related blood flukes but we find extreme losses of genes and pathways that are ubiquitous in other animals, including 34 homeobox families and several determinants of stem cell fate. Tapeworms have specialized detoxification pathways, metabolism that is finely tuned to rely on nutrients scavenged from their hosts, and species-specific expansions of non-canonical heat shock proteins and families of known antigens. We identify new potential drug targets, including some on which existing pharmaceuticals may act. The genomes provide a rich resource to underpin the development of urgently needed treatments and control.

Therapeutic efficacy of nanocompounds in the treatment of cystic and alveolar echinococcoses: challenges and future prospects.

Echinococcus granulosus sensu lato and E. multilocularis are the causative agents of life-threatening cystic and alveolar echinococcoses (CE and AE), respectively, which lead to serious public health concerns across the globe. Benzimidazoles (BMZs) are the drugs of choice for the treatment of human CE and AE. Presently, the chemotherapeutic failures of BMZs against CE and AE are caused by their low aqueous solubility, poor absorption, and consequently their erratic bioavailability. Among the BMZ compounds used for CE/AE treatment, albendazole (ABZ) and mebendazole (MBZ) are the only drugs licensed for human use. Nevertheless, the administration of these BMZs for a long period of time leads to undesirable adverse effects. Therefore, there is an urgent need for designing new formulations of BMZs with increased bioavailability. To bridge these therapeutic gaps, nanoparticle enantiomers of ABZ and drug delivery systems based on nanostructured entities currently provide an interesting new formulation of already existing drugs to improve the pharmacokinetic effects of BMZs. This study provides an overview of the tested nanocompounds against E. granulosus and E. multilocularis, including their effective dose, type of nanoparticles (NPs), assay setting, and therapeutic outcomes. This review suggests that BMZ derivatives loaded in NPs can significantly improve the scolicidal and cysticidal activities compared with single BMZ. Moreover, BMZ-loaded polymeric NPs show a tendency to increase mortality rate against protoscoleces and microcysts compared with metallic formulations, nanoemulsions, lipid nanocapsules, solid lipid NPs, liposomes, and nanocrystals. In the future, the use of the newly structured entities, attained by bridging ligands to the modified surface of NPs, as well as the electromagnetically produced nanodrugs could be helpful for developing fine-tuned formulations as an alternative to the already existing drugs against these neglected parasitic infections.

Evaluation of kinase-inhibitors nilotinib and everolimus against alveolar echinococcosis in vitro and in a mouse model.

Infection with the larval stage (metacestode) of the fox tapeworm Echinococcus multilocularis leads to a primary hepatic disease referred to as alveolar echinococcosis (AE). The progressive disease can be lethal if untreated. In cases where complete parasite resection by surgery is not feasible, the current treatment regimens of AE consist of chemotherapy with the parasitostatic benzimidazoles albendazole or mebendazole over decades. Kinase-inhibitors currently administered in various cancer treatments are of increasing interest also as anti-parasitic drugs due to previous promising in vitro results. In order to search for novel drug targets and treatment regimens, nilotinib (AMN107; Tasigna®), an Abl-tyrosine kinase inhibitor and everolimus (RAD001; Afinitor®), a serine/threonine-kinase inhibitor, were tested for their treatment efficacy against metacestode vesicles of E. multilocularis in vitro and in BALB/c mice. In vitro treatment with 200 µM nilotinib caused drug-induced alterations after 12 days, and everolimus exerted parasite damage at concentrations dosing from 40 to 100 µM after 5 and 12 days of in vitro exposure. Nilotinib (100 mg/kg) + erythromycin (to increase nilotinib plasma levels: 10 mg/kg intraperitoneal) or everolimus (5 mg/kg) were formulated in honey and administered daily for three weeks and subsequently twice a week for an additional three weeks in experimentally infected mice. Treatments did not result in any reduction of parasite growth compared to untreated control groups, whereas oral treatment with albendazole (200 mg/kg) was highly effective. Combined application of the kinase-inhibitors with albendazole did not lead to a synergistic or additive treatment efficacy compared to albendazole treatment alone. These results show that neither nilotinib nor everolimus represent valuable alternatives to the current treatment regimens against AE.

To see or not to see: non-invasive imaging for improved readout of drug treatment trials in the murine model of secondary alveolar echinococcosis.

Alveolar echinococcosis (AE) is an emerging zoonotic disease caused by the cestode Echinococcus multilocularis. The secondary infection model of AE is based on intraperitoneal injection of disease-causing metacestodes into the peritoneal cavity of mice, which allows investigations on novel drugs or immunotherapeutical treatment options in vivo. So far, such in vivo studies assessed exclusively the parasite weight at the endpoint of a given treatment period. We here developed an ultrasound (US)-based scoring system that allows to follow-up parasite development in the living animal, and provides insights into parasite growth during the treatment phase. By this method a statistically significant difference between untreated and medicated mice with E. multilocularis infection was observed at 2 months post-infection, and the growth curve of the parasite load was described by a linear mixed model. High correlation and similar levels of variation were observed for the standard method based on parasite weight measurement, the novel US-based scoring system, as well volume segmentation by post-mortem magnetic resonance imaging. Thus, US-based scoring in the live animal has the potential to assist the 3R concept by contributing to the refinement and reduction of animal use in experimental echinococcosis.

Immunoregulation in larval Echinococcus multilocularis infection.

Alveolar echinococcosis (AE) is a clinically very severe zoonotic helminthic disease, characterized by a chronic progressive hepatic damage caused by the continuous proliferation of the larval stage (metacestode) of Echinococcus multilocularis. The proliferative potential of the parasite metacestode tissue is dependent on the nature/function of the periparasitic immune-mediated processes of the host. Immune tolerance and/or down-regulation of immunity are a marked characteristic increasingly observed when disease develops towards its chronic (late) stage of infection. In this context, explorative studies have clearly shown that T regulatory (Treg) cells play an important role in modulating and orchestrating inflammatory/immune reactions in AE, yielding a largely Th2-biased response, and finally allowing thus long-term parasite survival, proliferation and maturation. AE is fatal if not treated appropriately, but the current benzimidazole chemotherapy is far from optimal, and novel options for control are needed. Future research should focus on the elucidation of the crucial immunological events that lead to anergy in AE, and focus on providing a scientific basis for the development of novel and more effective immunotherapeutical options to support cure AE by abrogating anergy, anticipating also that a combination of immuno- and chemotherapy could provide a synergistic therapeutical effect.

A Rapid and Convenient Method for in Vivo Fluorescent Imaging of Protoscolices of Echinococcus multilocularis.

Human and animal alveolar echinococcosis (AE) are important helminth infections endemic in wide areas of the Northern hemisphere. Monitoring Echinococcus multilocularis viability and spread using real-time fluorescent imaging in vivo provides a fast method to evaluate the load of parasite. Here, we generated a kind of fluorescent protoscolices in vivo imaging model and utilized this model to assess the activity against E. multilocularis protoscolices of metformin (Met). Results indicated that JC-1 tagged E. multilocularis can be reliably and confidently used to monitor protoscolices in vitro and in vivo. The availability of this transient in vivo fluorescent imaging of E. multilocularis protoscolices constitutes an important step toward the long term bio-imaging research of the AE-infected mouse models. In addition, this will be of great interest for further research on infection strategies and development of drugs and vaccines against E. multilocularis and other cestodes.