Hydatid Cyst Protoscoleces Research Articles

Evaluation of the scolicidal activities of eugenol essential oil and its nanoemulsion against protoscoleces of hydatid cysts.

BackgroundCystic echinococcosis caused by the cestode Echinococcus granulosus remains a serious helminthic zoonosis affecting humans and animals in many endemic developing countries. Surgical intervention is the best management choice, although it is associated with high recurrence rates and serious complications. Also, the commonly used chemotherapeutics exhibited serious side effects. This study aimed to evaluate the protoscolicidal effects of eugenol (Eug) essential oil and its nanoemulsion (Eug-NE) against protoscoleces (PCs) of hydatid cysts in vitro.MethodsEug-NE was prepared and characterized. Their cytotoxicity on macrophages was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. E. granulosus PCs were treated with various concentrations of Eug and Eug-NE at different exposure times. The viability of protoscoleces was evaluated by the eosin exclusion test, and the changes in the morphology of protoscoleces were assessed. Albendazole (ABZ) was used as a positive control.ResultsThe cellular cytotoxicity of Eug and Eug-NE on macrophage cells, in minimum and maximum concentrations (0.2 and 1 μl/mL), were nearly negligible ranging from 4.7% to 8.3% and 3.7% to 7.2%, respectively. The results showed highly significant activity of Eug-NE and Eug against hydatid PCs compared to ABZ (P < 0.05). Eug and Eug-NE have similar protoscolicidal effects at all used concentrations. Their highest scolicidal activity (100% mortality rate) was recorded at 1 μl/ml after 30 min incubation (LC50 = 0.298—LC90 = 0.521 and LC50 = 0.309—LC90 = 0.646, respectively). Both formulations showed time- and dose-dependent effects.ConclusionsThis study suggested the potent scolicidal activities of Eug and Eug-NE as promising alternative scolicidal agents. Future studies are recommended to explore the mechanism of action and treatment response in vivo and clinical settings.

Iranian Herbal Medicines Against Hydatid Cyst Protoscoleces: A Systematic Review.

In recent years, more attention has been focused in the practice of both crude extract of medicinal plants, and the screening plant-derived compounds as substitute scolicidal agents during hydatid cyst surgery. The present study was designed to review the protoscolicidal effects of some Iranian herbal medicines against hydatid cyst protoscoleces. English databases, including PubMed, Google Scholar, Web of Science, EBSCO, Science Direct, and Scopus were searched for publications worldwide related to protoscolicidal effects of Iranian herbal medicines without date limitation so that identify all published articles (in vitro, in vivo, clinical and case-control) have studied. Keywords included "Protoscolicidal", "Scolicidal", "Herbal medicines", "Extract", "Essential oil", "Plant", "In vitro", and "Iran". Moreover, the language of data collection was limited to English. In total, 40 papers up to 2020 were included in the present systematic review. The most studies were conducted on protoscolicidal activity of methanolic extracts (17 studies) followed by essential oils (15 studies), and aqueous extract (3 studies). The most commonly used part of herbs were leaves (21 herbs), seeds (8 herbs), and fruit (6 herbs), respectively. Moreover, the most prevalent herbal family was Lamiaceae (6 studies), followed by Apiaceae (5 studies) and Liliaceae (5 studies). Considering the high efficacy of Iranian herbal medicines against hydatid cyst protoscoleces, it can be concluded that Iranian herbal medicines have ability to consider as new alternative protoscolicidal agents during hydatid cyst surgery; however, more clinical studies are required to discover the precise protoscolicida activity of Iranian medicines in animal and human subjects.

Chemical Composition, Apoptotic Activity, and Antiparasitic Effects of Ferula macrecolea Essential Oil against Echinococcus granulosus Protoscoleces

Background: Today, the present protoscolicidals used to minimize the serious risks during hydatid cyst surgery are not completely safe and have various adverse side effects. The present study aimed to evaluate the chemical composition and apoptotic activity of Ferula macrecolea essential oil (FMEO) as well as its in vitro and ex vivo protoscolicidal effects against hydatid cyst protoscoleces. Methods: Gas chromatography/mass spectrometry (GC/MS) analysis was performed to determine the chemical composition of FMEO. Protoscoleces of hydatid cysts were collected from liver fertile hydatid cysts of infected sheep and were then treated with various concentrations of the essential oil (75, 150, and 300 µL/mL) for 5–60 min in vitro and ex vivo. Then, by using the eosin exclusion test, the viability of the protoscoleces was studied. The caspase-3-like activity of the FMEO-treated protoscoleces was also evaluated through the colorimetric protease assay Sigma Kit based on the manufacturer’s instructions. Results: According to GC/MS, the main constituents of the essential oil were terpinolene (77.72%), n-nonanal (4.47%), and linalool (4.35%), respectively. In vitro, the maximum protoscolicidal activity of FMEO was observed at the concentrations of 150 and 300 µL/mL, such that 100% of the protoscoleces were killed after 30 and 20 min of exposure, respectively. Based on the obtained findings, the results demonstrate that FMEO required a longer time to kill protoscoleces ex vivo; after 12 min of exposure to FMEO, only 13.4% of the protoscoleces remained alive. After 48 h of the treatment of protoscoleces, FMEO, in a dose-dependent manner and at doses of 75, 150, and 300 µL/mL, induced the activation of the caspase enzyme by 24.3, 35.3, and 48.3%, respectively. Conclusions: Our findings demonstrate the potent protoscolicidal effects of FMEO in vitro and ex vivo; however, further studies are required to assess the safety and the efficiency of FMEO as a promising scolicidal agent in a preclinical model and clinical setting.

Copper nanoparticles: Biosynthesis, characterization, and protoscolicidal effects alone and combined with albendazole against hydatid cyst protoscoleces

BackgroundSurgery remains the preferred treatment option for hydatid cyst (cystic echinococcosis); however, recent studies have demonstrated that the current protoscolicidal agents used during surgery are associated with some adverse side effects such as biliary fibrosis, hepatic necrosis, and cirrhosis. The present study aims to evaluate the in vitro and ex vivo anti-parasitic effects of copper nanoparticles (CuNPs) alone and combined with albendazole on hydatid cyst protoscoleces. MethodsCuNPs was green synthesized using C. spinosa extract. Various concentrations of CuNPs (250, 500, and 750 mg/mL) alone and combined with albendazole (ALZ, 200 mg/mL) were exposed to protoscoleces collected from the liver fertile hydatid cysts of infected sheep for 5−60 min in vitro and ex vivo. Next, the eosin exclusion test was applied to determine the viability of protoscoleces. Caspase-3 like activity of CuNPs-treated protoscoleces was then evaluated using the colorimetric protease assay Sigma Kit based on the manufacturer’s instructions. ResultsScanning electron microscopy (SEM) results showed that the particle size of CuNPs was 17 and 41 nm with the maximum peak at the wavelength of 414 nm. The maximum protoscolicidal activity of CuNPs was observed at the concentration of 750 mg/mL in vitro, so that 73.3 % of protoscoleces were killed after 60 min of exposure. Meanwhile, the mortality of protoscoleces was 100 % after 10 min of exposure to 750 mg/mL of CuNPs along with ALZ (200 mg/mL). Nevertheless, the findings proved that CuNPs even in combination with ALZ required a longer time to kill protoscoleces ex vivo. After 48 h of treating protoscoleces, CuNPs in a dose-dependent manner and at doses of 250, 500, and 750 mg/mL induced the caspase enzyme activation by 20.5 %, 32.3 %, and 36.1 %, respectively. ConclusionThe findings of the present investigation showed potent protoscolicidal effects of CuNPs, especially combined with albendazole, which entirely eliminated the parasite after 10−20 min of exposure. The results also showed that although the possible protoscolicidal mechanisms of CuNPs are not clearly understood, the inducing apoptosis through caspases is one of the main protoscolicidal mechanisms of CuNPs. However, supplementary studies, especially in animal models and clinical settings, are needed to approve these results.

Customizing the Protoscolicidal Activity by a Drug Delivery System: Application of Guar Gum in Electrospun Nanofibers.

Background:The present study aimed to control mebendazole drug release from ethyl cellulose nanofibers containing guar gum produced by Electrospinning Method (ESM) on mortality of hydatid cyst protoscoleces under laboratory conditions.Methods:The study was conducted in Arak Islamic Azad University, 2019. After preparation of ethyl cellulose nanofibers containing guar gum with concentrations 10, 250, 50 and 500 ppm with ESM, the uniformity and fineness of nanofibers were investigated by electron microscope. By determining the absorption of nanofibers during 312 h via spectrophotometry method, the amount of drug release was obtained. Then, the mortality of live protoscoleces in-vitro with nanofibers made with different concentrations was studied during 13 days.Results:Guar gum nanofiber with four concentrations of 10, 50, 250 and 500 ppm had 0.78512, 0.83729, 1.0098 and 1.0633 absorption respectively and showed drug release 42.09%, 39.95%, 33.05% and 30.96% after 312 hours. Therefore, the survival of protoscoleces in the presence of guar gum with four concentrations was zero after 3, 6, 11 and 13 days (P<0.05).Conclusion:To produce nanofibers carrying the drug for research related to the treatment of hydatid cysts, the electrospinning technique can be considered as a reliable method.

IN-VITRO SCOLICIDAL EFFECTS OF GRAPE AND APPLE VINEGAR ON THE HYDATID CYST PROTSCOLECES

At present, surgery remains the preferred treatment for hydatid cyst. Various chemical scolicidal agents have been used for inactivation of protoscoleces during surgery, but most of them are associated with adverse side effects. In this study we investigated the effect of two commercial grape and apple vinegar on the viability of hydatid cyst protoscoleces. The protoscoleces of E. granulosus were aspirated from the infected livers and lungs of sheep slaughtered at Tiaret abattoir. 0.5 mL of two commercial grape and apple vinegar was used for 5, 10, and 15 min in the experiments. 0.1% eosin staining assessed viability of protoscoleces. All tests were carried in triplicate. The mortality rates of protoscoleces were 100% after 5min, 10 min and 15 min of incubation with grape vinegar and apple vinegar. To conclude, the results of the present survey indicated high scolicidal activity of apple vinegar and grape vinegar against protoscoleces of hydatid cyst and can be used in hydatid cyst surgery. However, further studies will be needed to confirm these results by checking this scolicidal activity in an in-vivo model.

In Vitro and Ex Vivo Evaluation of Capparis spinosa Extract to Inactivate Protoscoleces During Hydatid Cyst Surgery.

Hydatidosis is one of the most dangerous zoonosis diseases in the world caused by the larval stage of the broad-worm or Echinococcus granulosus parasite. Today, cysts' rupture or content leakage during surgery and involvement of organs adjacent to the organ involved, and consequently secondary cysts, are the major concern for hydatid cyst surgeons. Therefore, using scolicidal substances such as hypertonic saline 20%, silver nitrate and formalin has been considered to reduce the risk of protoscoleces spread and recurrence of disease in recent years. The current work was designed to assess the antiparasitic effects of Capparis spinose L. extract against hydatid cyst protoscoleces. Collected protoscoleces from liver fertile hydatid cysts of infected sheep were exposed to the different concentrations of the essential oil (150, 300, 600 mg/mL) for 5-60 min in vitro and ex vivo. Then by using the eosin exclusion assay, the viability of protoscoleces was studied. The primary phytochemical analysis of the C. spinosa extract was done to assess the presence of tannins, alkaloids, saponins, flavonoids, terpenoids and glycosides. C. spinosa extract exhibited a powerful protoscolicidal activity in vitro so at the dose of 300 and 600 mg/ml, it entirely eliminated the parasite after 10 and 5 minutes; whereas at lower doses, it demonstrated weak protoscolicidal activity. In ex vivo assay, no similar effect to in vitro assay was observed, so more time was required to show a potent protoscolicidal activity. C. spinosa extract, at the concentrations of 300 and 600 mg/mL after an exposure time of 20 and 12 min, killed 100% of protoscoleces within the hydatid cyst, respectively. The findings of primary phytochemical screening of the C. spinosa extract demonstrated the existence of flavonoids, tannins, terpenoids, glycosides and alkaloids in this plant. The obtained results in vitro and ex vivo exhibited potent protoscolicidal effects of C. spinosa extract particularly at the concentrations of 600 and 300 mg/ml, which entirely eliminated the parasite after 5-20 min exposure. However, more supplementary works are required to verify these findings through assessing in animal models and clinical subjects.

Correction to: Main compounds and in vitro effectiveness of Syzygium aromaticum essential oil on protoscoleces of hydatid cyst

In the original version of this article, the first name of the corresponding author Sidi was incorrectly spelled as “Sid”.

Efficacy and Safety Curcuma zadoaria L. to Inactivate the Hydatid Cyst Protoscoleces.

Background:The present work aimed to evaluate the chemical composition of Curcuma zadoaria essential oil and to investigate its efficacy and safety against hydatid cyst protoscoleces.MethodsCollected protoscoleces from liver fertile hydatid cysts of infected sheep were exposed to different concentrations of the essential oil (75, 150, 300 µl/mL) for 5-30 min in vitro and ex vivo. Then, by using the eosin exclusion assay, the viability of protoscoleces was studied. In the next step, 24 male NMRI mice were examined to assess the toxicity of C. zadoaria essential oil by measuring the biochemical and hematological parameters.ResultsBased on the obtained results, the LD50 value of intraperitoneal injection of the C. zadoaria essential oil was 1.76 mL/kg of body weight and the maximum non-fatal dose was 0.96 mL/kg of body weight. C. zadoaria essential oil had a strong proto scolicidal activity in vitro so that at the 300 and 150 µl/ml entirely eliminates the parasite after 5 and 10 minutes; whereas, weak proto scolicidal activity was observed at lower doses. Ex vivo assay, no similar effect with in vitro was observed, therefore, more time is required to show a potent proto scolicidal activity. C. zadoaria essential oil at the concentrations of 300 and 150 µl/mL after an exposure time of 7 and 12 min, killed 100% of protoscoleces within the hydatid cyst, respectively. After intraperitoneal injection of the C. zadoaria essential oil for 2 weeks, no significant difference (p > 0.05) was observed in the clinical chemistry and hematologic parameters at the doses of 0.15, 0.3, 0.6 mL/kg.ConclusionThe obtained results in vitro and ex vivo exhibited that C. zadoaria essential oil had a favorable proto scolicidal activity on hydatid cyst protoscoleces. However, more supplementary works are required to verify these findings by assessing clinical subjects.

Constituents of essential oil of Origanum minutiflorum and its in vitro antioxidant, scolicidal and anticancer activities

ObjectiveOriganum minutiflorum essential oil was evaluated for its chemical components, in vitro antioxidant, scolicidal and anticancer activities. MethodsOil was extracted and its components were identified with spectrometry and flame ionisation gas chromatography. The oil antioxidant activity was evaluated by DPPH radical scavenging and β-Carotene/linoleic acid bleaching assay. Scolicidal effect of the oil against protoscolices of hydatid cysts was evaluated at different concentrations and exposure times. Also, different oil concentrations were testififed for its anticancer activities on breast cancer cell line (MCF-7), human lung cancer cell line (A-549) and human hepatocellular carcinoma cell line (HepG2) ResultsAnalyses of the O. minutiflorum oil identified 64 components with carvacrol being the predominant compound. The oil showed a weak free radical scavenging capacity (IC50 at 310 ± 4.2 µg/mL) whilst it was moderately active in term of the suppression of linoleic acid oxidation (64.6%). All the tested oil concentrations, however, revealed a significant scolicidal activity on the protoscoleces of hydatid cysts. The anticancer activity of O. minutiflorum oil was found more in HepG2 cells followed by A-549 and MCF-7 cells and this was evidenced by the LC50 values. ConclusionsThe present work strongly suggests that O. minutiflorum could serve as a potential source of scolicidal and anticancer agents.

Cell-mediated and humoral immune profile to hydatidosis among naturally infected farm animals.

Background and Aim:Cystic echinococcosis (CE) is a widespread parasitic disease caused by Echinococcus granulosus tapeworm infect different intermediate hosts including sheep, cattle, and camels. The intermediate host’s immune response to the hydatid cyst is still conflict and complex. The current study was designed to evaluate the immune response in sera of hydatid naturally infected sheep, cattle, and camels in the form of features of inflammatory cell infiltrations, levels of Th1 and Th2 cytokines, besides the humoral specific immunoglobulin G (IgG) responses.Materials and Methods:Thirty-nine sheep, 74 cattle, and 79 camels’ sera were collected and considered as CE naturally infected and ten samples from each species were graded as non-infected. Lung specimens were collected for histopathological examination. The quantitative concentrations of tumor necrosis factor-α, interleukin (IL)-6, IL-4, and IL-10 were determined. Different antigens were prepared from hydatid cyst; hydatid cyst fluid of lung origin hydatid cyst fluid of liver origin, hydatid cyst protoscoleces of lung origin (HCP-g), hydatid cyst protoscoleces of liver origin, hydatid cyst germinal layer of lung origin, and hydatid cyst germinal layer of liver origin; and characterized by gel electrophoresis and Western blotting analysis. The total specific IgG level against E. granulosus infection was measured using an indirect enzyme-linked immunosorbent assay.Results:The results indicated that the cellular immune response in the infected tissues was characterized by inflammatory cell penetration. The pro-inflammatory Th1 cytokine profile was predominant in infected animals in comparison with non-infected ones. However, the humoral immune response was seen as a high level of IgG in infected animals. The presented data approved that the HCP-g antigen could be considered as a delegate antigen for all other prepared antigens with an immunoreactive band at molecular weights 32 kDa.Conclusion:This study provides a fundamental insight into the events that manipulate cellular and humoral immune profiles in an intermediate host; sheep, cattle, and camel that naturally infected with CE. Hence, it was concluded that CE is a constant disease and confirm the reactivity Th1 in combating hydatid cyst. Besides, it could lead to the activation of the humoral immune response in the form of a high level of IgG.

The High Potential of Ozone Gas to Inactivate Echinococcus granulosus Protoscoleces During Hydatid Cyst Surgery.

In medicine, ozone therapy is effectively used in a broad spectrum of diseases. Reviews have shown that ozone gas demonstrates potent antimicrobial effects against a wide range of pathogenic microorganisms, such as oral bacteria, fungi, viruses, and parasite even in resistant strains. The present investigation was designed to assess the protoscolicidal effects of ozone gas on hydatid cysts protoscoleces in vitro and in vivo. Hydatid cyst protoscoleces were acquired from sheep livers that were slaughtered at Kerman slaughterhouse, Iran. The viability of protoscoleces was assessed by the eosin exclusion examination after exposure with ozone gas for 1 to 14 min in vitro and ex vivo. In this study, in vitro assay showed that ozone gas at the concentration of 20 mg/L killed 85 and 100% of hydatid cyst protoscoleces after 4 and 6 min of treatment, respectively. However, in the ex vivo analysis, a longer time was needed to confirm a potent protoscolicidal activity such that ozone gas after an exposure time of 12 min, 100% of the protoscoleces were killed within the hydatid cyst. In conclusion, the findings of the present study showed that ozone gas at low concentrations (20 mg/L) and short times (4-6 min) might be used as a novel protoscolicidal drug for use in hydatid cyst surgery. However, more clinical surveys are required to discover the precise biological activity of ozone gas in animal and human subjects.

Efficacy and safety of Curcuma longa essential oil to inactivate hydatid cyst protoscoleces

BackgroundOne of the most important ways to cure hydatid cysts is surgery. Today, available chemical drugs have shown severe complications during hydatid cyst surgery. Here we investigated theefficacy and safety of Curcuma longa (C. longa) essential oil (CLEO) against hydatid cysts protoscoleces.MethodsHere, the collected protoscoleces from fertile hydatid cysts were exposed to different concentrations of the CLEO (50–200 μl/mL) for 5–30 min in vitro and ex vivo. In addition, male NIH mice (48 mice) were applied to examine the safety of CLEO.ResultsAll protoscoleces were completely killed in 5th min of treatment to the doses of 200 μl /mL of CLEO. On the other hand, in the 10th min of treatment, CLEO entirely killed 100% of protoscoleces at the dose of 100 μL/mL. Other doses of CLEO, but, revealed a postponed activity. Although CLEO at the doses of 50, and 100 μL/mL exhibited no similar effect in the ex vivo analysis; but, at the dose of 200 μL/mL and an exposure time of 5 min, approximately 100% of protoscoleces were destroyed into the cyst. We found that after intra-peritoneal injection of the CLEO for 14 days, although blood and biochemical parameters were changed; but there was no significant difference in comparison with the control group (p > 0.05).ConclusionThis research demonstrated that C. longa revealed the promising scolicidal effects against protoscoleces in vitro and in vivo, of course, after additional tests; it might be considered as an herbal scolicidal drug in order to decrease the threat of discharge of protoscoleces through hydatid cyst surgery. Nevertheless, supplementary studies will be desired to prove the current findings by examination the essential oil in a clinical setting.

The Lethal Effect of a Nano Emulsion of Satureja hortensis Essential Oil on Protoscoleces and Germinal Layer of Hydatid Cysts

New scolicidal agents and novel therapeutic drugs are essential for better management of the zoonotic infection, hydatid disease. This study evaluated the effect of a nanoemulsion (NE) of Satureja hortensis essential oil (SHEO) on protoscoleces and germinal layer of hydatid cysts. This study was conducted from July to October 2016 in Shiraz University, Shiraz, Iran. Gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS) were performed to identify the main components of SHEO. To determine the scolicidal power of the NE of SHEO, live protoscoleces of hydatid cysts were exposed to two concentrations (1 and, 2 mg/mL) of the NE and incubated at 37 °C for 10 and 20 min. To evaluate the anti-hydatid effect of the NE of SHEO, the collected hydatid cysts from the abdominal cavities of the experimentally infected mice were immersed in the NE (0.5 mg/ml) and incubated at 22 °C for 24 h. Carvacrol and γ-terpinene were the major components of the SHEO. NE of SHEO at the concentrations of 1 and 2 mg/mL showed 100% scolicidal power after 20 and 10 min respectively. Exposure of the hydatid cysts to the NE of SHEO resulted in crumpling of their germinal layer and detachment of this layer from the laminated layer. NE of SHEO showed a strong scolicidal activity as well as a profound lethal effect on the germinal layer of hydatid cysts. Accordingly, this product may be used as a natural scolicidal agent in hydatid cyst surgery. Furthermore, it may be used as a therapeutic tool for treatment of hydatid disease.

In Vitro and in Vivo Scolicidal Activities of Holothuria leuco-spilota Extract and CeO2 Nanoparticles against Hydatid Cyst

We aimed to investigate the scolicidal effects of Holothuria leucospilota extract and CeO2 nanoparticles against protoscoleces of hydatid cysts in-vitro and in-vivo. Hydatid cysts were collected from, Urmia slaughterhouses between years 2016-2017 and the hydatid fluid aspirated from the fertile cysts. Various concentration of H. leucospilota extract, CeO2 NPs and combination of CeO2-NPs/H. leucospilota were used for 10-60 min to evaluate the viability of protoscoleces by 0.1% eosin method. CASPASE -3 activity measured for assessment of cell apoptosis in treated protoscoleces. BALB/c mice were infected intraperitoneally with 2000 viable protoscoleces and treated daily for 4 wk by intragastrical inoculation with H. leucospilota, CeO2 NPs, combination of CeO2 NPs/H. leucospilota and Albendazole. Cyst development was macroscopically analyzed. H. leucospilota extract and combination of CeO2 NPs/H. leucospilota have potent scolicidal activity at concentration of 20 mg/ml and 15 mg/ml after 60 min treatment. Maximum caspase-3 activity was observed when protoscoleces expose with H. leucospilota and combination H. leucospilota & CeO2 NPs. After treatment of cyst infected mice with extract and CeO2 NPs, combination of CeO2 NPs/H leucospilota and albendazole, a significant decrease in number of cysts, size and volume of cyst (P<0.05) was observed. This result shows an antihydatic and scolicidal effects of H. leucospilota extract and CeO2 NPs.

In vitro and ex vivo scolicidal effects of Olea europaea L. to inactivate the protoscolecs during hydatid cyst surgery.

BackgroundNowadays, surgery is considered as one of the most important treatments for hydatidosis. Due to laceration, the cyst and spread of the content within it (protoscoleces) during the surgery that can put the patient at the risk of re-infection, anaphylaxis shock and even death, surgeons use some chemical drugs as protoscolicidal agents. The study is aimed to evaluate the scolicidal effects of olive (Olea europaea L.) leaf extract on hydatid cyst protoscoleces in vitro and ex vivo.MethodsAfter the collection of protoscoleces from sheep livers infected with fertile hydatid cysts, they were treated with various concentrations of olive leave extract (75–300 mg/mL) for 5–30 min in vitro and ex vivo. Finally, the mortality of protoscoleces was assessed by the eosin exclusion test (0.1% eosin staining).ResultsThe mean of the mortality of protoscoleces was 100% after 10 min of incubation with the concentration of 300 mg/ml of O. europaea leaves extract. On the other hand, the mean of the mortality of protoscoleces after 20 min of incubation with the concentration of 150 mg/ml of O. europaea leaves extract was 100%. After injection of O. europaea leaves extract directly into the hydatid cyst (ex vivo), the mean of the mortality of protoscoleces was 100% after 12 and 25 min of incubation with the concentration of 300 and 150 mg/ml of O. europaea leaves extract, respectively; indicating that the extract requiring a further time to display a potent protoscolicidal effects.ConclusionBased on the findings of the study, it can be concluded that the extract of olive leaf had a significant scolicidal activity on hydatid cyst protoscoleces. However, further research, especially in human and animal subjects, are required to reach this conclusion.

In vitro evaluation on the scolicidal effect of Myrtus communis L. and Tripleurospermum disciforme L. methanolic extracts

Hydatid disease, a zoonotic disease, is still endemic in many developing countries that is caused by the metacestode of Echinococcus (E.) granulosus. Surgical management is one of the best choices for the treatment of the hydatidosis and using effective scolicidal agents during hydatid surgery is essential to prevent the secondary infection. The aim of the present in vitro study was to evaluate the scolicidal effect of the methanolic extract of Myrtus communis and Tripleurospermum disciforme against protoscoleces of hydatid cyst. Protoscoleces of E. granulosus were aspirated aseptically from infected livers. Various concentrations of M. communis and T. disciforme extracts at different exposure times were examined for their scolicidal activity. Normal saline and silver nitrate were used as negative and positive groups, correspondingly. The viability of protoscoleces was evaluated by 0.1% eosin. The result of the current study indicated that the highest scolicidal effect (100%) of M. communis was obtained at 100 and 50 mg/ml concentrations and LC50 in 10, 20 and 30 min were 11.64 mg/ml, 7.62 mg/ml, and 6.47 mg/ml respectively. The scolicidal activity of T. disciforme was negligible even at high concentration. The findings have shown that the scolicidal activity of M. communis against echinococcosis protoscoleces was strong, while the T. disciforme extract showed fewer effects. However, further studies are required for identification of the active ingredients in the extract and its safety on cells in effective concentrations.

The scolicidal activity of the essential oil obtained from the needles of Pinus nigra Arn. subsp. pallasiana (Lamb.) Holmboe on hydatid cyst

Ethnopharmacological relevanceThis study aimed to evaluate the in vitro activity of the essential oil of Pinus nigra subsp. pallasiana against protoscoleces of Echinococcus granulosus sensu lato. In traditional Turkish folk medicine, the essential oils and resin of P. nigra and P. sylvestris are used externally for wound healing. They are also used in cattle for treating worms and as boiling essence against cold and stomachache. Protoscoleces were aseptically aspirated from sheep livers having hydatid cysts. Materials and methodsThree concentrations (10, 30, and 50 mg/mL) of the essential oil from P. nigra subsp. pallasiana cones were used for 10–60 min. The essential oil was applied to protoscoleces of hydatid cyst to determine the scolicidal activity of the essential oil. Viability of protoscoleces was observed by 0.1% eosin staining. ResultsThe rate of dead protoscoleces was found to be 2.27% in the control group, which was not treated with essential oil. The rate of dead protoscoleces increased to 61.69%, 69.39%, 75.98% and 71.76% respectively after, 10, 20, 30, and 60 min when the protoscoleces were exposed to the P. nigra subsp. pallasiana essential oil at 10 mg/mL concentration. When protoscoleces were exposed to the essential oil of P. nigra subsp. pallasiana for 10, 20, 30, and 60 min at 30 mg/mL concentration, the mortality rates of protoscoleces were determined as 3.82%, 91.33%, 94.67% and 96.74%, respectively. The mortality rate of 100% was determined at 50 mg/mL concentration after 60 min of exposure. According to statistical analysis, P. nigra subsp. pallasiana essential oil at all concentrations had significant scolicidal effects compared to the control group at all exposure times. ConclusionsThe essential oil of P. nigra subsp. pallasiana needles showed scolicidal activity and it could be a new scolicidal agent for using in hydatid cyst treatment.

Lethal effects of gold nanoparticles on protoscolices of hydatid cyst: in vitro study

Recurrence of hydatidosis may results from cyst fluid spillage during surgery. To inactivate the cyst content, different scolicidal agents have been introduced. Nevertheless, novel and more effective treatments are needed due to the associated complications. In the current study, we examined the scolicidal effects of gold nanoparticles on Echinococcus granulosus protoscoleces. Hydatid cysts of sheep liver were collected in this study. The cyst fluid containing protoscoleces was aspirated aseptically. The protoscoleces were exposed to gold nanoparticles at different concentrations (250, 500, 1000, 2000, and 4000 μg/mL). Eosin staining method was applied to determine the viability of protoscoleces at 5, 10, 20, 30, and 60 min. Moreover, cytotoxicity analysis, scanning electron microscopy (SEM) of ultrastructural changes, DNA fragmentation, were performed, and protoscoleces were studied following treatment. The significant scolicidal effects of gold nanoparticles were observed at all concentrations compared to the control group. A total of 4000 μg/mL of gold nanoparticles could destroy 76% of protoscoleces in 60 min. Cytotoxic effects of gold nanoparticle on J774 macrophage cell line, in minimum and maximum concentration (500 and 4000 μg/mL), were 1 and 11%, respectively. After treatment of protoscoleces with gold nanoparticles in different times and concentrations, considerable alteration in size, ultrastructure changes in tegument and shape of sucker, as well as DNA fragmentation of parasite were seen. Based on the results, gold nanoparticles could exert scolicidal effects on hydatid cyst protoscoleces; therefore, they can be applied in hydatid cyst treatment.

BALB/c Mice Immunity to Hydatidosis Induced by In-vitro Reared Echinococcus granulosus Adult Worm Antigens.

Echinococcosis is a zoonotic parasitic disease caused by the larval stage of Echinococcus granulosus. Several native and recombinant antigens, derived from different stages of E. granulosus life cycle, have been used for vaccine trials. In vitro reared adult worms are good candidates for vaccination as they do not produce fertile egg/s and do not have any risk of contamination for researchers. To evaluate different antigens derived from in vitro reared E. granulosus adult worms for the immunization of BALB/c mice against secondary hydatidosis. Viable protoscoleces (PCSs) of sheep hydatid cyst were cultivated in S.10E.H media. Excretory secretory (E/S) and crude antigens were prepared from reared adult worms. A total of fifty BALB/c mice, each 8-weeks-old, were divided into 5 groups of 10 mice. Three groups were subcutaneously immunized with crude, E/S and immunodominant antigens on days 1 and 28. The fourth group received only PBS and the fifth group had no injection. Three weeks following the second immunization, all groups were challenged, intraperitoneal, with viable PSCs. After the autopsy of the mice and opening their abdominal wall, cysts were counted and measured followed by histopathological observations. The highest protective immunity (98.7%) against hydatidosis was induced by crude antigen, followed by E/S and immunodominant antigens. Antigens (crude antigens in particular) derived from in vitro reared E. granulosus adult worms, and their different protein components are suitable candidates for the vaccination of intermediate hosts against hydatidosis.