Significant Reduction In Worm Burden Research Articles

Eficácia terapêutica da Ferula asafetida contra Hymenolepis nana

ABSTRACT Hymenolepis nana is a common intestinal tapeworm that affects humans. Drugs, including praziquantel (PZQ), are essential for managing this infection. Natural products are now considered as an alternative agent to control hymenolepiasis. Three doses of the herb Ferula asafetida (FAH) (100-150-200 mg/ml) were used to assess the appropriate dose and right time to eliminate H. nana. It was found that 150 mg/ml gives no movement in 5 min and is an appropriate dose affecting H. nana. This study showed that FAH completely controls mature worms. Treatment with FAH induced a significant reduction in worm burden and complete healing after 14 days relative to a single dose PZQ drug. Moreover, histological studies for the infected-treated mice with FAH demonstrated improvement in the intestinal tissue and less accumulation of inflammation relative to those treated with PZQ. In addition, the cestodal infection significantly upregulated the inflammatory cytokines. This increase in mRNA expression of TNF-��, iNOS, and IL-2 was 6.80, 5.65, and 8.95-fold, respectively, which significantly downregulated upon treatment. Collectively, F. asafetida is a promising medicinal plant with anti-cestodal and anti-inflammatory activities and could be used for the treatment of hymenolepiasis.

Evaluation of schistosomula lung antigen preparation and soluble egg antigen vaccines on experimental schistosomiasis mansoni.

Schistosomiasis causes significant morbidity and mortality worldwide. This study aimed to assess the effect of schistosomula lung antigen preparation (SLAP) and soluble egg antigen (SEA) on a murine schistosomiasis mansoni model. Ninety laboratory-bred male Swiss albino mice were divided into 6 groups. Two doses of the vaccine were given at 2-week intervals. All mice were subcutaneously infected with 80±10 Schistosoma mansoni cercariae 2 weeks after the last vaccination dose. They were sacrificed 7 weeks post-infection. Parasitological and histopathological studies were conducted to assess the effect of inoculated antigens (single or combined). The results showed that the combination of SLAP and SEA (combination group) led to a significant reduction in worm burden (65.56%), and liver and intestine egg count (59% and 60.59%, respectively). The oogram pattern revealed a reduction in immature and mature eggs (15±0.4 and 10±0.8, respectively) and an increased number of dead eggs in the combination group (P<0.001). In terms of histopathological changes, the combination group showed notably small compact fibrocellular egg granuloma and moderate fibrosis in the liver. A high percentage of destroyed ova was observed in the intestine of the combination group. This study demonstrates for the first time the prophylactic effect of combined SLAP and SEA vaccine. The vaccine induced a significant reduction in the parasitological and pathological impacts of schistosomiasis mansoni in hepatic and intestinal tissues, making it a promising vaccine candidate for controlling schistosomiasis.

Loading praziquantel within solid lipid nanoparticles improved its schistosomicidal efficacy against the juvenile stage

• Characterization of praziquantel solid lipid nanoparticles was done. • Loading of praziquantel within solid lipid nanoparticles improve its effect against the invasive and juvenile of S.mansoni . • This improvement was obvious by significant reduction in worm burden, intestinal egg count and hepatic granuloma size and number. • Marked changes in tegument by scanning electron microscopy were also evident.

Characterization of aspartyl aminopeptidase from Schistosoma japonicum

The tegument of schistosomes is the interface between the worm and the host environment. Some molecules distributed on the tegument participate in host–parasite interactions. Aspartyl aminopeptidase (AAP), identified on the tegument of Schistosoma japonicum (S. japonicum), facilitate protein turnover by acting in concert with other aminopeptidases. In this study, the gene encoding S. japonicum aspartyl aminopeptidase (SjAAP) was cloned, expressed and characterized. Quantitative real-time PCR analysis showed that SjAAP was expressed in all studied developmental stages. The transcript level was higher in 8, 14, 21, and 28 days old worms than the other detected stages. Moreover, the level of expression in 42-day-old male worms was significantly higher than that in females. The recombinant SjAAP (rSjAAP) was expressed as both supernatant and inclusion bodies in Escherichia coli BL21 cells. The enzymatic activity of rSjAAP was 4.45 U/mg. The Km and Vmax values for H-Asp-pNA hydrolysis were discovered to be 5.93 mM and 0.018 mM·min–1. Immunofluorescence analysis revealed that SjAAP is primarily distributed on the tegument and parenchyma of schistosomes. Western blot showed that rSjAAP possessed good immunogenicity. Although specific antibodies were produced in BALB/c mice vaccinated with rSjAAP emulsified with ISA 206 adjuvant, no significant reduction of worm burden and number of eggs in the liver was observed. Therefore, rSjAAP may not be suitable to act as a potential vaccine candidate against schistosomiasis japonica in mice. However, this study provides some foundation for further exploration of the biological function of this molecule.

Mucosal Vaccination With Recombinant Tm-WAP49 Protein Induces Protective Humoral and Cellular Immunity Against Experimental Trichuriasis in AKR Mice.

Trichuriasis is one of the most common neglected tropical diseases of the world’s poorest people. A recombinant vaccine composed of Tm-WAP49, an immunodominant antigen secreted by adult Trichuris stichocytes into the mucosa of the cecum to which the parasite attaches, is under development. The prototype is being evaluated in a mouse model of Trichuris muris infection, with the ultimate goal of producing a mucosal vaccine through intranasal delivery. Intranasal immunization of mice with Tm-WAP49 formulated with the adjuvant OCH, a truncated analog of alpha-GalCer with adjuvanticity to stimulate natural killer T cells (NKT) and mucosal immunity, induced significantly high levels of IgG and its subclasses (IgG1 and IgG2a) in immunized mice. This also resulted in a significant reduction of worm burden after challenge with T. muris-infective eggs. The addition of QS-21 adjuvant to this vaccine formulation further reduced worm counts. The improved protection from the dual-adjuvanted vaccine correlated with higher serum antibody responses (IgG, IgG1, IgG2a, IgA) as well as with the induction of antigen-specific IgA in the nasal mucosa. It was also associated with the robust cellular responses including functional subsets of CD4 T cells producing IL-4, and cytotoxic CD8 T cells expressing granzyme B. The worm reduction achieved by mucosal immunization was higher than that induced by subcutaneous immunization. Intranasal immunization also induced a significantly higher nasal mucosa-secreted antigen-specific IgA response, as well as higher functional cellular responses including CD4+IL4+ (Th1) and CD8+GnzB+ (Th2) T cells, and antigen-specific INFγ-producing T cells in both spleen and MLNs and antibody-producing B cells (CD19+B220+/B220+GL7+). Mucosal immunization further induced long-term T lymphocyte memory with increased central (CD62L+CD44+) and effector (CD62L-CD44+) memory subsets of both CD4 and CD8 T cells at 60 days after the last immunization. In summary, intranasal immunization with recombinant Tm-WAP49 protein induced strong protection versus murine trichuriasis. It represents a promising vaccination approach against intestinal nematodes.

Efficacy of carvacryl acetate in vitro and following oral administration to mice harboring either prepatent or patent Schistosoma mansoni infections.

Schistosomiasis is a major public health problem that afflicts more than 240 million individuals globally, particularly in poor communities. Treatment of schistosomiasis relies heavily on a single oral drug, praziquantel, and there is interest in the search for new antischistosomal drugs. This study reports the anthelmintic evaluation of carvacryl acetate, a derivative of the terpene carvacrol, against Schistosoma mansoni ex vivo and in a schistosomiasis animal model harboring either adult (patent infection) or juvenile (prepatent infection) parasites. For comparison, data obtained with gold standard antischistosomal drug praziquantel are also presented. Initially in vitro effective concentrations of 50% (EC50) and 90% (EC90) were determined against larval and adult stages of S. mansoni. In an animal with patent infection, a single oral dose of carvacryl acetate (100, 200, or 400mg/kg) caused a significant reduction in worm burden (30-40%). S. mansoni egg production, a process responsible for both life cycle and pathogenesis, was also markedly reduced (70-80%). Similar to praziquantel, carvacryl acetate 400mg/kg had low efficacy in pre-patent infection. In tandem, although carvacryl acetate had interesting in vitro schistosomicidal activity, the compound exhibited low efficacy in terms of reduction of worm load in S. mansoni-infected mice.

Oral Peptide Vaccine against Hookworm Infection: Correlation of Antibody Titers with Protective Efficacy.

Approximately 0.4 billion individuals worldwide are infected with hookworm. An effective vaccine is needed to not only improve the health of those affected and at high risk, but also to improve economic growth in disease-endemic areas. An ideal anti-hookworm therapeutic strategy for mass administration is a stable and orally administered vaccine. Oral vaccines are advantageous as they negate the need for trained medical staff for administration and do not require strict sterility conditions. Vaccination, therefore, can be carried out at a significantly reduced cost. One of the most promising current antigenic targets for hookworm vaccine development is the aspartic protease digestive enzyme (APR-1). Antibody-mediated neutralization of APR-1 deprives the worm of nourishment, leading to reduced worm burdens in vaccinated hosts. Previously, we demonstrated that, when incorporated into vaccine delivery systems, the APR-1-derived p3 epitope (TSLIAGPKAQVEAIQKYIGAEL) was able to greatly reduce worm burdens (≥90%) in BALB/c mice; however, multiple, large doses of the vaccine were required. Here, we investigated a variety of p3-antigen conjugates to optimize antigen delivery and establish immune response/protective efficacy relationships. We synthesized, purified, and characterized four p3 peptide-based vaccine candidates with: (a) lipidic (lipid core peptide (LCP)); (b) classical polymeric (polymethylacrylate (PMA)); and (c) novel polymeric (polyleucine in a branched or linear arrangement, BL10 or LL10, respectively) groups as self-adjuvanting moieties. BL10 and LL10 induced the highest serum anti-p3 and anti-APR-1 IgG titers. Upon challenge with rodent hookworms, the highest significant reduction in worm burden was observed in mice immunized with LL10. APR-1-specific serum IgG titers correlated with worm burden reduction. Thus, we provide the first vaccine-triggered immune response-protection relationship for hookworm infection.

Effect of celecoxib against different developmental stages of experimental Schistosoma mansoni infection

Due to the high prevalence of schistosomiasis and the wide use of praziquantel solely for mass drug administration to control the disease, there is a great concern about the potential emergence of reduced susceptibility strains. This, together with the concern that praziquantel is ineffective against juvenile worms highlight the importance of developing an alternative anti-schistosomal drug. Using nonsteroidal anti-inflammatory drugs against schistosome infection is considerable. The present study evaluated the effect of oral administration of five days celecoxib regimen (20 mg/kg/day) against different developmental stages of Schistosoma mansoni infection. This regimen induced significant reduction in worm burden, tissue egg count, individual female fecundity and the mean percentage of immature and mature eggs with increased mean percentage of dead eggs. More importantly, celecoxib was more potent than praziquantel in all these parasitological parameters (except in the worm burden when given against the adult stage where the difference was statistically non-significant). Scanning and transmission electron microscopy of the adult worms revealed severe tegumental damage, laceration of the muscular layers and oedema of the syncytial layer. There was disruption of the testicular, ovarian and vitelline glandular tissues with signs of apoptosis and abnormalities of the spermatozoa and the oocytes. Additionally, celecoxib induced reduction in the number and the size of the hepatic granulomata and also amelioration of the hepatic tissue pathology.

Fifteen Years of Sm-p80-Based Vaccine Trials in Nonhuman Primates: Antibodies From Vaccinated Baboons Confer Protection in vivo and in vitro From Schistosoma mansoni and Identification of Putative Correlative Markers of Protection.

Recent advances in systems biology have shifted vaccine development from a largely trial-and-error approach to an approach that promote rational design through the search for immune signatures and predictive correlates of protection. These advances will doubtlessly accelerate the development of a vaccine for schistosomiasis, a neglected tropical disease that currently affects over 250 million people. For over 15 years and with contributions of over 120 people, we have endeavored to test and optimize Sm-p80-based vaccines in the non-human primate model of schistosomiasis. Using RNA-sequencing on eight different Sm-p80-based vaccine strategies, we sought to elucidate immune signatures correlated with experimental protective efficacy. Furthermore, we aimed to explore the role of antibodies through in vivo passive transfer of IgG obtained from immunized baboons and in vitro killing of schistosomula using Sm-p80-specific antibodies. We report that passive transfer of IgG from Sm-p80-immunized baboons led to significant worm burden reduction, egg reduction in liver, and reduced egg hatching percentages from tissues in mice compared to controls. In addition, we observed that sera from Sm-p80-immunized baboons were able to kill a significant percent of schistosomula and that this effect was complement-dependent. While we did not find a universal signature of immunity, the large datasets generated by this study will serve as a substantial resource for further efforts to develop vaccine or therapeutics for schistosomiasis.

THE POSSIBLE ANTISCHISTOSOMAL EFFECT OF ALLICIN IN SCHISTOSOMA MANSONI INFECTED ALBINO MICE

Schistosomiasis is endemic mainly in Africa and the Middle East Countries, causing acute and chronic clinical pathogenicity. This study evaluated the potential antischistosomal efficiency of allicin against S. mansoni infected mice. Twenty CD1 male Albino mice were divided into two groups (10 mice/group). G1 (infected and treated with 0.9% NaCl), and G2 (infected and treated with 20mL/kg of allicin). Seven weeks post-infection, each group was intraperitoneally injected with allicin (3 times/week), sacrificed at the end of the 8th-week. Liver was harvested for studing worm burden, egg load, and oogram pattern. The recovered worms from both groups were subject to SEM and DNA fragmentation tests to monitor the differences. The results showed a significant reduction in worm burden and changes in oogram pattern ofG2 mice as compared to G1 (control). Allicin gave marked deformation of oral, ventral suckers and atrophy of the tubercles as well as marked fragmentation of the worms DNA.

THE POSSIBLE ANTISCHISTOSOMAL EFFECT OF ALLICIN IN SCHISTOSOMA MANSONI INFECTED ALBINO MICE

Schistosomiasis is endemic mainly in Africa and the Middle East Countries, causing acute and chronic clinical pathogenicity. This study evaluated the potential antischistosomal efficiency of allicin against S. mansoni infected mice. Twenty CD1 male Albino mice were divided into two groups (10 mice/group). G1 (infected and treated with 0.9% NaCl), and G2 (infected and treated with 20mL/kg of allicin). Seven weeks post-infection, each group was intraperitoneally injected with allicin (3 times/week), sacrificed at the end of the 8th-week. Liver was harvested for studing worm burden, egg load, and oogram pattern. The recovered worms from both groups were subject to SEM and DNA fragmentation tests to monitor the differences. The results showed a significant reduction in worm burden and changes in oogram pattern ofG2 mice as compared to G1 (control). Allicin gave marked deformation of oral, ventral suckers and atrophy of the tubercles as well as marked fragmentation of the worms DNA.

Promethazine exhibits antiparasitic properties in vitro and reduces worm burden, egg production, hepato-, and splenomegaly in a schistosomiasis animal model.

The treatment and control of schistosomiasis, a neglected disease that affects more than 200 million people worldwide, rely on the use of a single drug, praziquantel. A vaccine has yet to be developed and since new drug design and development is a lengthy and costly process, drug repurposing is a promising strategy. In this study, the efficacy of promethazine, a first-generation antihistamine, was evaluated against Schistosoma mansoni ex vivo and in a murine model of schistosomiasis. In vitro assays demonstrated that promethazine affected parasite motility, viability, and it induced severe tegumental damage in schistosomes. The LC50 of the drug was 5.84 μM. Similar to promethazine, schistosomes incubated with atropine, a classical anticholinergic drug, displayed reduced motor activity. In an animal model, promethazine treatment was introduced at an oral dose of 100 mg/kg for five successive days at different intervals from the time of infection, for the evaluation of the stage-specific susceptibility (pre-patent and patent infections). Various parasitological criteria indicated the in vivo antischistosomal effects of promethazine: there were significant reductions in worm burden, egg production, and hepato- and splenomegaly. The highest worm burden reduction was achieved with promethazine in patent infections (> 90%). Taken together, considering the importance of the repositioning of drugs in infectious diseases, especially those related to poverty, our data revealed the possibility of promethazine repositioning as an antischistosomal agent.

The phosphodiesterase-4 inhibitor roflumilast impacts Schistosoma mansoni ovipositing in vitro but displays only modest antischistosomal activity in vivo

Praziquantel (PZQ) is the sole drug used to treat schistosomiasis, and the probability of developing resistance is growing the longer it is relied upon, justifying the search for alternatives. Phosphodiesterases (PDEs), particularly the PDE4 family, have attracted considerable attention as drug targets, including in Schistosoma mansoni, and especially SmPDE4A. This study investigates the potential antischistosomal activity of human PDE4 and potent SmPDE4A inhibitor roflumilast, either alone or combined with PZQ. In vitro, roflumilast resulted in a significant, concentration-dependent reduction in egg production but not of worm viability. In vitro exposure to roflumilast in combination with a low concentration of PZQ was less effective than PZQ alone, pointing to antagonism. S. mansoni-infected mice treated with roflumilast showed significant reductions in worm burden (27%) as well as hepatic and intestinal egg burdens (~28%) two weeks post treatment. Scanning EM of worms isolated from roflumilast-treated and untreated mice did not reveal noticeable changes to their tegument. S. mansoni-infected mice treated with a fixed dosage of roflumilast and a variable dosage of PZQ resulted in a higher reduction in worm burden, reduced hepatic egg counts, absence of immature eggs and a marked increase in dead eggs, compared to PZQ alone. However, the combination resulted in increased animal mortality, probably attributable to pharmacodynamic interactions between the two drugs. Although this study marks the first report of in vivo antischistosomal potential by a PDE inhibitor, an important proof of concept, we conclude that the antischistosomal effects of roflumilast are insufficient to warrant further development.

Evaluation of the effect of some plant molluscicides on the infectivity of Schistosoma mansoni cercariae

Objectives To study the effect of sublethal concentrations (LC50) of methanol extracts from Solanum nigrum and Callistemon citrinus leaves on the infectivity of Schistosoma mansoni cercariae by an in-vivo study. Background The high cost of synthetic molluscicides and their negative impacts on the environment necessitated search for an alternative approach by using plant extracts for the control of schistosomiasis. Using molluscicides could be more effective if these agents also possess cercaricidal activity. Materials and methods Methanol extracts of S. nigrum and C. citrinus leaves were prepared and their LC50on S. mansoni cercariae were determined. The experimental mice used in this study were divided into three groups. Group I including mice infected with S. mansoni cercariae preexposed to S. nigrum. Group II included mice infected with S. mansoni cercariae preexposed to C. citirnus. Group III included mice infected with nonexposed S. mansoni cercariae (control group). The attenuating effect of the plants on cercariae was evaluated through different parameters including parasitological, histopathological, and scanning electron microscopic examinations. Results LC50of S. nigrum and C. citrinus methanol extracts were 25 and 40 mg/l, respectively. Parasitological results showed a significant reduction in worm burden, mean number of ova per gram tissue in mice of group I and group II in comparison with those of group III (control group). These results were associated with decrease in the size and number of hepatic granulomas. All these obtained results were more significant with S. nigrum than C. citrinus. Obvious tegmental damages were also detected by scanning electron microscopic in both groups (I and II). Conclusion Methanol extracts of both S. nigrum and C. citrinus leaves exhibited cercariicidal activity but S. nigrum was found to be more potent. This study provides a considerable scope in exploiting local indigenous resources for schistosomiasis control.

Trichuris muris whey acidic protein induces type 2 protective immunity against whipworm.

Human whipworm (Trichuris trichiura) infects approximately 1 in 15 people worldwide, representing the leading infectious cause of colitis and subsequent, inflammatory bowel disease (IBD). Current control measures focused on mass deworming have had limited success due to low drug efficacies. Vaccination would be an ideal, cost-effective strategy to induce protective immunity, leading to control of infection and transmission. Here we report the identification of whey acidic protein, a whipworm secretory protein, as a strong immunogen for inducing protective efficacy in a surrogate mouse T. muris infection model. The recombinant WAP protein (rTm-WAP49), as well as a single, highly conserved repeat within WAP (fragment 8) expressed as an Na-GST-1 fusion protein (rTm-WAP-F8+Na-GST-1), generate a strong T helper type 2 (Th2) immune response when delivered as subcutaneous vaccines formulated with Montanide ISA 720. Oral challenge with T. muris infective eggs following vaccination led to a significant reduction in worm burden of 48% by rTm-WAP49 and 33% by rTm-WAP-F8+Na-GST-1. The cellular immune correlates of protection included significant antigen-specific production of Th2 cytokines IL-4, IL-9, and IL-13 by cells isolated from the vaccine-draining inguinal lymph nodes, parasite-draining mesenteric lymph nodes, and spleen in mice vaccinated with either rTm-WAP49 or rTm-WAP-F8+Na-GST-1. The humoral immune correlates included a high antigen-specific ratio of IgG1 to IgG2a, without eliciting an IgE-mediated allergic response. Immunofluorescent staining of adult T. muris with WAP antisera identified the worm’s pathogenic stichosome organ as the site of secretion of native Tm-WAP protein into the colonic mucosa. Given the high sequence conservation for the WAP proteins from T. muris and T. trichiura, the results presented here support the WAP protein to be further evaluated as a potential human whipworm vaccine candidate.

Efficacy of Ozoroa pulcherrima Schweinf methanolic extract against Schistosoma mansoni-induced liver injury in mice

The roots of Ozoroa pulcherrima Schweinf are used in traditional medicine to treat intestinal helminthiasis. The aim of this study was to assess the effect of Ozoroa pulcherrima roots methanolic extract (OPME) on liver injury induced by Schistosoma mansoni in mice. A preliminary phytochemical study of OPME was conducted. OPME was given daily and orally to S. mansoni-infected mice at 100, 200 or 400 mg/kg for 28 days, starting from the 36th day post-infection. Praziquantel was used as reference drug. Non-infected and infected-untreated mice served as controls. Worm burden and egg output, transaminases, total bilirubin, alkaline phosphatase and total protein; as well as malondialdehyde, catalase and reduced glutathione were evaluated. In OPME, total phenolic was 79.61 ± 0.25 mg gallic acid equivalent/g, while total flavonoid was 7.98 ± 0.04 mg rutin equivalent/g. Treatment of S. mansoni-infected mice with OPME produced significant reduction of worm burden and ova count in the faeces, liver and intestine. Significant reduction of alanine aminotransferase activity (p < 0.001) as well as significant increase of total protein content (p < 0.001) was recorded after OPME treatment at all doses. Total bilirubin level was also reduced (p < 0.01). Administration of OPME at all doses corrected the high malondialdehyde level (p < 0.001) induced by the infection. At 200 mg/kg, catalase activity and reduced glutathione concentration were significantly increased (p < 0.001). OPME at 200 mg/kg showed moderate schistosomicidal effect, but was effective as the standard drug praziquantel in restoring the liver function after S. mansoni infection.

Suppression of VAMP2 Alters Morphology of the Tegument and Affects Glucose uptake, Development and Reproduction of Schistosoma japonicum

Schistosomiasis caused by schsitosomes is a serious global public health concern. The tegument that surrounds the worm is critical to the schistosomes survival. The tegument apical membrane undergoes a continuous process of rupture and repair owing to membranous vacuoles fusing with the plasma membrane. Vesicle-associated membrane protein 2 (VAMP2), a member of soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNAREs) is required for membrane fusion. Here, we used RNA interference (RNAi) to knock down the expression of VAMP2 of Schistosoma japonicum (SjVAMP2), and both real-time PCR and western blot analysis confirmed the suppression of this molecule, as well as the suppression of the transcript levels of schistosome glucose transporters (SGTP1 and SGTP4), and insulin receptors (SjIR1 and SjIR2). SjVAMP2-suppressed worms exhibited a lower viability, and phenotypic alterations were also observed in the tegument. Moreover, the glucose consumption of SjVAMP2-suppressed worms decreased significantly in 4 and 6 days, respectively, as well as a significant reduction in egg production. We also observed a significant reduction in worm burden and hepatic eggs burden in two independent RNAi experiment in vivo, and minor pathological changes in mice treated with SjVAMP2 specific small interfering (si)RNA. These findings reveal that SjVAMP2 may play important roles in the maintenance of tegument, glucose uptake, worm development and egg production in schistosomes.

Anthelmintic effects of forage chicory (Cichorium intybus) against free-living and parasitic stages of Cooperia oncophora

Chicory shows great promise as an anthelmintic forage for grazing ruminants that can reduce reliance on anti-parasitic drugs. Recently, we reported potent anthelmintic effects of chicory-based diets in infected cattle with significant reductions in worm burdens of the abomasal nematode Ostertagia ostertagi, whilst no apparent activity was observed against the small intestinal parasite Cooperia oncophora. To explore this discrepancy, we investigated direct anthelmintic effects of forage chicory against C. oncophora in vitro. Chicory leaves (cultivar ‘Spadona’) were extracted with methanol in a Soxhlet apparatus and the resulting extract was purified by solid-phase extraction to concentrate bioactive phytochemicals such as sesquiterpene lactones. C. oncophora eggs and adult worms from mono-infected donor calves were exposed to decreasing concentrations of the chicory extract. In an egg hatch assay, the chicory extract induced a marked and dose-dependent inhibition of egg hatching, with 95% inhibition at 2500μg extract/mL (EC50=619 [95% CI: 530–722] μg extract/mL). In the adult motility inhibition assays, the chicory extract induced a potent and dose-dependent worm paralysis. At 12h of incubation, worms exposed to chicory showed a total paralysis at ≥500μg extract/mL, while after 48h of incubation a complete inhibition of worm motility was observed at ≥250μg extract/mL (EC50=80 [95% CI: 67–95] μg extract/mL). We have demonstrated that forage chicory can induce potent inhibitory effects on the egg hatching and exert direct anthelmintic activity against parasitic stages of C. oncophora. These results suggest that the previously reported absence of in vivo effects of chicory towards C. oncophora in infected animals may be related with host-mediated factors and/or inhibitory digestive conditions, rather than an inherent inactivity of chicory and its bioactive phytochemicals.

Characterization of Schistosoma japonicum tetraspanning orphan receptor and its role in binding to complement C2 and immunoprotection against murine schistosomiasis

BackgroundSchistosomiasis remains an important global public health problem, as millions of people are at risk of acquiring infection. An ideal method for sustainable control of schistosomiasis would be to develop an efficient vaccine. Schistosomes can survive in the host vascular system by immune evasion, regulating the host complement cascade. Schistosoma japonicum tetraspanning orphan receptor (SjTOR) is a complement regulator, which is a tegument membrane protein. To date there is no experimental evidence to explain the function of SjTOR.ResultsWe cloned the first extracellular domain of the SjTOR (SjTOR-ed1) gene and expressed the gene in Escherichia coli. The expression level of SjTOR in different developmental stages of S. japonicum was assessed by quantitative real-time RT-PCR. Western blotting showed that recombinant SjTOR-ed1 (rSjTOR-ed1) could be recognised by schistosome-infected mouse serum. Immunolocalization indicated that the protein was mainly distributed on the tegument of the parasite. Haemolytic assays and ELISA revealed that rSjTOR-ed1 could inhibit complement hemolysis and bind to complement C2. Purified rSjTOR-ed1 emulsified with ISA206 adjuvant could induce a significant reduction of worm burden from 24.51 to 26.51%, and liver egg numbers from 32.92 to 39.62% in two independent trials in mice.ConclusionsThe results of this study indicated that rSjTOR-ed1 could inhibit complement hemolysis and bind to complement C2, and it is a potential vaccine candidate that protects against S. japonicum infection.

Evaluation of Anthelmintic Activity and Composition of Pumpkin (Cucurbita pepo L.) Seed Extracts-In Vitro and in Vivo Studies.

A significant number of studies report growing resistance in nematodes thriving in both humans and livestock. This study was conducted to evaluate the in vitro and in vivo anthelmintic efficiency of Curcubita pepo (C. pepo) L. hot water extract (HWE), cold water extract (CWE) or ethanol extract (ETE) on two model nematodes: Caenorhabditis elegans (C. elegans) and Heligmosoides bakeri (H. bakeri). Methods: Raman, IR and LC-MS spectroscopy analyses were performed on the studied plant material to deliver qualitative and quantitative data on the composition of the obtained extracts: ETE, HWE and CWE. The in vitro activity evaluation showed an impact of C. pepo extracts on C. elegans and different developmental stages of H. bakeri. The following in vivo experiments on mice infected with H. bakeri confirmed inhibitory properties of the most active pumpkin extract selected by the in vitro study. All of the extracts were found to contain cucurbitine, aminoacids, fatty acids, and-for the first time-berberine and palmatine were identified. All C. pepo seed extracts exhibited a nematidicidal potential in vitro, affecting the survival of L1 and L2 H. bakeri larvae. The ETE was the strongest and demonstrated a positive effect on H. bakeri eggs hatching and marked inhibitory properties against worm motility, compared to a PBS control. No significant effects of pumpkin seed extracts on C. elegans integrity or motility were found. The EtOH extract in the in vivo studies showed anthelmintic properties against both H. bakeri fecal egg counts and adult worm burdens. The highest egg counts reduction was observed for the 8 g/kg dose (IC50 against H. bakeri = 2.43; 95% Cl = 2.01–2.94). A decrease in faecal egg counts (FEC) was accompanied by a significant reduction in worm burden of the treated mice compared to the control group. Conclusions: Pumpkin seed extracts may be used to control of Gastrointestinal (G.I.) nematode infections. This relatively inexpensive alternative to the currently available chemotherapeutic should be considered as a novel drug candidate in the nearest future.