In vitro and in vivo anthelmintic effect of essential oil obtained from Thymus capitatus flowers against Haemonchus contortus and Heligmosomoides polygyrus.

Sheep haemonchosis is a disease that causes serious losses in livestock production, particularly with the increase of cases of anthelmintic resistance around the world. This justifies the urgent need of alternative solutions. The aim of this study was to determine the chemical profile, in vitro, and, in vivo, anthelmintic properties of Thymus capitatus essential oil. To evaluate the, in vitro, anthelmintic activity of the T. capitatus EO on Haemonchus contortus, two tests were used: egg hatch assay (EHA) and adult worm motility (AWM) assay. The nematicidal effect of this oil was evaluated, in vivo, in mice infected artificially with Heligmosomoides polygyrus using faecal egg count reduction (FECR) and total worm count reduction (TWCR). Chromatographic characterization of T.capitatus composition using gas chromatography coupled to mass spectrometry (GC-MS) demonstrated the presence of carvacrol (81.16%), as the major constituents. The IC50 values obtained was 1.9 mg/mL in the EHT. In the AWM assay; T. capitatus essential oil achieved 70.8% inhibition at 1 mg/mL after 8 h incubation. The in vivo, evaluation on H. polygyrus revealed a significant nematicidal effect 7 days post-treatment by inducing 49.5% FECR and 64.5% TWCR, using the highest dose (1600 mg/kg). The results of present study, demonstrate that T.capitatus EO possess a significant anthelmintic properties. Furthermore, it could be an alternative source of anthelmintic agents against gastrointestinal infections caused by H. contortus.

Anthelmintic Activity and Cytotoxic Effects of Compounds Isolated from the Fruits of Ozoroa insignis Del. (Anacardiaceae).

Ozoroa insignis Del. is an ethnobotanical plant widely used in traditional medicine for various ailments, including schistosomiasis, tapeworm, and hookworm infections. From the so far not investigated fruits of Ozoroa insignis, the anthelmintic principles could be isolated through bioassay-guided isolation using Caenorhabditis elegans and identified by NMR spectroscopic analysis and mass spectrometric studies. Isolated 6-[8(Z)-pentadecenyl] anacardic (1), 6-[10(Z)-heptadecenyl] anacardic acid (2), and 3-[7(Z)-pentadecenyl] phenol (3) were evaluated against the 5 parasitic organisms Schistosoma mansoni (adult and newly transformed schistosomula), Strongyloides ratti, Heligmosomoides polygyrus, Necator americanus, and Ancylostoma ceylanicum, which mainly infect humans and other mammals. Compounds 1-3 showed good activity against Schistosoma mansoni, with compound 1 showing the best activity against newly transformed schistosomula with 50% activity at 1µM. The isolated compounds were also evaluated for their cytotoxic properties against PC-3 (human prostate adenocarcinoma) and HT-29 (human colorectal adenocarcinoma) cell lines, whereby compounds 2 and 3 showed antiproliferative activity in both cancer cell lines, while compound 1 exhibited antiproliferative activity only on PC-3 cells. With an IC50 value of 43.2 µM, compound 3 was found to be the most active of the 3 investigated compounds.

Synthesis, characterization and antiparasitic activity of organometallic derivatives of the anthelmintic drug albendazole.

Helminthiases, a group of neglected tropical diseases, affect more than one billion people mainly in tropical and subtropical regions. Moreover, major intestinal protozoa have a significant impact on global public health. Albendazole (ABZ) is a broad-spectrum anthelmintic recommended by the World Health Organisation (WHO). However, drug resistance is emerging due to its widespread use. In order to tackle this problem, taking into account the spectacular results obtained with ferroquine, an organometallic derivatization of the antimalarial drug chloroquine, we have prepared, in this study, a series of new ferrocenyl and ruthenocenyl derivatives of the organic drug ABZ and assessed their activity against different helminths and protozoans, namely Trichuris muris, Heligmosomoides polygygrus, Schistosoma mansoni, Giardia lamblia, Haemonchus contortus and Toxoplasma gondii. The ferrocene-containing ABZ analogue 2d exhibited over 70% activity against T. muris adults in vitro at 200 µM and no toxicity to mammalian cells (IC50 >100 µM). H. polygyrus adults were not affected by any of the derivatives tested. Against T. gondii, the ferrocene-containing ABZ analogues 1a and 2d showed better in vitro activity than ABZ and low toxicity to the host cells. The activity of the analogous ruthenocenyl compound 2b against S. mansoni and T. gondii in vitro might be attributed to its toxicity towards the host cells rather than a specific antiparasitic activity. These results demonstrate that the derivatives show a species specific in vitro activity and the choice of the organometallic moieties attached to the organic drug is playing a very important role. Two of our organometallic compounds, namely 1b and 2d, were tested in T. muris infected mice. At a 400 mg kg-1 dose, the compounds showed moderate worm burden reductions but low worm expulsion rates. Overall, this work, which is one of the first studies reporting the potential of organometallic compounds on a very broad range of parasitic helminths and protozoan, is a clear confirmation of the potential of organometallic complexes against parasites of medical and veterinary importance.

Hydro-Ethanolic Extract of Mentha pulegium Exhibit Anthelmintic and Antioxidant Proprieties In Vitro and In Vivo.

INTRODUCTION: During recent decades, the emergence of chemoresistance among synthetic anthelmintic drugs has increased the interest in screening novel natural anthelmintic compounds derived from plants. The current study is aimed to determine the chemical profile, anthelmintic and antioxidant properties of Mentha pulegium hydro-ethanolic extract. MATERIALS AND METHODS: Two tests were used to assess the in vitro anthelmintic activity of the hydro-ethanolic extract of M. pulegium against Haemonchus contortus; egg hatch assay (EHA) and adult worm motility (AWM) assay. M. pulegium extracts at the doses of 500, 1000, 2000 and 4000 mg/kg were evaluated in vivo in mice infected with Heligmosomoides polygyrus. The anthelmintic efficacy was monitored using faecal egg count reduction (FECR) and total worm count reduction (TWCR). The antioxidant activity of M. pulegium extract was evaluated by testing the total antioxidant capacity and the DPPH free radical-scavenging ability. RESULTS: Chromatographic characterization of M. pulegium composition using RP-HPLC revealed the presence of phenolic acids such as syringic acid, ferulic acid and the presence of flavonoid compounds, such as isorhamnetin-3-O-glucoside and kaempferol-3-O-rutinoside. We observed 91.58% inhibition in the EHA at 8 mg/mL after 48 h of incubation (IC50=1.82 mg/mL). In the AWM assay, M. pulegium extract achieved 65.2% inhibition at 8 mg/mL after 8 h. The highest dose (4000 mg/kg) showed a significant nematicidal effect 7 days post-treatment by inducing 60.39% FECR and 71.6% TWCR. We also report strong in vivo antioxidant capacity of the extract, as revealed by a significant increase of the enzymatic activities of catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPx) enzymes in mice infected with H. polygyrus. CONCLUSION: Together, the results in this paper suggest that M. pulegium possesses anthelmintic properties and could be a potential source of novel compounds for the control of helminth parasites as well as its associated oxidative damage.

Chamomile Methanolic Extract Mitigates Small Bowel Inflammation and ROS Overload Related to the Intestinal Nematodes Infection in Mice.

BACKGROUND: Chamomile (Matricaria recutita L.) is a plant which has been reported to be effective in treating several parasitic and digestive diseases. The present study was conducted to evaluate the anthelmintic activity of chamomile methanolic extract (CME). METHODS: In vitro, the anthelmintic activities of CME were investigated on the L3 larvae of Heligmosomoides polygyrus in comparison to albendazole. In vivo, Swiss albino mice were infected with infective third (L3) larval stage of H. polygyrus by intragastric administration. Moreover, the effect of CME and albendazole on worm eggs, adult worms, serum cytokine production, and oxidative stress was studied. RESULTS: All used doses of CME showed a potent anthelmintic activity both in vitro and in vivo and the effect being similar to treatment with albendazole. Moreover, H. polygyrus infestation was accompanied by an intestinal oxidative stress status characterized by an increased lipoperoxidation, a depletion of antioxidant enzyme activity, as well as an overload of hydrogen peroxide. We have also recorded an increase of pro-inflammatory mediator (TNF-α, IL-6, and IL-1ß) levels after treatment with CME (14 ± 0.8; 41 ± 2; 58 ± 4 pg/mg protein, respectively, with the concentration 800 mg/kg, body weight) when compared with infected control mice (20 ± 1; 59 ± 2, and 83 ± 4 pg/mg protein, respectively). However, extract treatment alleviated all the deleterious effects associated with H. polygyrus infection. CONCLUSION: These findings suggest that CME can be used in the control of gastrointestinal helminthiasis and associated oxidative stress.

Exposure of Heligmosomoides polygyrus and Trichuris muris to albendazole, albendazole sulfoxide, mebendazole and oxantel pamoate in vitro and in vivo to elucidate the pathway of drug entry into these gastrointestinal nematodes.

Millions of people are treated with anthelmintics to control soil-transmitted helminth infections; yet, drug distribution in the plasma and gastrointestinal tract compartments and the pathway of drug uptake into gastrointestinal nematodes responsible for the pharmacological effect are unknown. We assessed the distribution and uptake of albendazole, albendazole sulfoxide, albendazole sulfone in the hookworm Heligmosomoides polygyrus in vitro and in vivo as well as the distribution and uptake of albendazole, mebendazole, and oxantel pamoate in the whipworm Trichuris muris in vitro and in vivo. Oral and intraperitoneal treatments (100 mg/kg) were studied. Drug quantities in helminths and host compartments (stomach, the contents and mucosa of the small and large intestine, and the plasma) were determined using HPLC-UV/vis and anthelmintic activities were recorded using phenotypic readout. The influence of 1-aminobenzotriazole (ABT), an irreversible and unspecific cytochrome P450 inhibitor, on albendazole disposition in mice harboring H. polygyrus was evaluated. In vivo, albendazole was found in quantities up to 10 nmol per ten H. polygyrus and up to 31 nmol per ten T. muris. ABT did not change the levels of albendazole or its metabolites in the plasma of mice harboring H. polygyrus or in H. polygyrus, whereas drug levels in the gastrointestinal tract of host mice doubled. Mebendazole and oxantel pamoate quantities per ten T. muris were as high as 21 nmol and 34 nmol, respectively. Albendazole revealed a very dynamic distribution and high rate of metabolism, hence, H. polygyrus and T. muris are exposed to albendazole and both metabolites via multiple pathways. Diffusion through the cuticle seems to be the crucial pathway of oxantel pamoate uptake into T. muris, and likely also for mebendazole. No relationship between concentrations measured in helminths and concentrations in plasma, intestinal content and mucosa of mice, or drug efficacy was noted for any of the drugs studied.

Low-level regulatory T-cell activity is essential for functional type-2 effector immunity to expel gastrointestinal helminths.

Helminth infection is frequently associated with the expansion of regulatory T cells (Tregs) and suppression of immune responses to bystander antigens. We show that infection of mice with the chronic gastrointestinal helminth Heligmosomoides polygyrus drives rapid polyclonal expansion of Foxp3(+)Helios(+)CD4(+) thymic (t)Tregs in the lamina propria and mesenteric lymph nodes while Foxp3(+)Helios(-)CD4(+) peripheral (p)Treg expand more slowly. Notably, in partially resistant BALB/c mice parasite survival positively correlates with Foxp3(+)Helios(+)CD4(+) tTreg numbers. Boosting of Foxp3(+)Helios(+)CD4(+) tTreg populations by administration of recombinant interleukin-2 (rIL-2):anti-IL-2 (IL-2C) complex increased worm persistence by diminishing type-2 responsiveness in vivo, including suppression of alternatively activated macrophage and granulomatous responses at the sites of infection. IL-2C also increased innate lymphoid cell (ILC) numbers, indicating that Treg functions dominate over ILC effects in this setting. Surprisingly, complete removal of Tregs in transgenic Foxp3-DTR mice also resulted in increased worm burdens, with "immunological chaos" evident in high levels of the pro-inflammatory cytokines IL-6 and interferon-γ. In contrast, worm clearance could be induced by anti-CD25 antibody-mediated partial depletion of early Treg, alongside increased T helper type 2 responses and without incurring pathology. These findings highlight the overarching importance of the early Treg response to infection and the non-linear association between inflammation and the prevailing Treg frequency.

Anthelmintic activity of Securidaca longepedunculata (Family: Polygalaceae) root extract in mice, in vitro and in vivo.

OBJECTIVE: To elucidate the pharmacological bases of oral administration of Securidaca longepedunculata (S. longepedunculata) root extract as an anthelmintic in folkloric medicine. METHODS: Albino mice were infected with infective third (L3) larval stage of Heligmosomoides polygyrus (H. polygyrus) by esophageal intubation. Following establishment of the adult worms in the intestine, the mice were treated with 0-2 000 mg/kg body weight (bw) of methanolic root extract of S. longepedunculata and 100 mg/kg bw of pyrantel embonate, the reference drug in vivo. Bioactivity and larvicidal effects of the extract were tested by exposing brine shrimps (Artemia salina) to 0.00-1.00 mg/mL and the L3 stage of Heligmosomoidescontortus (H. contortus) and H. polygyrus to 0.00-2.50 mg/mL of the extract in vitro. RESULTS: The percentage yield of the extract was 7.13% w/w dry matter. The brine shrimps toxicity bioassay resulted in an LC50 of 74.18 µg/mL. The extract had a significant, dose-dependent larvicidal effect on the L3 stage of H. contortus and H. polygyrus with the terminal effect of 75% and 70% at the highest exposure concentrations, respectively. The extract however, did not affect the number of worm eggs per gram (epg) of fecal materials (P<0.05) and total worm burden (twb) of adult H. polygyrus in infected mice. Treatment with pyrantel embonate significant reduced both the fecal egg count and twb to 0 compared to the untreated control (P<0.05). CONCLUSIONS: These results indicate that S. longepedunculata root extract contains potent bioactive compounds and has larvicidal effect on L3 stage of H. contortus and H. polygyrus, substantiating its use as anthelmintic in alternative medicine.

Changes in Heligmosomoides polygyrus glycoprotein pattern by saponins impact the BALB/c mice immune response.

Saponins of marigold (Calendula officinalis), in particular derivatives of 3-O-monoglucuronide of oleanolic acid, are able to reduce infectivity of Heligmosomoides polygyrus in mice. The purpose of this study was to understand the immune activation provoked by third-stage larvae exposed to marigold glucuronides. We also examined the pattern of glycosylation of larval antigens which appeared to be crucial for induction of cytokine production in BALB/c mice; higher concentrations of IL-6, IFN-γ, IL-10 and TNF-α were observed in serum or intestine one week post infection. Three weeks later, in the chronic phase of infection, cells in culture were able to produce IL-6, IFN-γ, TNF-α and IL-17. Restimulation of cells with H. polygyrus antigen resulted in reduced production of IL-6, and TNF-α. The pattern of cytokine production co-existed with reduced expression of terminal glucose, α-linked mannose, N-acetyl-galactosamine, ß-galactose, N-acetyl-glucosamine and α-fucose in several protein bands. Galactose, as a new terminal carbohydrate residue appeared in 20-24kDa protein bands. The number of immunogenic epitopes in parasitic antigens was reduced; only three protein bands of 56, 26 and 12kDa were recognized by IgG1. These studies provide a model system to find the glycosylated molecules expressed on nematodes that improve establishment and survival and characterize cytokine production in mice infected with larvae exposed to saponin. Identification of these molecules is the first step in the recognition of key antigenic epitopes able to induce protective or tolerogenic immune responses.

Efficacy of ivermectin versus dual infections of Haemonchus contortus and Heligmosomoides polygyrus in the mouse.

This report describes a novel assay for the detection of gastrointestinal anthelmintics using mice infected with Haemonchus contortus and adapted to the 1 animal/test group protocol. Mice infected with both H. contortus and Heligmosomoides polygyrus were fed ivermectin-medicated diets for 6 days. A dietary level of 0.09375 ppm was 98.1% effective against the 0- to 6-day-old abomasal stomach worm of sheep, whereas a level of 0.75 ppm reduced the 3- to 9-day-old H. polygyrus worm burden by 94.0%. H. contortus was approximately 8-fold more sensitive to ivermectin than was H. polygyrus in this model. The sensitivity of this assay rivals that of the gerbil-Trichostrongylus colubriformis model while utilizing a more economical host.

Anthelmintic constituents of Clonostachys candelabrum.

Five diastereomeric polyketide glycosides, roselipins 3A-3E (1-5), have been isolated from the acetone extract of Clonostachys candelabrum on the basis of their positive anthelmintic activity. The structures of these compounds were elucidated by comparison of their NMR and MS data to those of previously reported roselipins and related structures, and were confirmed by 2D-NMR spectral analysis. Known compounds linoleic acid (6) and aurantiogliocladin (7) were also isolated as active anthelmintic components, although much less potent than the roselipins.

In vitro nematicidal activity of extracts of Canthium mannii (Rubiaceae), on different life-cycle stages of Heligmosomoides polygyrus (Nematoda, Heligmosomatidae).

The increasing prevalence of anthelmintic-resistant strains of helminths, drug residues in animal products and high cost of conventional anthelmintics has created an interest in studying medicinal plants as an alternative source of anthelmintic. The potential nematicidal activities of four extracts from the bark of Canthium mannii (Rubiaceae) stem were investigated in vitro. Extracts were diluted in distilled water (DW) to obtain five different concentrations (1.5, 2.0, 2.5, 3.0 and 3.5 mg/ml) and put in contact with eggs and larvae of Heligmosomoides polygyrus. The different stages of the life cycle were also put in contact with the same concentration of mebendazole (MBZ, positive control). One millilitre of each extract at different concentrations and control were added to 1 ml solution containing 30-40 eggs or 10-15 larvae (L1, L2 and L3) and distributed in different Petri dishes. The eggs and larvae were incubated at 24 degrees C and exposure times were: 48 h for un-embryonated eggs, 6 h for embryonated eggs; 2, 4, 6 and 24 h for L1 and L2 larvae, 24-48 h for infective larvae (L3), and 5 days for the larval development test (from L1 to L3). DW and 1% dimethyl sulphoxide (DMSO) were used as placebo and DMSO control, respectively. Significant effects were obtained with three of the four extracts, and differences were observed depending on the parasite stage. Cold water extract (CWE), hot water extract (HWE) and ethanol extract (ETE) inhibited embryonic development (40, 45 and 10%) and hatching of embryonated eggs (40, 85 and 80%), respectively, at 3.5 mg/ml. Only ETE killed L1 (97.18%) and L2 (92.68%) larvae of H. polygyrus after 24 h at 3.5 mg/ml and drastically reduced the production rate (6% at 3.0 and 3.5 mg/ml) of infective larvae (L3) after 5 days of incubation compared to other extracts (P < 0.05). However, the infective larvae of H. polygyrus were resistant to the effect of each of the tested products (extracts and mebendazole). These in vitro results suggested that extracts of C. mannii, used by traditional healers in Dschang, Western Region of Cameroon (Central Africa) to cure intestinal helminthiasis and abdominal pains of their patients, possess nematicidal properties. The active principles responsible for the activity could be secondary metabolites such as alkaloids and saponins present in the extracts. It is suggested that further experiments incorporating in vivo purification of extracts and toxicological investigations should be carried out.

Anthelmintic macrolactams from Nonomuraea turkmeniaca MA7381.

A new macrolactam, fluvirucin B0 (1), and two known macrolactams, Sch 38516/fluvirucin B1 (2) and Sch 39185/fluvirucin B3 (3), have been isolated from an acetone extract of a strain of Nonomuraea turkmeniaca. These compounds were isolated by bioassay-guided fractionation as part of our search for new anthelmintics. The structures of these compounds were elucidated by comparison of their NMR and MS data to those of previously reported fluvirucins, and confirmed by 2D-NMR. 1approximately 3 exhibited in vitro activity (EC90 <1.0 approximately 1.7 microg/ml) against Haemonchus contortus larvae, but were ineffective in reducing worm counts in vivo against Heligmosomoides polygyrus in mice at 50 mg/kg dosed intramuscularly.

The anthelmintic efficacy of plant-derived cysteine proteinases against the rodent gastrointestinal nematode, Heligmosomoides polygyrus, in vivo.

Gastrointestinal (GI) nematodes are important disease-causing organisms, controlled primarily through treatment with synthetic drugs, but the efficacy of these drugs has declined due to widespread resistance, and hence new drugs, with different modes of action, are required. Some medicinal plants, used traditionally for the treatment of worm infections, contain cysteine proteinases known to damage worms irreversibly in vitro. Here we (i) confirm that papaya latex has marked efficacy in vivo against the rodent gastrointestinal nematode, Heligmosomoides polygyrus, (ii) demonstrate the dose-dependent nature of the activity (>90% reduction in egg output and 80% reduction in worm burden at the highest active enzyme concentration of 133 nmol), (iii) establish unequivocally that it is the cysteine proteinases that are the active principles in vivo (complete inhibition of enzyme activity when pre-incubated with the cysteine proteinase-specific inhibitor, E-64) and (iv) show that activity is confined to worms that are in the intestinal lumen. The mechanism of action was distinct from all current synthetic anthelmintics, and was the same as that in vitro, with the enzymes attacking and digesting the protective cuticle. Treatment had no detectable side-effects on immune cell numbers in the mucosa (there was no difference in the numbers of mast cells and goblet cells between the treated groups) and mucosal architecture (length of intestinal villi). Only the infected and untreated mice had much shorter villi than the other 3 groups, which was a consequence of infection and not treatment. Plant-derived cysteine proteinases are therefore prime candidates for development as novel drugs for the treatment of GI nematode infections.

Synthesis of anthelmintically active N-methylated amidoxime analogues of the cyclic octadepsipeptide PF1022A.

The N-methylated amidoxime analogues of the cyclic octadepsipeptide PF1022A represent novel derivatives with activity against Trichinella spiralis Owen and Nippostrongylus brasiliensis Lane in vitro and against parasitic nematodes in mice and sheep. Some of them show better activity against Hymenolepis nana Siebold, Heterakis spumosa Schneider and Heligmosomoides polygyrus Dujardin in mice than the natural product PF1022A. In particular an improved efficacy against Haemonchus contortus Rudolphi and Trichostrongylus colubriformis Giles in sheep compared to the potent cyclic octadepsipeptide PF1022A and its mono-thionated derivative has been observed. Here we report on a specific modification at the N-methyl amide linkage by using the mono-thionated PF1022A, resulting in novel anthelmintically active backbone analogues of PF 1022A.

Effect of oxygen radicals and differential expression of catalase and superoxide dismutase in adult Heligmosomoides polygyrus during primary infections in mice with differing response phenotypes.

The ability of oxygen radicals to kill Heligmosomoides polygyrus adult worms was examined by assessing parasite survival following incubation with hydrogen peroxide and acetaldehyde/xanthine oxidase, generators of H2O2 and H2O2/O2(-), respectively. H. polygyrus worms could tolerate levels of < 0.25 mM hydrogen peroxide and < 0.5 mM/20 mU acetaldehyde/xanthine oxidase for 20 h, but, at higher concentrations, marked sex-dependent susceptibility was observed, with males being more sensitive to H2O2 and O2(-) than female worms. The ability to evade free radical-mediated damage was also evaluated by measuring superoxide dismutase (SOD) and catalase levels in worms isolated at different time points from four strains of mice with differing resistance phenotypes. Levels of both catalase and SOD in female worms isolated from 'rapid'[(SWRxSJL)F1], 'fast' (SWR) or 'intermediate' (BALB/c), but not 'slow' (C57BL/10), responder mice showed a strain-dependent increase with time. Moreover, male worms were rejected faster than female worms in the 'rapid', 'fast' and 'intermediate' responder strains of mice. The results suggest that host-derived free radicals can damage adult worms and that female worms can increase production of their scavenging enzymes in response to the immune onslaught that eventually leads to worm expulsion in mice with 'fast', 'rapid' or 'intermediate' response phenotypes.

Activity of the cyclic depsipeptide emodepside (BAY 44-4400) against larval and adult stages of nematodes in rodents and the influence on worm survival.

The present investigations deal with the activity of the cyclic depsipeptide emodepside (BAY 44-4400) against larval and adult stages of three rodent nematodes. While emodepside acts strongly against the adult stages of the rat nematodes Nippostrongylus brasiliensis and Strongyloides ratti, as well as against the mouse nematode Heligmosomoides polygyrus, its actions against the larval stages of these nematodes vary according to the species. Thus, emodepside is highly effective against the lung and intestine larval stages of N. brasiliensis and S. ratti. By contrast. the larval stages of H. polygyrus in the intestine are only partly affected by higher emodepside dosages.

Zinc deficiency impairs immune responses against parasitic nematode infections at intestinal and systemic sites.

Research on the complex interactions among host nutritional status, parasitic infection and immune responsiveness has focused on the detrimental consequences of parasitic infections on host nutritional status and on mechanisms by which malnutrition impairs immunocompetence. Curiously, relatively few studies have examined the effects of malnutrition on the immune response in the parasite-infected host, and even fewer have considered the events occurring at the intestinal level, where absorption of nutrients occurs, intestinal parasites reside, and the gastrointestinal-associated lymphoid tissues play a role in directing both the local and the more systemic immune responses. Our work using a zinc-deficient nematode-infected mouse model reveals that parasites are better able to survive in the zinc-deficient hosts than in well-nourished hosts; that the production of interleukin-4 in the spleen of zinc-deficient mice is depressed, leading to depressed levels of IgE, IgG(1) and eosinophils; and that the function of T cells and antigen-presenting cells is impaired by zinc deficiency as well as by energy restriction. Given the paramount role of the gastrointestinal-associated lymphoid tissues in inducing and regulating immune responses to intestinal parasites and in orchestrating responses in the spleen and peripheral circulation, we conclude that zinc deficiency (in association with energy restriction) exerts profound effects on the gut mucosal immune system, leading to changes in systemically disseminated immune responses and, importantly, to prolonged parasite survival.

The in vitro secretion of acetylcholinesterase by adult stages of Heligmosomoides polygyrus: the effects of broad-spectrum anthelmintics.

The secretion of acetylcholinesterase (AChE) by female and male Heligmosomoides polygyrus was studied in different in vitro culture media. AChE secretion was increased in the presence of fetal calf serum or bovine serum albumin (BSA). In the absence of crowding effects, specific AChE activity in excretion/secretion products was higher for male (2.41 +/- 0.07 mumol min-1 l-1 mg-1) than for female (0.56 +/- 0.04 mumol min-1 mg-1) worms but on a per nematode basis both sexes showed comparable rates of secretion. Acetylthiocholine iodide was the favoured substrate of the enzyme. When the nematodes were incubated in vitro with albendazole (ABZ), ricobendazole (RBZ), mebendazole (MBZ), levamisole (LVM), morantel (MRT) or ivermectin (IVM), at concentrations from 1 mM to 10 nM, in RPMI medium for 2 or 6 h and then transferred to a drug-free medium (RPMI medium supplemented with 0.5% BSA) for 24 h or continuously exposed to the drugs in supplement-free medium (24 h), the concentration- and time-dependent inhibitory effects on AChE secretion were observed. The continued exposure to the drugs for all incubation periods (with a single exception for LVM 1 mM) produced the highest levels of inhibition. Under these conditions, the concentrations inhibiting the secretion of AChE by 50% (IC50) relative to drug-free controls were estimated. The IC50 values ranged from 0.012 microM (IVM) to 2.96 microM (MRT). The potential of this bioassay for the selective primary evaluation of new compounds with broad-spectrum anti-nematodal activity is discussed.

Synergistic action of a cyclic depsipeptide and piperazine on nematodes.

The present study describes the synergistic effects of the cyclic depsipeptide BAY 44-4400 and piperazine in the treatment against the nematodes Trichinella spiralis, Heligmosomoides polygyrus, and Heterakis spumosa. The in vitro anthelmintic activity of a combination of the two compounds (1.7 motility units) against T. spiralis larvae was significantly higher than the sum of the individual drug effects (1.3 motility units). With regard to the rate of expulsion of H. polygyrus worms from the intestine of infected mice, an additive effect was observed; piperazine alone exerted an efficacy of 54.4% and BAY 44-4400 alone, one of 44.4%, whereas the combination of these compounds had an efficacy of 97.5%. With regard to the expulsion of H. spumosa worms, the effect of the combination was more than 5 orders of magnitude greater than the sum of the effects of the single compounds, i.e., there was a considerable potentiation of the actions of BAY 44-4400 and piperazine. Moreover, the combination exerted a significantly higher degree of degenerative effects on the intestine and on the nerve chords of H. spumosa as compared with the single compounds.