Molecular cloning and purification of Ac-TMP, a developmentally regulated putative tissue inhibitor of metalloprotease released in relative abundance by adult Ancylostoma hookworms.

A cDNA encoding a putative tissue inhibitor of metalloprotease was cloned from an Ancylostoma caninum adult hookworm cDNA library by immunoscreening with anti-hookworm secretory products antiserum. Ac-TMP (A. caninum tissue inhibitor of metalloproteinase) is encoded by a 480-bp mRNA with a predicted open reading frame of 140 amino acids (molecular weight, 16,100 Da) that contains one potential N-linked glycosylation site and an N-terminal Cys-X-Cys consensus sequence. The open reading frame corresponds to a putative hookworm tissue inhibitor of metalloproteases (TIMP) with 33% identity and 50% similarity to the N-terminal domain of human TIMP-2. Analysis by reverse transcriptase-polymerase chain reaction indicates that transcription of Ac-tmp is restricted to the adult stage. The protein was isolated from A. caninum adult secretory products by reverse-phase high-performance liquid chromatography and identified as one of the most abundant proteins released by the parasite. To our knowledge, this is the first description of a TIMP from a parasitic invertebrate.

Randomized, double-blind, placebo-controlled trial of oral artemether for the prevention of patent Schistosoma haematobium infections.

Artemether is an efficacious antimalarial drug that also displays antischistosomal properties. Laboratory studies have found that artemether curtails the development of adult worms of Schistosoma japonicum, S. mansoni and S. haematobium, and thus prevents morbidity. These findings have been confirmed in clinical trials for the former two parasites; administered orally once every 2-3 weeks, artemether significantly reduced the incidence and intensity of patent infections. Here, we present the first randomized, double-blind, placebo-controlled trial of artemether against S. haematobium, done in a highly endemic area of Côte d'Ivoire. Urine specimens from 440 schoolchildren were examined over 4 consecutive days, followed by two systematic praziquantel treatments 4 weeks apart. S. haematobium-negative children were randomized to receive 6 mg/kg artemether (N = 161) or placebo (N = 161). Medication was administered orally for a total of six doses once every 4 weeks. Adverse events were assessed 72 hours after medication, and perceived illness episodes were monitored throughout the study period. Incidence and intensity of S. haematobium infections, and microhematuria and macrohematuria were assessed 3 weeks after the final dosing. We also monitored malaria parasitemia and treated positive cases with sulfadoxine-pyrimethamine (SP). Oral artemether was well tolerated. The incidence of patent S. haematobium infections in artemether recipients was significantly lower than in placebo recipients (49% versus 65%, protective efficacy: 0.25, 95% CI: 0.08-0.38, P = 0.007). The geometric mean infection intensity in the artemether group was less than half that of the placebo recipients (3.4 versus 7.4 eggs/10 mL urine, P < 0.001). Heavy S. haematobium infections, microhematuria and macrohematuria, and the incidence of malaria parasitemia were all significantly lower in artemether recipients. In conclusion, previous findings of efficacy of artemether against S. japonicum and S. mansoni were confirmed for S. haematobium, although the protective efficacy was considerably lower. These findings enlarge the scope and potential of artemether and further contribute to discussions of its role as an additional tool for integrated schistosomiasis control.

Albendazole-soybean oil emulsion for the treatment of human cystic echinococcosis: evaluation of bioavailability and bioequivalence.

A single 12.5 mg/kg dose of albendazole (Abz) in tablet form (AbzT) followed 2 weeks later by an equivalent dose of Abz emulsified in 30% soybean oil (AbzE) was administered orally 2 h after the first morning meal to 7 male adult patients with cystic echinococcosis caused by Echinococcus granulosus. Serum samples were taken 1, 3, 5, 7, 8, 9, 11, 14, 18, 24, 36, and 48 h post medication from each patient to measure the serum concentrations of albendazole sulfoxide (AbzSOX), the principal bioactive metabolite of Abz. AbzSOX concentrations were measured by reverse phase HPLC. The data were subjected to pharmacokinetic analysis to compare the relative bioavailability and bioequivalence of AbzT and AbzE. The results demonstrated that the mean peak concentrations (C(max)) for AbzT and AbzE were 1.06+/-0.38 mg/l and 1.71+/-0.47 mg/l, respectively; the area under the concentration-time curves (AUC) were 13.24+/-4.93 mg x h/l and 21.01+/-7.54 mg x h/l, respectively. The relative bioavailability of AbzE was F(Flu)=1.59. Two one-sided tests procedure and (1-2 alpha) 90% confidence interval methods were used to evaluate the bioequivalence of AbzE and AbzT. The results demonstrated that the bioavailability of AbzE was greater than AbzT.

Augmented bioavailability and cysticidal activity of albendazole reformulated in soybean emulsion in mice infected with Echinococcus granulosus or Echinococcus multilocularis.

The anthelminthic drug, albendazole (Abz), was reformulated in a soybean oil emulsion and evaluated as a therapeutic agent for the treatment of Echinococcus granulosus and Echinococcus multilocularis in mice. Abz emulsified with 30% soybean oil (AbzE-30) resulted in higher circulating plasma concentrations of the major bioactive Abz metabolite, Abz sulfoxide (AbzSOX), after oral administration, compared with an Abz suspension. The soybean oil-emulsified Abz compound was also noted to penetrate into the hydatid cyst wall and produced higher hydatid cyst concentrations of AbzSOX. The emulsion was superior to Abz suspension in reducing the size of hydatid cysts caused by E. granulosus protoscolices collected from naturally infected sheep in Urumchi, Xinjiang Uygar Autonomous Region. In contrast, the reformulated compound's ability to reduce E. multilocularis cyst masses was only marginally superior to Abz suspension. AbzE-30 exhibited increased bioavailability and bioactivity in the treatment of murine Echinococcus hydatid cyst infections. The compound has the potential for improving therapeutic outcomes for human echinococcosis.

Observations on clinical efficacy of albendazole emulsion in 264 cases of hepatic cystic echinococcosis.

Two regimens of albendazole emulsion (AbzE), a novel formulation, were used in the treatment of 264 cases of hepatic cystic echinococcosis. AbzE 10 mg/kg per day (calculated by albendazole base) was administered orally to 71 cases for 6 months to over 1 year. Imaging evaluation at the end of courses showed overall efficacy in 97.2%, (cure rate 60.6%, and inefficacy rate 2.8%); The follow-up study on 62 cases 3-4 years post therapeutic courses showed overall efficacy in 92.0% (cure rate 83.9%, ineffective rate 1.5% and recurrence rate 6.5%); Abz 12.5 mg/kg per day was administered orally to 193 cases for 3 months to over 1 year, resulting in an overall efficacy of 97.9%, (cure rate 75.1% and inefficacy rate 2.1%). The follow-up study in 139 cases 2-4 years post treatment demonstrated efficacy in 89.2%, (cure rate 84.2% and recurrence rate 10.8%); Mild reversible adverse reactions were observed in 14.4% of the patients. Retreatment of recurrent hydatidosis patients with AbzE provided promising results. AbzE is considered to be superior to the albendazole tablet or capsule formulations currently used in treatment of liver cystic hydatid disease.

Effect of Artemether on the Tegument of Adult Schistosoma haematobium Recovered from Mice / 中国寄生虫学与寄生虫病杂志

Objective To assess the effect of artemether on the tegument of adult Schistosoma haematobium harbored in mice. Methods Ten mice were infected subcutaneously with 100-120 S. haematobium cercariae each. At day 81 post-infection, 8 mice were treated orally with 400 mg/kg artemether. Mice were sacrificed at 1, 3, 7 and 14 days post-treatment, and schistosomes were collected by the perfusion technique, fixed and examined under a scanning electron microscope. Schistosomes obtained from the 2 untreated mice served as a control. Results Twenty-four hours post-treatment, tubercles on the tegument of male worms showed lesions, characterized by enlargement, collapse and partial peeling off from the border with the tegument. In both male and female worms, the tegument showed focal or extensive swelling, fusion, vacuolization, erosion, peeling, and destruction of sensory structures. Three days post-treatment,tegumental alterations further aggravated; particularly severe damage was the swelling or collapse of the oral sucker observed in both sexes. In addition, extensive swelling, erosion and peeling of tegumental ridges and destruction of discoidlike sensory structures were seen in female worms. Seven to 14 days post-treatment, moderate-to-severe damage was still evident in some worms, whereas other worms surviving the treatment showed apparent recovery in most parts of their tegument. Conclusion Artemether causes extensive and severe tegumental damage in adult S. haematobium.

Ultrastructural Alterations of Adult Schistosoma haematobium Harbored in Mice Following Artemether Administration / 中国寄生虫学与寄生虫病杂志

Objective To perform a temporal examination of ultrastructural alterations in adult Schistosoma haematobium due to artemether Methods Eight mice infected with 100-120 S. haematobium cercariae for 81 days were treated intragastrically with 400 mg/kg artemether. At 24 hours, 3, 7 and 14 days post-treatment, groups of 2 mice were sacrificed and schistosomes collected by the perfusion technique. Worm samples were fixed and examined by transmission electron microscopy. Schistosomes were also obtained from 2 untreated mice that served as control.Results Typical ultrastructural alterations included swelling, lysis and vacuolization of the tegumental matrix, and disappearance of basal membrane. In sensory organelles and tubercles, there was extensive or local lysis of internal structure. In the musculature, parenchymal tissues, syncytium and gut epithelial cells, focal or extensive lysis, decrease in granular endoplasmic reticulum, vacuolization and degeneration of mitochondria were observed. These alterations became apparent both in male and female worms 24 hours post-treatment. In female worms, severe damage to the vitelline cells was also observed, resulting in the emergence of vacuoles, a decrease in granular endoplasmic reticulum,fusion of vitelline balls or even collapse of damaged vitelline cells. The most extensive tegumental alterations were observed 3-7 days post-treatment. Whilst 14 days post-treatment ultrastructural damage was still apparent, the tegument of some worms showed similar features to those recovered from untreated control mice. Conclusion Administration of artemether to mice infected with adult S. haematobium results in extensive damage to the ultrastructure in the tegument and subtegument tissues of the worms, confirming previous results with other schistosome species.

Progress in Anthelmintic Agent Study since the Founding of the People′s Republic of China and Current Challenges / 中国寄生虫学与寄生虫病杂志

This paper summarizes the progress in the study on anthelmintics,including nematocide,trematocide and cestocide since the founding of the People′ s Republic of China and the roles that these agents played in the control of parasitic diseases.Meanwhile,views are given to the challenges faced in the further study on anthelmintics.

Impact of Host Factors on the Schistosome-Killing Process Induced by Praziquantel / 中国寄生虫学与寄生虫病杂志

Experimental studies indicated that the killing process of schistosomes induced by praziquantel comprises two aspects, i.e. the direct effect of praziquantel on schistosomes and the host immune reaction. The former one appears in stimulation of worm activity, spasmodic contraction of worm musculatures and severe damage to the tegument, which results in hepatic shift of schistosomes, influence on the nutrition absorption, excretion/secretion and defense function of the tegument, followed by the secondary interference with the worm metabolism. While the latter one involves the destruction of the host concomitant immune mechanism after tegumental damage and peeling, which is unfavorable for worm survival. Particularly, the exposure of the worm surface antigen provides a target which can be attacked by specific antibodies. Therefore, the antischistosomal activity of praziquantel is immune-dependent. In this paper some host factors involved in the killing process of schistosomes induced by praziquantel were summarized.

Study Progress on the Mode of Action of Praziquantel Against Schistosomes / 中国寄生虫学与寄生虫病杂志

Praziquantel is the only effective drug of choice against five huaman species of schistosomes. Main adva-ntages of praziquantel include convenient oral administration, high safety and efficacy as well as short treatment course. To better understand the mode of action of praziqantel against schistosomes would be helpful for further development of new broad-spectrum anthelminthics. This paper summarizes the 30 years′ research progress on the mode of action of the drug against schistosomes proceeded by domestic and abroad laboratories.

Experimental Study Progress on Tribendimidine,Artemether and Artesuante Against Clonorchis sinensis and other Trematodes / 中国寄生虫学与寄生虫病杂志

Currently praziquantel is one of the major drugs used in treatment of schistosomiasis and other trematode infections.Recent experimental studies indicate that a new anthelmintic,tribendimidine,is used in the treatment of intestinal nematodes,also possesses effect against several species of trematodes including Clonorchis sinensis,Opisthorchis viverrini and Echinostoma caproni.Tribendimidine is even more effective against C.sinensis in rats that a single 300 mg/kg oral dose cures almost all of the animals treated,a lower cure dose than praziquantel(375-500 mg/kg).The anti-malarial drugs artemether and artesunate are not only effective in the prevention of schistosomiasis,but also effective against several species of trematodes,especially C.sinensis.The single oral dose of both drugs to cure or achieve high efficacy in infected rats is 75 mg/kg.This review summarized research progress on tribendimidine,artesunate,and artemether in experimental animals infected with C.sinensis and other species of trematodes.

Reduction of Total Antioxidant Capacity in Artemether-treated Female Schistosoma japonicum / 中国寄生虫学与寄生虫病杂志

Objective To study the effect of artemether (Art) on total antioxidant capacity (T-AOC) in adult Schistosoma japonicum. Methods In vitro, the T-AOC was determined in five-week old worms incubated without or with Art and/or hemin for 24 h, and the worms were continuously incubated for 96 h, then worm survival was assessed. In vivo, T-AOC was determined in worms freshly recovered from mice 6 - 24 h after treatment with Art 300 mg/kg. Results Throughout 96 h incubation no worms were killed by 50 μmol/L Art or 50 μmol/L hemin alone, but approximatdy 80% of them were killed by Art plus hemin. Addition of reduced glutathione and vitamin E could significantly block the cidal action of the combined treatment. No effect on T-AOC was seen in the worms exposed to Art or heroin alone for 24 h, but the combined treatment led to a pronounced T-AOC reduction in female worms in vitro. Such a drug effect on female worms was demonstrated in vivo. After female worms were exposed to Art for 6 - 24 h in vivo, their T-AOC was significantly reduced by 40% - 64%. However, no drug effect on male worms' T-AOC was observed in vivo and in vitro exposed to Art plus hemin. Conclusion Art-induced T-AOC reduction in female worms may sensitize them to lethal damages of endogenous and exogenous oxygen radicals.

ODNSIRUCTION AND ANALYSIS OF cDNA LIBRARY OF NECATOR AMERICANUS THIRD STAGE LARVAE / 中国寄生虫学与寄生虫病杂志

[Objective] To obtain the genetic information on Necator americanus and to search for the purpose genes.[Methods] Mrna was isolated from the third stage larvae of Necator americanus maintained in hamsters.Double strand Cdna was synthesized and ligated to ΛzapⅡ vector to construct the Cdna library.Expresed se-quence tages (ESTs) were obtained by single pass sequencing of randomly isolated Cdna clones from the es-tablished library.[Results] A Cdna librazy of N.americanus was successfully constructed with high recombi-nant efficiency.The titer of unamplified library was 1×107.The insert size was about 750～3000bp.Of 11 ESTs obtained from the library,7 have a significant homology with certain functional genes.[Conclunsion]A high quality and high representative Cdna library of N.americanus was constructed at the first time and ome functional genes were identified from the library by ESTs.

Therapeutic effect of artemether and artesunate in mice infected with Schistosoma mansoni / 中国血吸虫病防治杂志

Objective To explore the therapeutic effect of artemether and artesunate on adult Schistosoma mansoni in experimental mice.Methods The mice were administered intragastrically with artemether or artesunate 46 days after being infected with cercariae of S.mansoni subcutaneously. On Day 1,a dose of 400, 300, 200 mg/kg of artemether or artesunate was administered to the mice. From Day 2 to Day 7, a half above dose was administered. On Day 7, the single-dose groups were administered with artemether or artesunate at the dose of 1600, 1200, 800 mg/kg, meanwhile, an infected group of mice was served as control, untreated. Results With 7-day therapy of artemether at the dosage of 1600, 1200, 800 mg/kg, the worm reduction rates were 53%, 49% and 53%, respectively, and female worm reduction rates were from 78%-82%, compared with the control group.The therapeutic effects of artemether on single-dose groups were similar. The worm reduction rates, with 7-day therapy of artemether at the dosage of 1600, 1200, 800 mg/kg , were 16%,37% and 49%, respectively, compared with the control group. Conclusion The efficacy of therapy with artemether and artesunate on S.mansoni infection mice were relatively well. Concerning the therapeutic effect and toxicity, artemether is slightly better than artesunate.

Clinical efficacy of albendazole emulsion in treatment of 212 cases of liver cystic hydatidosis / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To evaluate the clinical efficacy of a new formulation of albendazole emulsion (AbzE) in cases of liver cystic hydatidosis.</p><p><b>METHODS</b>Two regimens of AbzE (10 mg.kg(-1).d(-1) and 12.5 mg.kg(-1).d(-1)) were given to 212 patients with liver cystic hydatidosis in courses ranging from 3 months to more than one year. Assessment of drug efficacy was essentially based on imaging signs with ultrasonography as the main tool. Assessments were performed at the end of different courses and in the follow-up study of 1 - 4 years after the cessation of therapy.</p><p><b>RESULTS</b>At the end of therapeutic courses, the overall cure rate of the 212 cases was 74.5%, with a 99.1% effective rate. In the follow-up study, the cure rate was 83.1%, effective rate was 89.3%, ineffective rate was 0.6%, and recurrence rate was 10.2%. The highest cure rate was observed in cases receiving AbzE 12.5 mg.kg(-1).d(-1) for 9 months. Retreatment of recurrent cases with AbzE obtained satisfactory results.</p><p><b>CONCLUSIONS</b>AbzE surpassed other currently used antihydatidosis drugs or formulations with its promising efficacy and mild side effects, and could be recommended as a drug of choice in the treatment of cystic hydatidosis.</p>