Microcapsulated biocides for the targeted control of invasive bivalves.

Invasive alien species (IAS) are one of the greatest drivers of ecological change. Typically, control uses chemical agents that often are ineffective, harmful to non-target organisms, and environmentally persistent. Bivalves are frequently high impact IAS, but have proven particularly hard to control due to their valve-closing response when exposed to conventional control agents. Microencapsulation of biocides with edible coatings represents a highly targeted delivery route, bypassing avoidance responses and accumulating in bivalves through their prodigious filter feeding. Uneaten microcapsules degrade and become biologically inactive within hours thus reducing potential impacts on non-target biota. We manufactured two new formulations of microcapsules (BioBullets). Particles were designed to mimic natural food particles (algae) in terms of size (9.5 ± 0.5 to 19.4 ± 1.3 SE µm diameter), buoyancy (near neutral) and shape (spherical). Laboratory exposures demonstrated that two formulations effectively controlled the Gulf wedge clam Rangia cuneata, an IAS currently spreading rapidly through Europe. A single dose of 2-6 mg L-1 of the active ingredient in a static system achieved 90% mortality after 30 days of exposure. Microencapsulation offers an effective and targeted management tool for rapid responses following the early detection of both Gulf wedge clams and many other filter-feeding IAS, and may be especially effective in closed systems or where populations remain very localised.

Molluscicidal effectiveness of Luo-Wei, a novel plant-derived molluscicide, against Oncomelania hupensis, Biomphalaria alexandrina and Bulinus truncatus.

BACKGROUND: Control of snail intermediate hosts has been proved to be a fast and efficient approach for interrupting the transmission of schistosomiasis. Some plant extracts have shown obvious molluscicidal activity, and a new compound Luo-Wei, also named tea-seed distilled saponin (TDS), was developed based on the saponins extracted from Camellia oleifera seeds. We aimed to test the molluscicidal activity of 4% TDS against the intermediate host snails in China and Egypt, and evaluate its environmental safety to non-target organisms. METHODS: In the laboratory, Oncomelania hupensis, Biomphalaria alexandrina and Bulinus truncatus were exposed to 4% TDS, and the median lethal concentration (LC50) was estimated at 24, 48 and 72 h. In the field, snail mortalities were assessed 1, 2, 3 and 7 d post-immersion with 2.5 g/m3 4% TDS and 1, 3, 7 and 15 d post-spraying with 5 g/m2 4% TDS. In addition, the acute toxicity of 4% TDS to Japanese quail (Coturnix japonica), zebrafish (Brachydanio rerio) and freshwater shrimp (Macrobrachium nipponense) was assessed by estimations of LC50 or median lethal dose (LD50). RESULTS: In the laboratory, the LC50 values of 4% TDS for O. hupensis were 0.701, 0.371 and 0.33 mg/L at 24, 48 and 72 h, respectively, and 4% TDS showed a 1.975 mg/L [corrected] 24 h LC50 against B. alexandrina, and a 1.396 mg/L 24 h LC50 against B. truncatus. Across all study regions, the pooled mortalities of O. hupensis were 72, 86, 94 and 98% at 1, 2, 3 and 7 d, following field immersion of 4% TDS at a dose of 2.5 g/m3, and were 69, 77, 85 and 88% at 1, 3, 7 and 15 d, following field spraying at 5 g/m2, respectively. 4% TDS had moderate toxicity to Japanese quail (7 d LD50 > 60 mg/kg) and to shrimp (96 h LC50 = 6.28 mg/L; 95% CI: 3.53-11.2 mg/L), whereas its toxicity to zebrafish was high (96 h LC50 = 0.15 mg/L; 95% CI: 0.14-0.17 mg/L). CONCLUSIONS: 4% TDS is active against O. hupensis, B. alexandrina and B. truncatus under laboratory and field conditions, and it may be a candidate molluscicide of plant origin.

Double impact: natural molluscicide for schistosomiasis vector control also impedes development of Schistosoma mansoni cercariae into adult parasites.

BACKGROUND: Schistosomiasis has been reported in 78 endemic countries and affects 240 million people worldwide. The digenetic parasite Schistosoma mansoni needs fresh water to compete its life cycle. There, it is susceptible to soluble compounds that can affect directly and/or indirectly the parasite's biology. The cercariae stage is one of the key points in which the parasite is vulnerable to different soluble compounds that can significantly alter the parasite's life cycle. Molluscicides are recommended by the World Health Organization for the control of schistosomiasis transmission and Euphorbia milii latex is effective against snails intermediate hosts. METHODOLOGY/PRINCIPAL FINDINGS: We used parasitological tools and electron microscopy to verify the effects of cercariae exposure to natural molluscicide (Euphorbia milii latex) on morphology, physiology and fitness of adult parasite worms. In order to generate insights into key metabolic pathways that lead to the observed phenotypes we used comparative transcriptomics and proteomics. CONCLUSIONS/SIGNIFICANCE: We describe here that the effect of latex on the adult is not due to direct toxicity but it triggers an early change in developmental trajectory and perturbs cell memory, mobility, energy metabolism and other key pathways. We conclude that latex has not only an effect on the vector but applies also long lasting schistosomastatic action. We believe that these results are of interest not only to parasitologists since it shows that natural compounds, presumably without side effects, can have an impact that occurred unexpectedly on developmental processes. Such collateral damage is in this case positive, since it impacts the true target of the treatment campaign. This type of treatment could also provide a rational for the control of other pests. Our results will contribute to enforce the use of E. milii latex in Brazil and other endemic countries as cheap alternative or complement to mass drug treatment with Praziquantel, the only available drug to cure the patients (without preventing re-infection).

Apropos: critical analysis of molluscicide application in schistosomiasis control programs in Brazil.

Schistosomiasis is a snail-transmitted infectious disease affecting over 200 million people worldwide. Snail control has been recognized as an effective approach to interrupt the transmission of schistosomiasis, since the geographic distribution of this neglected tropical disease is determined by the presence of the intermediate host snails. In a recent Scoping Review published in Infectious Diseases of Poverty, Coelho and Caldeira performed a critical review of using molluscicides in the national schistosomiasis control programs in Brazil. They also described some chemical and plant-derived molluscicides used in China. In addition to the molluscicides described by Coelho and Caldeira, a large number of chemicals, plant extracts and microorganisms have been screened and tested for molluscicidal actions against Oncomelania hupensis, the intermediate host of Schistosoma japonicum in China. Here, we presented the currently commercial molluscicides available in China, including 26% suspension concentrate of metaldehyde and niclosamide (MNSC), 25% suspension concentrate of niclosamide ethanolamine salt (SCNE), 50% niclosamide ethanolamine salt wettable powder (WPN), 4% niclosamide ethanolamine salt dustable powder (NESP), 5% niclosamide ethanolamine salt granule (NESG) and the plant-derived molluscicide "Luowei". These molluscicides have been proved to be active against O. hupensis in both laboratory and endemic fields, playing an important role in the national schistosomiasis control program of China. Currently, China is transferring its successful experiences on schistosomiasis control to African countries. The introduction of Chinese commercial molluscicides to Africa, with adaptation to local conditions, may facilitate the progress towards the elimination of schisosomiasis in Africa.

Altered Gene Expression in the Schistosome-Transmitting Snail Biomphalaria glabrata following Exposure to Niclosamide, the Active Ingredient in the Widely Used Molluscicide Bayluscide.

In view of the call by the World Health Organization (WHO) for elimination of schistosomiasis as a public health problem by 2025, use of molluscicides in snail control to supplement chemotherapy-based control efforts is likely to increase in the coming years. The mechanisms of action of niclosamide, the active ingredient in the most widely used molluscicides, remain largely unknown. A better understanding of its toxicology at the molecular level will both improve our knowledge of snail biology and may offer valuable insights into the development of better chemical control methods for snails. We used a recently developed Biomphalaria glabrata oligonucleotide microarray (31K features) to investigate the effect of sublethal exposure to niclosamide on the transcriptional responses of the snail B. glabrata relative to untreated snails. Most of the genes highly upregulated following exposure of snails to niclosamide are involved in biotransformation of xenobiotics, including genes encoding cytochrome P450s (CYP), glutathione S-transferases (GST), and drug transporters, notably multi-drug resistance protein (efflux transporter) and solute linked carrier (influx transporter). Niclosamide also induced stress responses. Specifically, six heat shock protein (HSP) genes from three super-families (HSP20, HSP40 and HSP70) were upregulated. Genes encoding ADP-ribosylation factor (ARF), cAMP response element-binding protein (CREB) and coatomer, all of which are involved in vesicle trafficking in the Golgi of mammalian cells, were also upregulated. Lastly, a hemoglobin gene was downregulated, suggesting niclosamide may affect oxygen transport. Our results show that snails mount substantial responses to sublethal concentrations of niclosamide, at least some of which appear to be protective. The topic of how niclosamide's lethality at higher concentrations is determined requires further study. Given that niclosamide has also been used as an anthelmintic drug for decades and has been found to have activity against several types of cancer, our findings may be of relevance in understanding how both parasites and neoplastic cells respond to this compound.

Resistance to niclosamide in Oncomelania hupensis, the intermediate host of Schistosoma japonicum: should we be worried?

As the currently only available molluscicide, niclosamide has been widely used for snail control for over 2 decades in China. There is therefore a concern about the emergence of niclosamide-resistant snail populations following repeated, extensive use of the chemical. The purpose of this study was to investigate the likelihood of niclosamide resistance in Oncomelania hupensis in China. Active adult O. hupensis snails derived from 20 counties of 10 schistosomiasis-endemic provinces of China, of 10 snails in each drug concentration, were immersed in solutions of 1, 0.5, 0.25, 0.125, 0.063, 0.032, 0.016 and 0.008 mg L-1 of a 50% wettable powder of niclosamide ethanolamine salt (WPN) for 24 and 48 h at 25 °C, and the median lethal concentration (LC50) was estimated. Then, the 24- and 48-h WPN LC50 values were compared with those determined in the same sampling sites in 2002. The results indicated that the 24- and 48-h WPN LC50 values for O. hupensis were not significantly different from those determined in 2002 (P = 0.202 and 0.796, respectively). It is concluded that the current sensitivity of O. hupensis to niclosamide has not changed after more than 2 decades of repeated, extensive application in the main endemic foci of China, and there is no evidence of resistance to niclosamide detected in O. hupensis.

Schistosomicidal and molluscicidal activities of aminoalkylamino substituted neo- and norneocryptolepine derivatives.

CONTEXT: The cryptolepines originate from the roots of the climbing shrub Cryptolepis sanguinolenta (Lindi) Schitr(Periplocaeae) which is used in Central and West Africa in traditional medicine for the treatment of malaria. OBJECTIVES: Evaluation for the first time of a series of chloro- and aminoalkylamino derivatives of neo- and norneocryptolepines for potential schistosomicidal and molluscicidal activities. MATERIALS AND METHODS: A series of chloro- and aminoalkylamino substituted neo- and norneocryptolepine derivatives were synthesized. They were tested in vitro against viable Schistosoma mansoni Sambon mature worms in culturemedium with fetal serum and antibiotics and in dechlorinated water against the snail vector Biomphalaria alexandrina Ehrenberg. Active compounds were further subjected to determination of their IC50 values. RESULTS: Results showed that six neocryptolepine and two norneocryptolepine derivatives had in vitro schistosomicidal activity on Egyptian and Puerto Rican strains of S. mansoni. The most effective derivative (2-chloro-5-methyl-N-(2-morpholin-4-ethyl)-5H-indolo[2,3b]quinoline-11-amine) has IC50 and IC90 1.26 and 4.05 µM and 3.54 and 6.83 µM with the Egyptian and Puerto Rican strains of Schistosoma, respectively. All eight derivatives showed molluscicidal activity against the vector snail B. alexandrina. The most active compound (2-chloro-11-(4-methylpiperazin-1-yl)-6H-indolo[2,3-b] quinoline) has LC50 0.6 and LC90 3.9 ppm after 24 h. DISCUSSION AND CONCLUSIONS: The findings demonstrate that introducing chloro- and aminoalkylamino side chain initiated both schistosomicidal and molluscicidal activities in these derivatives. The structure­activity relationship of this series of compounds is discussed.

[Evaluation on molluscicidal effect of suspension concentrate of metaldehyde and niclosamide].

OBJECTIVE: To evaluate the molluscicidal effect of suspension concentrate of 26% metaldehyde and niclosamide (MNSC) in laboratory and field. METHODS: The experiment of MNSC against Oncomelania snails were performed by immersion and spray methods with different time in laboratory and field, and the wettable powder of 50% niclosamide ethanolamine salt (WPN) and fresh water were set as the controls. RESULTS: In laboratory, by immersion method (immersion for 24 h), LC50 of MNSC was 0.058 3 mg/L, which was lower than 0.094 7 mg/L of WPN. For the concentrations of 2.0 mg/L and 1.0 mg/L of MNSC by immersion method (immersion for 24, 48 h and 72 h, respectively), the death rates of snails were all 100%. In laboratory, by the spray method, the snail death rates were higher than 97% on the 3rd and 7th day when the dosage of MNSC was higher than 1.0 g/m2. In the field, by the immersion method, the snail death rates of MNSC (2 g/m3) for 24, 48 h and 72 h were 97.78% -100% in Hubei Province and 97.50% - 100% in Anhui Province, respectively, the rates of WPN were 98.89% - 100% and 97.56% - 100%, respectively, and the snail death rates of MNSC and WPN were both 100% in Jiangsu Province. In the field, by the spray method, the snail death rates of MNSC for 1, 3 d and 7 d were 83.12% - 94.64% in Jiangsu Province, 67.81% - 86.67% in Hubei Province and 72.80% - 100% in Anhui Province, respectively. The snail death rates of WPN for 1, 3 d and 7 d were 78.40% - 86.11%, 77.27% - 87.50% and 83.81% - 98.51%, respectively. In Hunan Province, the snail death rates of MNSC and WPN after spraying for 7 d were 94.94% and 94.20%, respectively, after spraying for 15 d, the rates were 86.67% and 89.38%, respectively. CONCLUSIONS: The molluscicidal effect of MNSC is as the same as WPN by immersion and spray methods in laboratory and field. MNSC has the characteristics of good water solubility, quick dispersing and easy using. In addition, the efficiency component of MNSC is half of WPN. So MNSC is a novel form of molluscicide which is worthy of promotion in field.

[Toxicity of niclosamide with plastic film mulching to fish].

OBJECTIVE: To observe the toxicity of niclosamide with plastic film mulching to fish. METHODS: A ditch with Oncomelania snails was selected as the study site, and it was divided into a control group and a molluscicide group. After the river closure, the molluscicidal group was sprayed with niclosamide with a dosage of 2 g/m2, and then covered with plastic film; no molluscicide measure was implemented in the control group. The toxicity of the molluscicide to carp fish in resting and flowing water environment was observed. RESULTS: In flowing water, the death rates of carp fry of the molluscicidal and control groups showed no statistical difference 7 d after the film mulching (P = 0. 680), and no carp fry died in the 2 groups after plastic film taking off for 4 d. The death rates of carp fry between the 3 sites (with a distance of 50, 100 and 150 m from the molluscicidal group, respectively) and the control group had no statistical difference 7 d after film mulching (P = 0.955), and no dead fry was discovered 4 d after taking off the film. In downstream, for the death rates of carp fry, there was no difference between the molluscicidal group and the control group (P = 0.376). The death rates of snails before and after the molluscicidal experiment showed a significant difference (P < 0.01). CONCLUSIONS: The technology of niclosamide molluscicide with plastic film mulching can not only improve the molluscicidal effect, but also avoid the toxicity to fish, which is suitable for special environment such as a fish pond.

Molluscicidal activity of Solanum species of the Northeast of Brazil on Biomphalaria glabrata.

Seven species of Solanum were screened for their molluscicidal properties against Biomphalaria glabrata, the intermediated host of Schistosoma mansoni, Solanum agrarium, S. jabrense, S. melissarum, S. megalonyx, S. paludosum, S. paraibanum and S. stipulaceum. Four extracts showed molluscicidal activity with LC(50) from 22 to 56 microg/ml.

Screening of Echinops ellenbeckii and Echinops longisetus for biological activities and chemical constituents.

Members of the genus Echinops in the family of Asteraceae are widely used in Ethiopian herbal medicine for the treatment of various diseases and illness such as migraine, diarrhea, heart pain, different forms of infections, intestinal worm infestation and hemorrhoid. Hydroalcoholic extracts of the root, flower head, leaf and stem of Echinops ellenbeckii O. Hoffm. and Echinops longisetus A. Rich were investigated for their chemical constituents and biological activities. The presence of alkaloids, saponins, phytosterols, polyphenols and carotenoids in the different parts of the plants was observed whilst anthraquinones were not detected. The leaf extracts of both plants and stem extract of E. longisetus showed strong inhibitory activity against cultures of Staphylococcus aureus. None of the extracts were found to be active against Gram-positive organisms. The flower extract of E. ellenbeckii showed strong inhibitory activity against Candida albicans. Root and flower extracts of the plants investigated showed lethal activity against earthworms. Moreover, the extracts of the roots of both plants showed molluscicidal activity against schistosome-transmitting snail hosts. The biological activities observed were dose dependent.

Biological screening of Nigella damascena for antimicrobial and molluscicidal activities.

The essential oil, various extracts at different polarity, fractions, and pure compounds obtained from Nigella damascena plants and seeds were screened for biological activity. Antimicrobial tests showed the essential oil to be active only against Gram positive bacteria; among the extracts, the BuOH was active against Pseudomonas aeruginosa and Staphylococcus aureus. Molluscicidal activity was absent.

Molluscicidal and antimicrobial activity of Solanum aculeastrum.

The methanolic extract of the fresh root bark and berries of Solanum aculeastrum showed significant activity against host snails of schistosomiasis. The berries extract was more potent with 100% snail kill at 50 ppm. Fractionation increased activity with 100% mortality at 20 ppm. The aqueous and methanolic extracts of the berries showed moderate antimicrobial activity, increasing by fractionation.

Molluscicidal activity of Lawsonia inermis and its binary and tertiary combinations with other plant derived molluscicides.

Molluscicidal activity of leaf, bark and seed of Lawsonia inermis against Lymnaea acuminata and Indoplanorbis exustus was studied. Highest toxicity was observed in the seed of Lawsonia inermis. Toxicity of binary (1:1) and tertiary (1:1:1) combinations of the essential oil of cedar (Cedrus deodara Roxh) and neem (Azadirachta indica A. Juss), powder from bulb of garlic (Allium sativum Linn), and oleoresin extracted from rhizome of ginger (Zingiber officinale Rosc) with Lawsonia inermis and Embelia ribes fruit powder were studied against L. acuminata and I. exustus. L. inermis seed powder in combination with Cedrus deodara oil and Azadirachta indica oil was more toxic than their individual components and other combinations.

Sublethal concentration of Ambrosia maritima(Damsissa) affecting compatibility of Biomphalaria alexandrina snails to infection with Schistosoma mansoni through disturbing the glycolytic pathway.

High glycolytic flux as an emergency pathway for generating ATP was recorded as the most important metabolic pathway required for the success of Biomphalaria-Schistosome sporocyst interaction. Effect of LC25 of dry powdered Ambrosia maritima (Damsissa) as plant molluscicide on hexokinase (HK), pyruvate kinase(PK), glucose phosphate isomerase(GPI) was tested. It resulted in a significant inhibition of the three investigated enzymes. Treatment of snails with LC10 concentrations of A. maritima reduced considerably the infection rate of Biomphalaria alexandrina with Schistosoma mansoni to be 34% compared to an infection rate of 80% in control non-treated snails. Longer prepatent period and remarkable decrease in cercarial production was also recorded in snails treated with the sublethal concentrations of this molluscicide.

Natural rubber base matrix containing Calendula officinalis plant as a source of molluscicdal saponin.

Natural rubber (NR) has been used as a binding matrix for Calendula officinalis, where it is used as a vulcanizate. Various doses of the plant covering the range from 100 up to 200 parts per 100 of rubber (phr) were found suitable for processing conditions using conventional rubber mixing equipment. The release rate of saponin was extended to over 4 months. Incorporation of fillers, such as HAF and Hisil, in the compounded rubber decreased the release of saponin; especially in the first period of immersion. The amount of saponin released was affected by the environmental temperature and the type of fillers present in the formulations. The scanning electron micrographs illustrate the presence of a tortuous pass and pore structure that formed after soaking in water and leaching process occurred.

Evaluation of the molluscicidal properties of Euphorbia splendens var. hislopii (N.E.B.) Latex: experimental test in an endemic area in the state of Minas Gerais, Brazil

Following the positive results obtained regarding the molluscicidal properties of the latex of Euphorbia splendens that were corroborated in laboratory and field tests restrited conditions, a field study was conducted in experimental streams located in an endemic area. After recording the average annual fluctuation of vectors in three streams, a solution of E. splendens latex at ppm was applied in stream A, a solution of niclosamide at 3 ppm that applied in stream B and a third stream (C) remained untreated for negative control. Applications of E. splendens and niclosamide resulted in a mortality of 100 per cent among the snails collected in the streams A and B. No dead snails were found in the negative control stream. A monthly follow-up survey conducted during three consecutive months confirmed the return of vectors to both experimental streams treated with latex and niclosamide. This fact has called for a need to repeat application in order to reach the snails that remained buried in the mud substrate or escaped to the water edge, as well as, newly hatched snails that did not respond to the concentration of these molluscicides. Adults snails collected a month following treatment led us to believe that they had migrate from untreated areas of the streams to those previously treated.

Phenobarbital-type P-450 inducers protect rats against metaldehyde toxicity.

The mechanism of metaldehyde toxicity is unclear. It may be due to the compound itself or, at least in part, to acetaldehyde resulting from the hydrolysis of metaldehyde in the stomach. In this study, we orally dosed rats with twice the LD50 of metaldehyde following no pretreatment (control) or pretreatment with 1 of 3 different cytochrome P-450 inducers either phenobarbital or o,p'-DDD (inducers of cytochromes P-450 IIB and IIIA) or 3-methylcholanthrene (an inducer of P-450 IA). Our results show strong protection against metaldehyde poisoning afforded by the phenobarbital-DDD P-450 inducers, but only weak protection with 3-methylcholanthrene pretreatment. Acetaldehyde administered at the same molarity failed to produce the clinical signs of metaldehyde toxicity and no clinical differences were observed between any of the pretreated groups.

Field tests of a village-based mollusciciding programme for the control of snail hosts of human schistosomes in Cameroon.

The feasibility of using unskilled workers at the district or village level to perform mollusciciding activities in arid regions of northern Cameroon was investigated. A simplified procedure for application to temporary ponds, the principal sites of schistosomiasis transmission in that region, was tested. This consisted of mixing the required quantity of niclosamide (Bayluscide, Bayer AG, Leverkusen) with pond water in a plastic watering can and applying the mixture around the circumference. To aid dispersion, the can was repeatedly swept from the perimeter toward the centre of the pond. Two ponds each were treated once near the end of the rainy season at 1 of 3 concentrations: 0.5, 1.0 and 1.5 p.p.m. Following treatment, 100% snail mortality was recorded at all concentrations. A simple, rapid method for determining the quantity of niclosamide needed to treat temporary ponds was also devised. Physical characteristics were determined for 16 potential schistosomiasis transmission sites in the region. An integration of those characteristics was used to develop a simplified scale indicating the quantity of niclosamide needed to treat ponds based on their circumference. The operational feasibility of a village based mollusciciding programme using those simplified procedures was then evaluated in 2 villages. Village health workers demonstrated a high accuracy (87%) in identifying potential schistosomiasis transmission foci and snail host populations were virtually eliminated from treated sites.

The plant molluscicide Millettia thonningii (Leguminosae) as a topical antischistosomal agent.

The West African legume Millettia thonningii is used in Ghana as an anthelmintic and as a purgative agent. A chloroform extract of the seeds of Millettia thonningii which is known to be molluscicidal and cercaricidal was topically applied to mouse skin 2 and 24 h prior to exposure to Schistosoma mansoni cercariae. The presence of Millettia thonningii extract components on the surface of the skin appeared to be effective in preventing subsequent establishment of infection. The compound responsible for the activity is thought to be the isoflavonoid alpinumisoflavone.