Versatile natural molluscicides against the snail Biomphalaria glabrata Say, 1818 (Pulmonata: Planorbidae), the principal vector of Schistosoma mansoni on the American continent

Versatile natural molluscicides against the snail Biomphalaria glabrata Say, 1818 (Pulmonata: Planorbidae), the principal vector of Schistosoma mansoni on the American continent | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Versatile natural molluscicides against the snail Biomphalaria glabrata Say, 1818 (Pulmonata: Planorbidae), the principal vector of Schistosoma mansoni on the American continent Deborah Bortolucci Hartmann, Renan Alberto Marim, Giuliana Zardeto, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4048314/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Schistosomiasis is a millennial disease that affects approximately 235 million people. Considering that most of the population affected by schistosomiasis faces different conditions of poverty, finding compounds from plants that can help control or eradicate this disease would be worthwhile especially if the plants are readily available for those who need to use them the most. This study aimed to evaluate the effects of fresh latex from fourteen locally cultivated Euphorbiaceae species on the freshwater snail Biomphalaria glabrata Say, 1818 (Gastropoda: Planorbidae), the principal vector of the Schistosoma mansoni trematode on the American continent. Adult B. glabrata snails were exposed to various concentrations of fresh latex from Euphorbiaceae species for 24 and 48 hours. Mortality data were analyzed using the probit method. To evaluate the latex ecotoxicity of the promising species against other aquatic organisms, lethality tests were performed on Poecilia reticulata Peter, 1859 (Cyprinodontiformes: Poeciliidae) fish and Artemia salina Leach (Anostraca: Artemiidae) nauplii. Three species presented significant molluscicide activity: Euphorbia milii var. breonii (LC 50 = 0.9 µg/mL), E. milii var. milii (LC 50 = 1.1 µg/mL) and E. umbellata (LC 50 = 9.1 µg/mL). Only E. milii (var. breonii and var. milii ) was not lethal to the evaluated fish (LC 50 of 9.9 µg/mL) and A. salina (LC 50 > 100.0 µg/mL). These results qualify both the E. milii variety of fresh latices as versatile (active at low concentrations, biodegradable, inexpensive, and obtained from renewable resources) and alternative molluscicidal agents against B. glabrata snails. Tropical Medicine Chemical Biology Health Policy Christ's crown ecotoxicity Euphorbiaceae helminthiasis plant molluscicide snail control 1 Introduction Schistosomiasis is a millennial disease of poverty that leads to chronic ill health. It is caused by trematodes in the Schistosoma genus and affects almost 235 million people worldwide, and more than 700 million people live in endemic areas. The infection is prevalent in tropical and subtropical regions and in poor communities without potable water or adequate sanitation. It is the second most important tropical disease and is responsible for mortality after malaria (WHO, 2023). Over the years, there have been attempts to eradicate this disease. However, the number of cases of this disease has been increasing with progressive consolidation in endemic areas (Coelho & Caldeira, 2016 ). On the American continent, the causative agent of human schistosomiasis is the Schistosoma mansoni trematode. The life cycle of this parasite involves an intermediate host represented by freshwater snails from the genus Biomphalaria , and the B. glabrata snail is the principal vector in South America and the Caribbean (Coelho & Caldeira, 2016 ). The occurrence of mansonic schistosomiasis is closely related to precarious socioenvironmental conditions. The lack of adequate sanitary facilities is one of the major causes of the continuous emergence of new cases of this vermin disease because it is a waterborne disease (Augusto & Silva, 2018 ). Several medications have already been utilized for mansonic schistosomiasis treatment, among which oxamniquine and praziquantel are available in Brazil (Coelho & Caldeira, 2016 ). These substances are indicated for the treatment of individuals who eliminate viable eggs in their feces so that more severe forms can be avoided, disease propagation can be reduced, and a clinical cure can be reached (Augusto & Silva, 2018 ). However, there are too many side effects, and because the financial burden is low, few pharmaceutical companies have developed new drugs for treatment. While the prevalence of this disease is continuous, one of the reasonable options for reducing cases of this parasitic disease could be the control of the intermediate host using substances that have molluscicidal activity to interrupt the developmental cycle of the parasite and, consequently, the emergence of new cases of the disease (Coural & Amaral, 2004 ; Augusto & Silva, 2018 ). Among the known synthetic molluscicides, niclosamide (Bayluscide®) is the most efficient and is recommended by the World Health Organization (WHO). However, the high expense of its application in extensive areas makes its use prohibitive in most developing countries. In addition to the cost, concerns related to the toxicity of nontarget organisms and the possible development of resistance to snails (McCullough et al. 1980 ) have led to a demand for alternative molluscicides of vegetal origin (Clark et al., 1997 ; Alves et al., 2000 ; Augusto & Silva, 2018 ). Many species of tropical plants that have substances with molluscicidal activity, mainly among the Asteraceae, Euphorbiaceae, Fabaceae, and Phytolaccaceae, contain different kinds of substances involved in secondary vegetal metabolism with biocidal action (Kloss & Mccullough, 1982 ; Mendes et al., 1999 ; Luna et al., 2005 ). Considering that most of the population affected by schistosomiasis endures different classes of poverty, finding compounds from plants that can help control or eradicate this disease would be worthwhile especially if the plants are readily available for those who need to use them the most. Therefore, this study aimed to evaluate the effects of fresh latex from fourteen locally cultivated Euphorbiaceae species (native and exotics) on B. glabrata snails. These species were chosen according to the following criteria: wide availability, easy adaptation to tropical regions, toxicity, easy cropping (latex), and easy preparation of the molluscicide solution (fresh latex). Additional tests on ecotoxicity against nontarget organisms at molluscicidal concentrations were performed with the latices of the species that presented potential molluscicidal activity. 2 Materials and Methods 2.1 Plant material All the species utilized in this study were collected in autumn (March-May) at the Medicinal Plant Garden of Paranaense University (Umuarama, Paraná State, Brazil), which is located at an altitude of 430 m above sea level (23 o 46’09.5” S-53 o 16’42.4” W). A voucher for each specimen was deposited at the Educational Herbarium of Paranaense University (“Herbário Educacional da Universidade Paranaense” - HEUP): Euphorbia cotinifolia L. (HEUP-2524); E. milii var. breonii (Nois.) Urch. & Leandri (HEUP-2471); E. milii Des Moul. var. milii (HEUP-2481); E. pulcherrima Willd ex. Klotzsch (HEUP-2552); E. tirucallii L. (HEUP-2529); E. tithymaloides L. [Syn. Pedilanthus tithymaloides (L.) Poit] (HEUP-2270); E. umbellata (Pax) Bruyns [Syn. Synadenium grantii Hook., f.] (HEUP-2476); Jatropha curcas L. (HEUP-2601); J. elliptica (Pohl) Oken (HEUP-2506); J. gossypiifolia L. (HEUP-2272); J. multifida L. (HEUP-2440); J. podagrica Hook. (HEUP-2271); Manihot utilissima Pohl. (HEUP-2603); and Ricinus communis L. (HEUP-2549). 2.2 Fresh latex processing The latex from each species was extracted through incisions in the stems or leaf petioles a little before carrying out the tests. The collected latex was immediately diluted in water at a ratio of 1:9 and quickly transported to the laboratory to prepare the other utilized dilutions. 2.3 Evaluation of molluscicidal activity In this study, Biomphalaria glabrata Say, 1818 (Pulmonata: Planorbidae) snails were utilized for tests of molluscicide activity. The snails were adapted to tanks at a temperature of 23–25°C and under natural light management, according to previous methods (Silva et al., 2008 ). Some samples of guppy fish ( Poecilia reticulata Peter, 1859; Cyprinodontiformes: Poeciliidae) and stems of Elodia canadensis (Hydrocharitaceae) aquatic plants were also adapted to the tank conditions, allowing them to make up the fish tank environment. Food was provided ad libitum and consisted of fresh lettuce ( Lactuca sativa ) for the snails and specific feed for the fish. The bioassay with adult B. glabrata specimens was performed according to the procedures recommended by the WHO (1965, 1983), with minor alterations (Silva et al., 2008 ). First, a stock solution was prepared at three concentrations (100.0, 50.0, and 25.0 µg/mL) for the evaluation of molluscicide potential in fresh latex. Groups of adult mollusks (shell diameter 10–15 mm) were randomly selected and transferred to mini tanks filled with different latex solutions. After 24 h of exposure, the snails were removed, washed with water, and subsequently moved to other mini tanks with dechlorinated water; after 24 h, the recovery time was maintained until mortality was recorded. The control groups were provided only fish tank water without additional substances in all the assays. Assays using niclosamide (Atenase®, UCI-Farma) as the reference molluscicide (0.5 µg/mL) were performed. The snail retraction within the shell or the release of hemolymph were adopted as the death criteria. Moreover, heartbeats were observed using a stereoscopic microscope to verify mollusk mortality. The extracts that caused 100% mortality at the lowest tested concentration (25.0 µg/mL) were selected for an additional test at lower concentrations (25.0, 5.0, 1.0, and 0.5 µg/mL) to determine lethal concentration values at 50% of the tested population (LC 50 ). 2.4 Ecotoxicity assays The ecotoxicity of fresh latices that presented promising molluscicide activity was verified against two nontarget organisms: guppy fish ( P. reticulata ) and brine shrimp ( Artemia salina Leach, Anostraca: Artemiidae). The procedure was like that for the bioassay involving B. glabrata . Fish and A. salina nauplii were separately exposed to different concentrations of promising latices (100.0, 25.0, and 5.0 µg/mL) for 24–48 h at 24–26°C. After this period, the lethality of the strains against populations of fish and nauplii was tested. 2.4 Statistical analysis The results relating to molluscicidal activity and ecotoxicity are expressed as the mean and percentage (%) for lethality tests on snails ( B. glabrata ), guppy fish ( P. reticulata ) and brine shrimp ( A. salina ). Comparisons between mortality rates were performed through analysis of variance (ANOVA) with a significance level of α = 0.05 to observe whether there was any difference between the fresh latices analyzed and the concentration. The estimated lethal concentration to kill 50% of the specimens (LC 50 ) was calculated from statistical software using probit analysis. 3 Results The first phase of this study consisted of evaluating the molluscicide activity of fresh latex from exotic or native species known for its toxicity. In this preliminary approach, B. glabrata snails were subjected to only three concentrations (100.0, 50.0, and 25.0 µg/mL) of fresh latices from fourteen Euphorbiaceae species to determine the most active latices for posterior LC 50 tests. Table 1 shows the mortality rates of the snails in this initial evaluation. The data revealed differences in the efficacy of the molluscicidal activities of the fourteen fresh latices species evaluated ( F = 12.91, p 0.05). Table 1 Preliminary evaluation of molluscicide activity of fresh latices from several exotic species of Euphorbiaceae against Biomphalaria glabrata snails after 24 h of exposure and recovery. Latex % lethality 100.0 µg/mL 50.0 µg/mL 25.0 µg/mL Euphorbia cotinifolia 0 0 0 Euphorbia milii var. breonii 100 100 100 Euphorbia milii var. milii 100 100 100 Euphorbia pulcherrima 0 0 0 Euphorbia tirucallii 100 10 0 Euphorbia tithymaloides 0 0 0 Euphorbia umbellata 100 100 100 Jatropha curcas 0 0 0 Jatropha elliptica 0 0 0 Jatropha gossypiifolia 0 0 0 Jatropha multifida 0 0 0 Jatropha podagrica 0 0 0 Manihot utilissima 100 6.7 0 Ricinus communis 0 0 0 Positive control: niclosamide (0.5 µg/mL). In this preliminary phase, B. glabrata snails were strongly affected when exposed to fresh latices from Euphorbia milii (var. breonii and var. milii ) and E. umbellata species. Due to their high molluscicide potential, these species were reevaluated at lower concentrations to determine the LC 50 value. Fresh latices from E. tirucallii and Manihot utilissima were also active but only at the highest tested concentration (100 µg/mL). The other species did not exhibit activity at the highest tested concentration. According to the recommendations of the WHO (1983), only aqueous or alcoholic extracts of vegetal materials that cause the death of 90% of the malacological population at concentrations lower than 20 µg/mL are considered potentially active and viable for field assays, while less active extracts may very well provide sources of new lead compounds with molluscicidal activities. Considering the observed results for E. milii var. breonii (LC 50 = 0.9 µg/mL), E. milii var. milii (LC 50 = 1.1 µg/mL) and E. umbellata (LC 50 = 9.1 µg/mL), their fresh latices can be classified as potential molluscicide agents (Table 2 ). No significant difference between these potent fresh latices was detected ( F = 3.10, p > 0.05); however, the lethality data as a function of concentration were significantly different ( F = 10.44, p < 0.05). Table 2 Complementary tests of the molluscicide activity of fresh latices from Euphorbia milii and Synadenium grantii against adult Biomphalaria glabrata snails after 24 h of exposure and recovery. Latex % lethality LC 50 (µg/mL) 25.0 µg/mL 5.0 µg/mL 1.0 µg/mL 0.5 µg/mL Euphorbia milii var. breonii 100 90 50 10 0.9 Euphorbia milii var. milii 100 90 50 0 1.1 Synadenium grantii 100 10 0 0 9.1 Positive control: niclosamide (0.5 µg/mL). Ecotoxicity tests were carried out to evaluate the toxicity of the fresh latices from E. milii (var. breonii and var. milii ) and E. umbellata plants against nontarget organisms such as fish and small crustaceans (Table 3 ). Guppy fish ( P. reticulata ) was used as the nontarget organism in this study because it is sensitive to contaminants and is easily found in rivers and lagoons in Brazil, providing results with great practical significance. A. salina , commonly known as brine shrimp, is a small crustacean that has been the subject of many physiological studies. The brine shrimp lethality assay is one of the most practical tools for the preliminary assessment of general toxicity (McLaughlin et al., 1991 ); moreover, it is a good option for ecotoxicity assays because it is a sensitive nontarget organism and is usually utilized in biomonitoring or detection of toxic or bioactive compounds in vegetal extracts. Table 3 Evaluation of the ecotoxic effects of different concentrations of fresh lattices from S. grantii and E. milii L. on guppy fish ( Poecilia reticulata ) and brine shrimp ( Artemia salina ) after 24 h of exposure and recovery. Latex Concentration (µg/mL) % lethality P. reticulata A. salina Euphorbia milii var. breonii 100.0 100 0 25.0 100 0 5.0 0 0 LC 50 9.9 µg/mL > 100 µg/mL Euphorbia milii var. milii 100.0 100 0 25.0 100 0 5.0 0 0 LC 50 9.9 µg/mL > 100 µg/mL Synadenium grantii 100.0 100 10 25.0 100 0 5.0 0 0 LC 50 9.9 µg/mL > 100 µg/mL E. umbellata and both varieties of E. milii ( breonii and milii ) were completely lethal to guppy fish at a concentration of 25.0 µg/mL, with an LC 50 = 9.9 µg/mL (Table 3 ). These fresh latices did not show toxicity against A. salina (LC 50 > 100 µg/mL). No statistically significant difference was found ( p > 0.05). 4 Discussion The control of snail vectors is an alternative way to prevent schistosomiasis. This strategy is particularly relevant in endemic localities in rural and urban areas, where the lack of sanitation and adequate housing show little potential for improvement in the future (Augusto & Silva, 2018 ). Efforts are being made to discover molluscicide products of natural origin that are potentially biodegradable to address the growing awareness of environmental pollution. In this sense, this work focused its attention on the search for plant-based molluscicides obtained by simple, inexpensive, and efficient extraction and application techniques that are capable of being used by needy communities affected by schistosomiasis, opening possibilities for sustainable improvements in the health of these communities. Generally, latex is an aqueous emulsion found in the vacuole of secretory cells and is composed of lipids, resins, sugars, proteins, and enzymes. It can be easily obtained and manipulated for the preparation of molluscicides, dispensing the use of solvents and equipment for the preparation of extracts or extraction of active components. The use of latex from locally grown species by communities associated with health education programmes can help keep schistosomiasis at low levels since eradication is not considered realistic under current conditions in some developing countries. Among the fourteen Euphorbiaceae species evaluated in this study, only the fresh latices of three species were potentially active as molluscicides: E. milii var. breonii , E. milii var. milii , and E. umbellata (Table 2 ). Originally from Madagascar, Euphorbia milii Des Moulin’s species is known in Brazil, where some varieties are cultivated as ornamental plants or hedges. In the present study, fresh latex solution of E. milii var. breonii had different molluscicidal effects than did the crude extract of leaves of this same species, which did not show molluscicidal activity in a previous study (LC 50 > 100 µg/mL) (Hartmann et al., 2011 ). The latex at a concentration of 5.0 µg/mL exhibited 100% lethality in the first 24 h of the experiment. After 48 h, the lethal concentration for 50% of the exposed population was calculated to be 0.9 µg/mL. This difference in activity between fresh latex and the crude extract (Hartmann et al., 2011 ) is probably due to the concentration of substances responsible for molluscicidal activity. The concentration of the active substance in the leaf extract was likely low, or the substance could have degraded or polymerized during extract preparation. Curiously, the results observed for the breonii variety were very close to those obtained for the milii variety (Table 1 ; LC 50 = 1.1 µg/mL). These concentrations are much lower than the WHO (1983) recommended concentrations for molluscicide activity. It was also noted that the calculated LC 50 value of the latex was very close to that reported in the literature for some species and varieties known as Christ's crown or crown-of-thorns (LC 50 = 0.12 to LC 50 = 2.0 µg/mL) (Vasconcelos & Schall, 1986 ; Baptista et al., 1994 ; Carvalho et al., 1998; Oliveira-Filho & Paumgartten, 2000 ; Coêlho et al., 2018 ). E. milii Des Moul. Ex Boiss var. hislopii (syn. E. splendens ) lyophilized latex is one of the most potent molluscicide plants tested in outlying areas. Zani et al. ( 1993 ) isolated eight categories of ingenol esters (miliamines) from E. milii var. hislopii and tested their molluscicide activities on B. glabrata , with miliamine L being the most efficient, with lethality occurring at concentrations as low as 1.0 ppb. The latex of this variety was also very toxic to other Planorbidae snails (Oliveira-Filho & Paumgartten, 2000 ), suggesting that fresh latex from E. milii var. milii and var. breoni could also be good molluscicides against other snail species. Another relevant aspect of extract concentrations is concern about other components of aquatic biota. Even though natural molluscicides are biodegradable at specific concentrations, these extracts may have risks even within the values demanded by the WHO. Therefore, toxicity tests showed that the use of latices from E. milii (var. breonii and var. milii ) as molluscicide agents would be favorable because both species are lethal to snails at lower concentrations (LC 50 ~ 1.0 µg/mL) than are the ones that are lethal to guppy fish (LC 50 = 9.9 µg/mL) and A. salina microcrustaceans (LC 50 > 100.0 µg/mL). Therefore, these latices can be considered atoxic to the environment at molluscicide concentrations (LC 50 ~ 1.0 µg/mL). These data corroborated those of Oliveira-Filho and Paumgartten ( 2000 ), who analyzed the ecotoxicity of lyophilized latex from E. milii var. hislopii . E. umbellata (syn. Synadenium grantii ) is an African species cultivated as an exotic ornamental plant in Brazil where it is considered highly toxic. The population utilizes latex to reduce warts, treat gastric diseases, and even treat cancer (Kinghorn, 1980 ). According to the literature, the chemical composition of latex from this species includes derivatives of phorbol esters (Kinghorn, 1980 ; Bagavathi et al., 1988 ), triterpenoids (Uzabakiliho et al., 1987 ), proteolytic enzymes (Menon et al., 2002 ), carboxylesterase enzymes (Govindappa et al., 1987 ), acetylcholinesterase isoenzymes (Govindappa et al., 1987 ) and glycoproteins (Rajesh et al., 2006 ). During the evaluation of the molluscicidal activity of fresh latex from this species, it was found to be potent (Table 2 ). As previously mentioned for E. milli var. breoni , the crude alcoholic extract of E. umbellata leaves is less active (LC 50 = 40.0 µg/mL; Hartmann et al., 2011 ) than fresh latex is. Regarding ecotoxicity, in the case of fresh E. umbellata latex, the lethal concentration for B. glabrata (LC 50 = 9.1 µg/mL) was close to that of guppy fish (LC 50 = 9.9 µg/mL) (Tables 2 and 3 ). Thus, although the latex from this species is considered a good molluscicide agent, its use requires ample care due to its toxicity to other organisms present in the aquatic environment at the same concentration, as latex is lethal to the vector snail of Schistosoma mansoni. These data differed from those achieved by Pereira et al. (2017), who reported an LC 50 = 1.36 µg/mL for lyophilized latex. These authors also performed assays with Danio regio (zebrafish) to investigate the toxicity of E. umbellata latex to nontarget organisms, and they found that this latex was less toxic to zebrafish (LC 50 = 7.22 µg/mL) than to B. glabrata (LC 50 = 1.36 µg/mL). The difference in activity observed between our results and those of the authors (Pereira et al., 2017) may be related to different factors: type of latex (fresh/lyophilized), latex harvesting period (autumn/spring), and geography (harvesting in the area close to the Tropic of Capricorn/harvesting in the equatorial area-coast). Another evaluated species in this study also presented latices with moderate molluscicidal activity. In Brazil, the African milkbush or aveloz plant ( E. tirucallii ), which is very caustic and popularly used for wart scaring, is known for its latex toxicity. As part of the screening of the molluscicide species, latex from this species was evaluated and found to have moderate activity against B. glabrata , with an LC 50 = 89.4 µg/mL. Cassava ( Manihot utilissima Pohl) is a native plant known for its great usefulness in the economy and history of Brazil. One of its varieties, “mandioca-brava”, is also known for poisoning cases in humans and animals. Considering the toxic character of your latex, its possible molluscicide activity was evaluated. Latex from leaf petioles of M. utilissima also presented moderate molluscicidal activity (LC 50 = 95.0 µg/mL). Latex from Barbados nuts ( Jatropha curcas ) did not exhibit activity at the highest tested concentration (100 µg/mL; Table 1 ) in the present study. Other authors have observed significant molluscicidal activity of J. curcas against different vector snails (Liu et al., 1997 ; Rug & Ruppel, 2000 ). According to Nath and Dutta ( 1991 ), latex from J. curcas has a protolithic enzyme called curcain. Another study indicated two significant activities for J. curcas latex: procoagulant and anticoagulant (Osoniyi & Onajobi, 2003 ). Therefore, even if latex contains interesting substances related to other activities, this does not indicate who is a good molluscicide. Latices from another species of this genus were evaluated in this study ( J. elliptica, J. gossypifolia L, J. multifida L, and J. podagrica ); however, none of them presented molluscicidal activity at the highest tested concentration (Table 1 ). 5 Conclusions Among the several Euphorbiaceae species evaluated in this study, only E. milii (var. breonii and var. milii ) and E. umbellata presented potent molluscicidal effects on B. glabrata snails. The use of fresh latices from E. milii varieties represents a feasible alternative strategy for schistosomiasis control because these plants can be efficiently grown in endemic areas (not requiring soil care, fertilizers, or water); additionally, these plants yield a relatively large amount of latex throughout the year, and the latex can be easily collected. In summary, these fresh latices are active at low concentrations, are biodegradable and inexpensive, and are obtained from renewable resources. The preparation of these molluscicides is straightforward, does not require organic solvents (typically used to prepare extracts) or equipment (rotary evaporator or freeze dryer), and can be easily prepared by the population most affected by schistosomiasis. The molluscicidal activity of fresh latex from E. milii var. breoni and E. milii var. milii latices is reported here for the first time. On the other hand, the fresh latex from E. umbellata was strongly lethal to B. glabrata snails (LC 50 = 9.1 µg/mL) and guppy fishes (LC 50 = 9.9 µg/mL). Consequently, their use as molluscicides is contraindicated because they require special attention due to their possible toxicity against nontarget organisms. Declarations Acknowledgments R. A. M. thanks Diretoria Executiva de Gestão da Pesquisa e da Pós-graduação - DEGPP (Executive Board of Research and Postgraduate Management)/UNIPAR for scholarship. The authors thank Dr. Felipe A. La Porta (UTFPR) for assistance with the statistical analyses. Author Contributions D. B. H., Z. C. G. and A. L. J. contributed to the conception and design of the experiments. E. J. contributed to the collection and identification of the botanical material . D. B. H., R. A. M., G. Z., I. A. S., and E. J. contributed to the data acquisition. D. B. H., R. A. M., and A. L. J. contributed to the analysis and interpretation of the data. A. L. J. supervised the project and wrote the manuscript with input from all the authors. All the authors read and agreed to the publication of this version of the article. Funding This project was financially supported by Paranaense University (UNIPAR - Proj: 15950 and 18004). Competing interests The authors declare no conflicts of interest. Ethical approval Does not apply. Informed consent Does not apply. References Alves, T. M. D., Silva, A. F., Brandão, M., Grandi, T. S. M., Smânia, E. F. A., Smânia, A., & Zani, C. L. (2000). Biological screening of Brazilian medicinal plants. Memórias do Instituto Oswaldo Cruz , 95 (3), 367-373. https://doi.org/10.1590/S0074-02762000000300012 Augusto, R. C., & Silva, C. C. M. (2018). Phytochemical Molluscicides and Schistosomiasis: What We Know and What We Still Need to Learn. Veterinary Sciences , 5 (4), 94. https://doi.org/10.3390/vetsci5040094 Bagavathi, R., Sorg, B., & Hecker, E. (1988). Tigliane-Type Diterpene Esters from Synadenium grantii. Planta Medica , 54 (6), 506-510. https://doi.org/10.1055/s-2006-962531 Baptista, D. F., Vasconcellos, M. C., Lopes, F. E., Silva I. P., & Schall V. T. 1994. Perspective of using Euphorbia splendens as a molluscicide in schistosomiasis control programs. Southeast Asian Journal of Tropical Medicine and Public Health , 25 (3), 419-424. Available at: https://www.arca.fiocruz.br/handle/icict/15715 Clark, T. E., Appleton, C. C., & Drewes, S. E. (1997). A semi-quantitative approach to the selection of appropriate candidate plant molluscicides a South African application. Journal of Ethnopharmacology , 56 (1), 1-13. https://doi.org/10.1016/S0378-8741(96)01495-X Coêlho, M. D. G., Lino, F. P. S., Akisue, G., Maciel, L. T. R., Bozo, L. S. O., & Coêlho, F. A. Z. (2018). Evaluation of toxicological and molluscicidal activities of lactiferous plants against Biomphalaria glabrata (Say, 1818). Scientia Parasitologica , 19 (1-2), 1-6. Available at: http://www.zooparaz.net/scientia/ Coelho, P. M. Z., & Caldeira, R. L. (2016). Critical analysis of molluscicide application in schistosomiasis control programs in Brazil. Infectious Diseases of Poverty , 5 , 57. https://doi.org/10.1186/s40249-016-0153-6 Coural, J. R., & Amaral, R. S. (2004). Epidemiological and control aspects of schistosomiasis in Brazilian endemic areas. Memórias do Instituto Oswaldo Cruz , 99 (suppl 1), 13-19. https://doi.org/10.1590/S0074-02762004000900003 Govindappa, T., Govardhan, L., Jyothy, P. S., & Veerabhadrappa, P. S. (1987). Purification and characterization of acetylcholinesterase isozymes from the latex of Synadenium-grantii Hook, F. Indian Journal of Biochemistry and Biophysics , 24 , 209-217. Hartmann, D.B., Marim, R.A., Silva, Y.L., Zardeto, G., Silva, I.Z., Mattos, D.A., & Laverde Junior, A. (2011). Letalidade do extrato de Synadenium grantii Hook. f. (Euphorbiaceae) frente a caramujos Biomphalaria glabrata Say, 1818 (Gastropoda, Planorbidae). Arquivos de Ciências Veterinárias e Zoológicas da UNIPAR , 14 (1), 5-11. Available at: https://revistas.unipar.br/index.php/veterinaria/article/view/3779/2456 Kinghorn, A.D. (1980). Major skin-irritant principle from Synadenium grantii. Journal of Pharmaceutical Sciences , 69 (12), 1446-1447. https://doi.org/10.1002/jps.2600691228 Kloss, H., & Mccullough, F. S. (1982). Plant molluscicides. Planta Medica , 46 (12), 195-209. https://doi.org/10.1055/s-2007-971215 Liu, S. Y., Sporer, F., Wink, M., Jourdane, J., Henning, R, Li, Y.L., & Ruppel, A. (1997). Anthraquinones in Rheum palmatum and Rumex dentatus (Polygonaceae), and pho.rbol esters in Jatropha curcas (Euphorbiaceae) with molluscicidal activity against the schistosome vector snails Oncomelania , Biomphalaria and Bulinus. Tropical Medicine and International Health , 2 (2), 179-188. https://doi.org/10.1046/j.1365-3156.1997.d01-242.x Luna, J. S., Santos, A. F., Lima, M. R. F, Omena, M. C., Mendonça, F. A. C., Bieber, L. W., & Sant’ana, A. E. G. (2005). A study of the larvicidal and molluscicidal activities of some medicinal plants from northeast Brazil. Journal of Ethnopharmacology , 97 (2), 199-206. https://doi.org/10.1016/j.jep.2004.10.004 McCullough, F. S., Gayral, P. H., Duncan, J., & Christie, J. D. 1980. Molluscicides in schistosomiasis control. Bulletin of the World Health Organization , 58 (5), 681-689. https://apps.who.int/iris/handle/10665/262070 McLaughlin, J. L., Chang, C. J., & Smith, D. L. (1991). “Bench-Top” bioassays for the discovery of bioactive natural products: an update. In: Atta-ur-Rahman (ed), Studies in Natural Products Chemistry , (Vol. 9, p. 383) Elsevier. Mendes, N. M., Queiroz, R. O., Grandi, T. S. M., Anjos, A. M. G., & Zani, C.L. (1999). Screening of Asteraceae (Compositae) plant extrats for molluscicidal activity. Memórias do Instituto Oswaldo Cruz , 94 (3), 411-412. https://doi.org/10.1590/S0074-02761999000300023 Menon, M., Vithayathil, P. J., Raju, S. M, & Ramadoss, C. S. (2002). Isolation and characterization of proteolytic enzymes from the latex of Synadenium grantii Hook, 'f'. Plant Science , 163 (1), 131-139. https://doi.org/10.1016/S0168-9452(02)00085-7 Nath, L. K., & Dutta, S. K. (1991). Extraction and purification of curcain, a protease from the latex of Jatropha curcas Linn. Journal of Pharmacy and Pharmacology , 43 (2), 111-114. https://doi.org/10.1111/j.2042-7158.1991.tb06642.x Oliveira-Filho, E. C., & Paumgartten, F. J. R. (2000). Toxicity of Euphorbia milli latex and Niclosamide to snails and non-target aquatic species. Ecotoxicology and Environmental Safety , 46 (3), 342-350. https://doi.org/10.1006/eesa.2000.1924 Osoniyi, O., & Onajobi, F. (2003). Coagulant and anticoagulant activities in Jatropha curcas latex. Journal of Ethnopharmacology , 89 (1), 101-105. https://doi.org/10.1016/s0378-8741(03)00263-0 Rajesh, R., Nataraju, A., Gowda, C. D. R., Frey, B. M., Frey, F. J., & Vishwanath, B. S. (2006). Purification and characterization of a 34-kDa, heat stable glycoprotein from Synadenium grantii latex: action on human fibrinogen and fibrin clot. Biochimie , 88 (10), 1313-1322. https://doi.org/10.1016/j.biochi.2006.06.007 Rug, M., & Ruppel, A. (2000). Toxic activities of the plant Jatropha curcas against intermediate snail hosts and larvae of schistosomes. Tropical Medicine and International Health , 5 (6), 423-430. https://doi.org/10.1046/j.1365-3156.2000.00573.x Silva, N. F. S., Cogo, J., Wiepieski, C. C. P., & Laverde Junior, A. (2008). Bioensaio de atividade moluscicida adaptado à avaliação de extratos de plantas medicinais. Arquivos de Ciências Veterinárias e Zoológicas da UNIPAR , 11 (2), 179-181. Available at: https://revistas.unipar.br/index.php/veterinaria/article/view/2575 Uzabakiliho, B., Largeau, C., & Casadevall, E. (1987). Latex constituents of Euphorbia candelabrum , Euphorbia grantii , Euphorbia tirucalli and Synadenium grantii . Phytochemistry, 26(11), 3041-3045. https://doi.org/10.1016/S0031-9422(00)84589-6 Vasconcelos, M. C., & Schall, V. T. (1986). Latex of “Coroa de Cristo” ( Euphorbia splendens ): an effective molluscicide. Memórias do Instituto Oswaldo Cruz , 81 (4), 475-476. https://doi.org/10.1590/S0074-02761986000400017 World Health Organization (WHO) (1965). Memoranda - molluscicide screening and evaluation. Bulletin of the World Health Organization , 33 (4), 567-581. Available at: https://apps.who.int/iris/handle/10665/262863 World Health Organization (WHO) (1983). Reports of the scientific working group on plant molluscicides. Bulletin of the World Health Organization , 61 , 927. Available at: https://apps.who.int/iris/handle/10665/60086 World Health Organization (WHO) (2023, February 1). Fact sheet Nº115: Schistosomiasis, Geneva, World Health Organization. Retrieved February 15, 2024, from: http://www.who.int/mediacentre/factsheets/fs115/en/ Zani, C. L., Marston, A., Hamburger, M., & Hostettmann, K. (1993). Molluscicidal milliamines from Euphorbia milii var. hislopii . Phytochemistry , 34 (1), 89-95. https://doi.org/10.1016/S0031-9422(00)90788-X Additional Declarations The authors declare no competing interests. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4048314","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":277512733,"identity":"a81c0b71-d09d-43dc-8605-3bbe7aabcdc3","order_by":0,"name":"Deborah Bortolucci Hartmann","email":"","orcid":"https://orcid.org/0009-0006-6866-1210","institution":"Paranaense University","correspondingAuthor":false,"prefix":"","firstName":"Deborah","middleName":"Bortolucci","lastName":"Hartmann","suffix":""},{"id":277515476,"identity":"bb1a3ee1-9645-43ae-a8c6-37f970f0a83a","order_by":1,"name":"Renan Alberto Marim","email":"","orcid":"https://orcid.org/0000-0003-3855-6971","institution":"Integrado University Center","correspondingAuthor":false,"prefix":"","firstName":"Renan","middleName":"Alberto","lastName":"Marim","suffix":""},{"id":277515477,"identity":"c2922af7-66da-4a0f-b69c-2647d156444e","order_by":2,"name":"Giuliana Zardeto","email":"","orcid":"https://orcid.org/0000-0003-1640-0714","institution":"Paranaense University","correspondingAuthor":false,"prefix":"","firstName":"Giuliana","middleName":"","lastName":"Zardeto","suffix":""},{"id":277515478,"identity":"4a017c72-8d15-40f1-b416-ca9dd2d01532","order_by":3,"name":"Isabela de Azevedo Silva","email":"","orcid":"https://orcid.org/0009-0005-1056-615X","institution":"Paranaense University","correspondingAuthor":false,"prefix":"","firstName":"Isabela","middleName":"de Azevedo","lastName":"Silva","suffix":""},{"id":277515479,"identity":"ae1ce46b-907e-4114-8a70-0ae76061ef4c","order_by":4,"name":"Ezilda Jacomassi","email":"","orcid":"https://orcid.org/0000-0003-0967-8427","institution":"Paranaense University","correspondingAuthor":false,"prefix":"","firstName":"Ezilda","middleName":"","lastName":"Jacomassi","suffix":""},{"id":277515480,"identity":"08fb7c43-6756-4496-9bd0-248a754ac0f0","order_by":5,"name":"Zilda Cristiani Gazim","email":"","orcid":"https://orcid.org/0000-0003-0392-5976","institution":"Paranaense University","correspondingAuthor":false,"prefix":"","firstName":"Zilda","middleName":"Cristiani","lastName":"Gazim","suffix":""},{"id":277515481,"identity":"5cb0a752-1ea3-49ce-872f-159a11cdd3c1","order_by":6,"name":"Antonio Laverde Junior","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA5UlEQVRIie3SPQrCMBTA8VeEdgl2TRH0Ci8UBLGHSSl0tW5uChlcPEC8hZOTQ6SgSw8QcCoFZw/gYPxEEKKjQ/5LHoEfb0gAXK5/LAAOgAChmU+3GwW+nbQeJJq1PPkruYfqVxIKOHaKIhnFerQV4w1025r7zcRCaAl5R2I+WOvME/IIcaR5wCrbmishWGJfZ0FDFKQrsyWaWUTvSWJpthgy/UrwSZDeCcdvhJWQDQnmSKuGCakoW1b1nNlId79ID+ScYDhPa1GopNfeZ7vaRgAIf43mjag5PDswP0a9E5fL5XJ9dgFMd0kfI1irHgAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0000-0001-8346-6217","institution":"Federal Technological University of Paraná","correspondingAuthor":true,"prefix":"","firstName":"Antonio","middleName":"Laverde","lastName":"Junior","suffix":""}],"badges":[],"createdAt":"2024-03-08 18:44:24","currentVersionCode":1,"declarations":{"humanSubjects":false,"vertebrateSubjects":false,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":false},"doi":"10.21203/rs.3.rs-4048314/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4048314/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":52482353,"identity":"6cd0beb7-d4c1-436e-97e2-808abedae3e1","added_by":"auto","created_at":"2024-03-12 06:23:10","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":314087,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4048314/v1/febff302-5fc4-42c0-a912-a932d4753036.pdf"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003e\u003cstrong\u003eVersatile natural molluscicides against the snail \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eBiomphalaria glabrata \u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003eSay, 1818 (Pulmonata: Planorbidae), the principal vector of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eSchistosoma mansoni\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e on the American continent\u003c/strong\u003e\u003c/p\u003e","fulltext":[{"header":"1 Introduction","content":"\u003cp\u003eSchistosomiasis is a millennial disease of poverty that leads to chronic ill health. It is caused by trematodes in the \u003cem\u003eSchistosoma\u003c/em\u003e genus and affects almost 235\u0026nbsp;million people worldwide, and more than 700\u0026nbsp;million people live in endemic areas. The infection is prevalent in tropical and subtropical regions and in poor communities without potable water or adequate sanitation. It is the second most important tropical disease and is responsible for mortality after malaria (WHO, 2023).\u003c/p\u003e \u003cp\u003eOver the years, there have been attempts to eradicate this disease. However, the number of cases of this disease has been increasing with progressive consolidation in endemic areas (Coelho \u0026amp; Caldeira, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). On the American continent, the causative agent of human schistosomiasis is the \u003cem\u003eSchistosoma mansoni\u003c/em\u003e trematode. The life cycle of this parasite involves an intermediate host represented by freshwater snails from the genus \u003cem\u003eBiomphalaria\u003c/em\u003e, and the \u003cem\u003eB. glabrata\u003c/em\u003e snail is the principal vector in South America and the Caribbean (Coelho \u0026amp; Caldeira, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). The occurrence of mansonic schistosomiasis is closely related to precarious socioenvironmental conditions. The lack of adequate sanitary facilities is one of the major causes of the continuous emergence of new cases of this vermin disease because it is a waterborne disease (Augusto \u0026amp; Silva, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eSeveral medications have already been utilized for mansonic schistosomiasis treatment, among which oxamniquine and praziquantel are available in Brazil (Coelho \u0026amp; Caldeira, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). These substances are indicated for the treatment of individuals who eliminate viable eggs in their feces so that more severe forms can be avoided, disease propagation can be reduced, and a clinical cure can be reached (Augusto \u0026amp; Silva, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). However, there are too many side effects, and because the financial burden is low, few pharmaceutical companies have developed new drugs for treatment. While the prevalence of this disease is continuous, one of the reasonable options for reducing cases of this parasitic disease could be the control of the intermediate host using substances that have molluscicidal activity to interrupt the developmental cycle of the parasite and, consequently, the emergence of new cases of the disease (Coural \u0026amp; Amaral, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2004\u003c/span\u003e; Augusto \u0026amp; Silva, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eAmong the known synthetic molluscicides, niclosamide (Bayluscide\u0026reg;) is the most efficient and is recommended by the World Health Organization (WHO). However, the high expense of its application in extensive areas makes its use prohibitive in most developing countries. In addition to the cost, concerns related to the toxicity of nontarget organisms and the possible development of resistance to snails (McCullough et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e1980\u003c/span\u003e) have led to a demand for alternative molluscicides of vegetal origin (Clark et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e1997\u003c/span\u003e; Alves et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Augusto \u0026amp; Silva, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eMany species of tropical plants that have substances with molluscicidal activity, mainly among the Asteraceae, Euphorbiaceae, Fabaceae, and Phytolaccaceae, contain different kinds of substances involved in secondary vegetal metabolism with biocidal action (Kloss \u0026amp; Mccullough, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e1982\u003c/span\u003e; Mendes et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e1999\u003c/span\u003e; Luna et al., \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2005\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eConsidering that most of the population affected by schistosomiasis endures different classes of poverty, finding compounds from plants that can help control or eradicate this disease would be worthwhile especially if the plants are readily available for those who need to use them the most. Therefore, this study aimed to evaluate the effects of fresh latex from fourteen locally cultivated Euphorbiaceae species (native and exotics) on \u003cem\u003eB. glabrata\u003c/em\u003e snails. These species were chosen according to the following criteria: wide availability, easy adaptation to tropical regions, toxicity, easy cropping (latex), and easy preparation of the molluscicide solution (fresh latex). Additional tests on ecotoxicity against nontarget organisms at molluscicidal concentrations were performed with the latices of the species that presented potential molluscicidal activity.\u003c/p\u003e"},{"header":"2 Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Plant material\u003c/h2\u003e \u003cp\u003eAll the species utilized in this study were collected in autumn (March-May) at the Medicinal Plant Garden of Paranaense University (Umuarama, Paran\u0026aacute; State, Brazil), which is located at an altitude of 430 m above sea level (23\u003csup\u003eo\u003c/sup\u003e46\u0026rsquo;09.5\u0026rdquo; S-53\u003csup\u003eo\u003c/sup\u003e16\u0026rsquo;42.4\u0026rdquo; W). A voucher for each specimen was deposited at the Educational Herbarium of Paranaense University (\u0026ldquo;Herb\u0026aacute;rio Educacional da Universidade Paranaense\u0026rdquo; - HEUP): \u003cem\u003eEuphorbia cotinifolia\u003c/em\u003e L. (HEUP-2524); \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e (Nois.) Urch. \u0026amp; Leandri (HEUP-2471); \u003cem\u003eE. milii\u003c/em\u003e Des Moul. var. \u003cem\u003emilii\u003c/em\u003e (HEUP-2481); \u003cem\u003eE. pulcherrima\u003c/em\u003e Willd ex. Klotzsch (HEUP-2552); \u003cem\u003eE. tirucallii\u003c/em\u003e L. (HEUP-2529); \u003cem\u003eE. tithymaloides\u003c/em\u003e L. [Syn. \u003cem\u003ePedilanthus tithymaloides\u003c/em\u003e (L.) Poit] (HEUP-2270); \u003cem\u003eE. umbellata\u003c/em\u003e (Pax) Bruyns [Syn. \u003cem\u003eSynadenium grantii\u003c/em\u003e Hook., f.] (HEUP-2476); \u003cem\u003eJatropha curcas\u003c/em\u003e L. (HEUP-2601); \u003cem\u003eJ. elliptica\u003c/em\u003e (Pohl) Oken (HEUP-2506); \u003cem\u003eJ. gossypiifolia\u003c/em\u003e L. (HEUP-2272); \u003cem\u003eJ. multifida\u003c/em\u003e L. (HEUP-2440); \u003cem\u003eJ. podagrica\u003c/em\u003e Hook. (HEUP-2271); \u003cem\u003eManihot utilissima\u003c/em\u003e Pohl. (HEUP-2603); and \u003cem\u003eRicinus communis\u003c/em\u003e L. (HEUP-2549).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Fresh latex processing\u003c/h2\u003e \u003cp\u003eThe latex from each species was extracted through incisions in the stems or leaf petioles a little before carrying out the tests. The collected latex was immediately diluted in water at a ratio of 1:9 and quickly transported to the laboratory to prepare the other utilized dilutions.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 Evaluation of molluscicidal activity\u003c/h2\u003e \u003cp\u003eIn this study, \u003cem\u003eBiomphalaria glabrata\u003c/em\u003e Say, 1818 (Pulmonata: Planorbidae) snails were utilized for tests of molluscicide activity. The snails were adapted to tanks at a temperature of 23\u0026ndash;25\u0026deg;C and under natural light management, according to previous methods (Silva et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). Some samples of guppy fish (\u003cem\u003ePoecilia reticulata\u003c/em\u003e Peter, 1859; Cyprinodontiformes: Poeciliidae) and stems of \u003cem\u003eElodia canadensis\u003c/em\u003e (Hydrocharitaceae) aquatic plants were also adapted to the tank conditions, allowing them to make up the fish tank environment. Food was provided \u003cem\u003ead libitum\u003c/em\u003e and consisted of fresh lettuce (\u003cem\u003eLactuca sativa\u003c/em\u003e) for the snails and specific feed for the fish. The bioassay with adult \u003cem\u003eB. glabrata\u003c/em\u003e specimens was performed according to the procedures recommended by the WHO (1965, 1983), with minor alterations (Silva et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). First, a stock solution was prepared at three concentrations (100.0, 50.0, and 25.0 \u0026micro;g/mL) for the evaluation of molluscicide potential in fresh latex. Groups of adult mollusks (shell diameter 10\u0026ndash;15 mm) were randomly selected and transferred to mini tanks filled with different latex solutions. After 24 h of exposure, the snails were removed, washed with water, and subsequently moved to other mini tanks with dechlorinated water; after 24 h, the recovery time was maintained until mortality was recorded. The control groups were provided only fish tank water without additional substances in all the assays. Assays using niclosamide (Atenase\u0026reg;, UCI-Farma) as the reference molluscicide (0.5 \u0026micro;g/mL) were performed. The snail retraction within the shell or the release of hemolymph were adopted as the death criteria. Moreover, heartbeats were observed using a stereoscopic microscope to verify mollusk mortality. The extracts that caused 100% mortality at the lowest tested concentration (25.0 \u0026micro;g/mL) were selected for an additional test at lower concentrations (25.0, 5.0, 1.0, and 0.5 \u0026micro;g/mL) to determine lethal concentration values at 50% of the tested population (LC\u003csub\u003e50\u003c/sub\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Ecotoxicity assays\u003c/h2\u003e \u003cp\u003eThe ecotoxicity of fresh latices that presented promising molluscicide activity was verified against two nontarget organisms: guppy fish (\u003cem\u003eP. reticulata\u003c/em\u003e) and brine shrimp (\u003cem\u003eArtemia salina\u003c/em\u003e Leach, Anostraca: Artemiidae). The procedure was like that for the bioassay involving \u003cem\u003eB. glabrata\u003c/em\u003e. Fish and \u003cem\u003eA. salina\u003c/em\u003e nauplii were separately exposed to different concentrations of promising latices (100.0, 25.0, and 5.0 \u0026micro;g/mL) for 24\u0026ndash;48 h at 24\u0026ndash;26\u0026deg;C. After this period, the lethality of the strains against populations of fish and nauplii was tested.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Statistical analysis\u003c/h2\u003e \u003cp\u003eThe results relating to molluscicidal activity and ecotoxicity are expressed as the mean and percentage (%) for lethality tests on snails (\u003cem\u003eB. glabrata\u003c/em\u003e), guppy fish (\u003cem\u003eP. reticulata\u003c/em\u003e) and brine shrimp (\u003cem\u003eA. salina\u003c/em\u003e). Comparisons between mortality rates were performed through analysis of variance (ANOVA) with a significance level of α\u0026thinsp;=\u0026thinsp;0.05 to observe whether there was any difference between the fresh latices analyzed and the concentration. The estimated lethal concentration to kill 50% of the specimens (LC\u003csub\u003e50\u003c/sub\u003e) was calculated from statistical software using probit analysis.\u003c/p\u003e \u003c/div\u003e"},{"header":"3 Results","content":"\u003cp\u003eThe first phase of this study consisted of evaluating the molluscicide activity of fresh latex from exotic or native species known for its toxicity. In this preliminary approach, \u003cem\u003eB. glabrata\u003c/em\u003e snails were subjected to only three concentrations (100.0, 50.0, and 25.0 \u0026micro;g/mL) of fresh latices from fourteen Euphorbiaceae species to determine the most active latices for posterior LC\u003csub\u003e50\u003c/sub\u003e tests. Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e shows the mortality rates of the snails in this initial evaluation. The data revealed differences in the efficacy of the molluscicidal activities of the fourteen fresh latices species evaluated (\u003cem\u003eF\u003c/em\u003e\u0026thinsp;=\u0026thinsp;12.91, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). However, no significant difference between the concentrations was observed (\u003cem\u003eF\u003c/em\u003e\u0026thinsp;=\u0026thinsp;2.17, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003ePreliminary evaluation of molluscicide activity of fresh latices from several exotic species of Euphorbiaceae against \u003cem\u003eBiomphalaria glabrata\u003c/em\u003e snails after 24 h of exposure and recovery.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eLatex\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"3\" nameend=\"c4\" namest=\"c2\"\u003e \u003cp\u003e% lethality\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003e100.0 \u0026micro;g/mL\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e50.0 \u0026micro;g/mL\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e25.0 \u0026micro;g/mL\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia cotinifolia\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia pulcherrima\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia tirucallii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia tithymaloides\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia umbellata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eJatropha curcas\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eJatropha elliptica\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eJatropha gossypiifolia\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eJatropha multifida\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eJatropha podagrica\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eManihot utilissima\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6.7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eRicinus communis\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003ePositive control: niclosamide (0.5 \u0026micro;g/mL).\u003c/p\u003e \u003cp\u003eIn this preliminary phase, \u003cem\u003eB. glabrata\u003c/em\u003e snails were strongly affected when exposed to fresh latices from \u003cem\u003eEuphorbia milii\u003c/em\u003e (var. \u003cem\u003ebreonii\u003c/em\u003e and var. \u003cem\u003emilii\u003c/em\u003e) and \u003cem\u003eE. umbellata\u003c/em\u003e species. Due to their high molluscicide potential, these species were reevaluated at lower concentrations to determine the LC\u003csub\u003e50\u003c/sub\u003e value. Fresh latices from \u003cem\u003eE. tirucallii\u003c/em\u003e and \u003cem\u003eManihot utilissima\u003c/em\u003e were also active but only at the highest tested concentration (100 \u0026micro;g/mL). The other species did not exhibit activity at the highest tested concentration.\u003c/p\u003e \u003cp\u003eAccording to the recommendations of the WHO (1983), only aqueous or alcoholic extracts of vegetal materials that cause the death of 90% of the malacological population at concentrations lower than 20 \u0026micro;g/mL are considered potentially active and viable for field assays, while less active extracts may very well provide sources of new lead compounds with molluscicidal activities. Considering the observed results for \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.9 \u0026micro;g/mL), \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.1 \u0026micro;g/mL) and \u003cem\u003eE. umbellata\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.1 \u0026micro;g/mL), their fresh latices can be classified as potential molluscicide agents (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). No significant difference between these potent fresh latices was detected (\u003cem\u003eF\u003c/em\u003e\u0026thinsp;=\u0026thinsp;3.10, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05); however, the lethality data as a function of concentration were significantly different (\u003cem\u003eF\u003c/em\u003e\u0026thinsp;=\u0026thinsp;10.44, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComplementary tests of the molluscicide activity of fresh latices from \u003cem\u003eEuphorbia milii\u003c/em\u003e and \u003cem\u003eSynadenium grantii\u003c/em\u003e against adult \u003cem\u003eBiomphalaria glabrata\u003c/em\u003e snails after 24 h of exposure and recovery.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eLatex\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c5\" namest=\"c2\"\u003e \u003cp\u003e% lethality\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eLC\u003csub\u003e50\u003c/sub\u003e\u003c/p\u003e \u003cp\u003e(\u0026micro;g/mL)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003e25.0 \u0026micro;g/mL\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5.0 \u0026micro;g/mL\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.0 \u0026micro;g/mL\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.5 \u0026micro;g/mL\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e90\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.9\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e90\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e50\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e1.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eSynadenium grantii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e9.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003ePositive control: niclosamide (0.5 \u0026micro;g/mL).\u003c/p\u003e \u003cp\u003eEcotoxicity tests were carried out to evaluate the toxicity of the fresh latices from \u003cem\u003eE. milii\u003c/em\u003e (var. \u003cem\u003ebreonii\u003c/em\u003e and var. \u003cem\u003emilii\u003c/em\u003e) and \u003cem\u003eE. umbellata\u003c/em\u003e plants against nontarget organisms such as fish and small crustaceans (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). Guppy fish (\u003cem\u003eP. reticulata\u003c/em\u003e) was used as the nontarget organism in this study because it is sensitive to contaminants and is easily found in rivers and lagoons in Brazil, providing results with great practical significance. \u003cem\u003eA. salina\u003c/em\u003e, commonly known as brine shrimp, is a small crustacean that has been the subject of many physiological studies. The brine shrimp lethality assay is one of the most practical tools for the preliminary assessment of general toxicity (McLaughlin et al., \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e1991\u003c/span\u003e); moreover, it is a good option for ecotoxicity assays because it is a sensitive nontarget organism and is usually utilized in biomonitoring or detection of toxic or bioactive compounds in vegetal extracts.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eEvaluation of the ecotoxic effects of different concentrations of fresh lattices from \u003cem\u003eS. grantii\u003c/em\u003e and \u003cem\u003eE. milii\u003c/em\u003e L. on guppy fish (\u003cem\u003ePoecilia reticulata\u003c/em\u003e) and brine shrimp (\u003cem\u003eArtemia salina\u003c/em\u003e) after 24 h of exposure and recovery.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eLatex\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eConcentration\u003c/p\u003e \u003cp\u003e(\u0026micro;g/mL)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c4\" namest=\"c3\"\u003e \u003cp\u003e% lethality\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cem\u003eP. reticulata\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cem\u003eA. salina\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e100.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e25.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003eLC\u003c/b\u003e\u003csub\u003e\u003cb\u003e50\u003c/b\u003e\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e9.9 \u0026micro;g/mL\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026gt;\u0026thinsp;100 \u0026micro;g/mL\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003e\u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e100.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e25.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003eLC\u003c/b\u003e\u003csub\u003e\u003cb\u003e50\u003c/b\u003e\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e9.9 \u0026micro;g/mL\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026gt;\u0026thinsp;100 \u0026micro;g/mL\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003e\u003cem\u003eSynadenium grantii\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e100.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e25.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003eLC\u003c/b\u003e\u003csub\u003e\u003cb\u003e50\u003c/b\u003e\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003e9.9 \u0026micro;g/mL\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e\u0026gt;\u0026thinsp;100 \u0026micro;g/mL\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cem\u003eE. umbellata\u003c/em\u003e and both varieties of \u003cem\u003eE. milii\u003c/em\u003e (\u003cem\u003ebreonii\u003c/em\u003e and \u003cem\u003emilii\u003c/em\u003e) were completely lethal to guppy fish at a concentration of 25.0 \u0026micro;g/mL, with an LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.9 \u0026micro;g/mL (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). These fresh latices did not show toxicity against \u003cem\u003eA. salina\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026gt;\u0026thinsp;100 \u0026micro;g/mL). No statistically significant difference was found (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05).\u003c/p\u003e"},{"header":"4 Discussion","content":"\u003cp\u003eThe control of snail vectors is an alternative way to prevent schistosomiasis. This strategy is particularly relevant in endemic localities in rural and urban areas, where the lack of sanitation and adequate housing show little potential for improvement in the future (Augusto \u0026amp; Silva, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Efforts are being made to discover molluscicide products of natural origin that are potentially biodegradable to address the growing awareness of environmental pollution. In this sense, this work focused its attention on the search for plant-based molluscicides obtained by simple, inexpensive, and efficient extraction and application techniques that are capable of being used by needy communities affected by schistosomiasis, opening possibilities for sustainable improvements in the health of these communities. Generally, latex is an aqueous emulsion found in the vacuole of secretory cells and is composed of lipids, resins, sugars, proteins, and enzymes. It can be easily obtained and manipulated for the preparation of molluscicides, dispensing the use of solvents and equipment for the preparation of extracts or extraction of active components. The use of latex from locally grown species by communities associated with health education programmes can help keep schistosomiasis at low levels since eradication is not considered realistic under current conditions in some developing countries. Among the fourteen Euphorbiaceae species evaluated in this study, only the fresh latices of three species were potentially active as molluscicides: \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e, \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e, and \u003cem\u003eE. umbellata\u003c/em\u003e (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eOriginally from Madagascar, \u003cem\u003eEuphorbia milii\u003c/em\u003e Des Moulin\u0026rsquo;s species is known in Brazil, where some varieties are cultivated as ornamental plants or hedges. In the present study, fresh latex solution of \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e had different molluscicidal effects than did the crude extract of leaves of this same species, which did not show molluscicidal activity in a previous study (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026gt;\u0026thinsp;100 \u0026micro;g/mL) (Hartmann et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). The latex at a concentration of 5.0 \u0026micro;g/mL exhibited 100% lethality in the first 24 h of the experiment. After 48 h, the lethal concentration for 50% of the exposed population was calculated to be 0.9 \u0026micro;g/mL. This difference in activity between fresh latex and the crude extract (Hartmann et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2011\u003c/span\u003e) is probably due to the concentration of substances responsible for molluscicidal activity. The concentration of the active substance in the leaf extract was likely low, or the substance could have degraded or polymerized during extract preparation. Curiously, the results observed for the \u003cem\u003ebreonii\u003c/em\u003e variety were very close to those obtained for the \u003cem\u003emilii\u003c/em\u003e variety (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.1 \u0026micro;g/mL). These concentrations are much lower than the WHO (1983) recommended concentrations for molluscicide activity. It was also noted that the calculated LC\u003csub\u003e50\u003c/sub\u003e value of the latex was very close to that reported in the literature for some species and varieties known as Christ's crown or crown-of-thorns (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.12 to LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;2.0 \u0026micro;g/mL) (Vasconcelos \u0026amp; Schall, \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e1986\u003c/span\u003e; Baptista et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e1994\u003c/span\u003e; Carvalho et al., 1998; Oliveira-Filho \u0026amp; Paumgartten, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Co\u0026ecirc;lho et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). \u003cem\u003eE. milii\u003c/em\u003e Des Moul. Ex Boiss var. \u003cem\u003ehislopii\u003c/em\u003e (syn. \u003cem\u003eE. splendens\u003c/em\u003e) lyophilized latex is one of the most potent molluscicide plants tested in outlying areas. Zani et al. (\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e1993\u003c/span\u003e) isolated eight categories of ingenol esters (miliamines) from \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003ehislopii\u003c/em\u003e and tested their molluscicide activities on \u003cem\u003eB. glabrata\u003c/em\u003e, with miliamine L being the most efficient, with lethality occurring at concentrations as low as 1.0 ppb. The latex of this variety was also very toxic to other Planorbidae snails (Oliveira-Filho \u0026amp; Paumgartten, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2000\u003c/span\u003e), suggesting that fresh latex from \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e and var. \u003cem\u003ebreoni\u003c/em\u003e could also be good molluscicides against other snail species.\u003c/p\u003e \u003cp\u003eAnother relevant aspect of extract concentrations is concern about other components of aquatic biota. Even though natural molluscicides are biodegradable at specific concentrations, these extracts may have risks even within the values demanded by the WHO. Therefore, toxicity tests showed that the use of latices from \u003cem\u003eE. milii\u003c/em\u003e (var. \u003cem\u003ebreonii\u003c/em\u003e and var. \u003cem\u003emilii\u003c/em\u003e) as molluscicide agents would be favorable because both species are lethal to snails at lower concentrations (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;~\u0026thinsp;1.0 \u0026micro;g/mL) than are the ones that are lethal to guppy fish (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.9 \u0026micro;g/mL) and \u003cem\u003eA. salina\u003c/em\u003e microcrustaceans (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026gt;\u0026thinsp;100.0 \u0026micro;g/mL). Therefore, these latices can be considered atoxic to the environment at molluscicide concentrations (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;~\u0026thinsp;1.0 \u0026micro;g/mL). These data corroborated those of Oliveira-Filho and Paumgartten (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2000\u003c/span\u003e), who analyzed the ecotoxicity of lyophilized latex from \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003ehislopii\u003c/em\u003e.\u003c/p\u003e \u003cp\u003e \u003cem\u003eE. umbellata\u003c/em\u003e (syn. \u003cem\u003eSynadenium grantii\u003c/em\u003e) is an African species cultivated as an exotic ornamental plant in Brazil where it is considered highly toxic. The population utilizes latex to reduce warts, treat gastric diseases, and even treat cancer (Kinghorn, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e1980\u003c/span\u003e). According to the literature, the chemical composition of latex from this species includes derivatives of phorbol esters (Kinghorn, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e1980\u003c/span\u003e; Bagavathi et al., \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e1988\u003c/span\u003e), triterpenoids (Uzabakiliho et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e1987\u003c/span\u003e), proteolytic enzymes (Menon et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2002\u003c/span\u003e), carboxylesterase enzymes (Govindappa et al., \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e1987\u003c/span\u003e), acetylcholinesterase isoenzymes (Govindappa et al., \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e1987\u003c/span\u003e) and glycoproteins (Rajesh et al., \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). During the evaluation of the molluscicidal activity of fresh latex from this species, it was found to be potent (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). As previously mentioned for \u003cem\u003eE. milli\u003c/em\u003e var. \u003cem\u003ebreoni\u003c/em\u003e, the crude alcoholic extract of \u003cem\u003eE. umbellata\u003c/em\u003e leaves is less active (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;40.0 \u0026micro;g/mL; Hartmann et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2011\u003c/span\u003e) than fresh latex is. Regarding ecotoxicity, in the case of fresh \u003cem\u003eE. umbellata\u003c/em\u003e latex, the lethal concentration for \u003cem\u003eB. glabrata\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.1 \u0026micro;g/mL) was close to that of guppy fish (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.9 \u0026micro;g/mL) (Tables\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e and \u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). Thus, although the latex from this species is considered a good molluscicide agent, its use requires ample care due to its toxicity to other organisms present in the aquatic environment at the same concentration, as latex is lethal to the vector snail of \u003cem\u003eSchistosoma mansoni.\u003c/em\u003e These data differed from those achieved by Pereira et al. (2017), who reported an LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.36 \u0026micro;g/mL for lyophilized latex. These authors also performed assays with \u003cem\u003eDanio regio\u003c/em\u003e (zebrafish) to investigate the toxicity of \u003cem\u003eE. umbellata\u003c/em\u003e latex to nontarget organisms, and they found that this latex was less toxic to zebrafish (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;7.22 \u0026micro;g/mL) than to \u003cem\u003eB. glabrata\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.36 \u0026micro;g/mL). The difference in activity observed between our results and those of the authors (Pereira et al., 2017) may be related to different factors: type of latex (fresh/lyophilized), latex harvesting period (autumn/spring), and geography (harvesting in the area close to the Tropic of Capricorn/harvesting in the equatorial area-coast).\u003c/p\u003e \u003cp\u003eAnother evaluated species in this study also presented latices with moderate molluscicidal activity. In Brazil, the African milkbush or \u003cem\u003eaveloz\u003c/em\u003e plant (\u003cem\u003eE. tirucallii\u003c/em\u003e), which is very caustic and popularly used for wart scaring, is known for its latex toxicity. As part of the screening of the molluscicide species, latex from this species was evaluated and found to have moderate activity against \u003cem\u003eB. glabrata\u003c/em\u003e, with an LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;89.4 \u0026micro;g/mL. Cassava (\u003cem\u003eManihot utilissima\u003c/em\u003e Pohl) is a native plant known for its great usefulness in the economy and history of Brazil. One of its varieties, \u0026ldquo;mandioca-brava\u0026rdquo;, is also known for poisoning cases in humans and animals. Considering the toxic character of your latex, its possible molluscicide activity was evaluated. Latex from leaf petioles of \u003cem\u003eM. utilissima\u003c/em\u003e also presented moderate molluscicidal activity (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;95.0 \u0026micro;g/mL).\u003c/p\u003e \u003cp\u003eLatex from Barbados nuts (\u003cem\u003eJatropha curcas\u003c/em\u003e) did not exhibit activity at the highest tested concentration (100 \u0026micro;g/mL; Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) in the present study. Other authors have observed significant molluscicidal activity of \u003cem\u003eJ. curcas\u003c/em\u003e against different vector snails (Liu et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e1997\u003c/span\u003e; Rug \u0026amp; Ruppel, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). According to Nath and Dutta (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e1991\u003c/span\u003e), latex from \u003cem\u003eJ. curcas\u003c/em\u003e has a protolithic enzyme called curcain. Another study indicated two significant activities for \u003cem\u003eJ. curcas\u003c/em\u003e latex: procoagulant and anticoagulant (Osoniyi \u0026amp; Onajobi, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). Therefore, even if latex contains interesting substances related to other activities, this does not indicate who is a good molluscicide. Latices from another species of this genus were evaluated in this study (\u003cem\u003eJ. elliptica, J. gossypifolia\u003c/em\u003e L, \u003cem\u003eJ. multifida\u003c/em\u003e L, and \u003cem\u003eJ. podagrica\u003c/em\u003e); however, none of them presented molluscicidal activity at the highest tested concentration (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e"},{"header":"5 Conclusions","content":"\u003cp\u003eAmong the several Euphorbiaceae species evaluated in this study, only \u003cem\u003eE. milii\u003c/em\u003e (var. \u003cem\u003ebreonii\u003c/em\u003e and var. \u003cem\u003emilii\u003c/em\u003e) and \u003cem\u003eE. umbellata\u003c/em\u003e presented potent molluscicidal effects on \u003cem\u003eB. glabrata\u003c/em\u003e snails. The use of fresh latices from \u003cem\u003eE. milii\u003c/em\u003e varieties represents a feasible alternative strategy for schistosomiasis control because these plants can be efficiently grown in endemic areas (not requiring soil care, fertilizers, or water); additionally, these plants yield a relatively large amount of latex throughout the year, and the latex can be easily collected. In summary, these fresh latices are active at low concentrations, are biodegradable and inexpensive, and are obtained from renewable resources. The preparation of these molluscicides is straightforward, does not require organic solvents (typically used to prepare extracts) or equipment (rotary evaporator or freeze dryer), and can be easily prepared by the population most affected by schistosomiasis. The molluscicidal activity of fresh latex from \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003ebreoni\u003c/em\u003e and \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e latices is reported here for the first time. On the other hand, the fresh latex from \u003cem\u003eE. umbellata\u003c/em\u003e was strongly lethal to \u003cem\u003eB. glabrata\u003c/em\u003e snails (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.1 \u0026micro;g/mL) and guppy fishes (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.9 \u0026micro;g/mL). Consequently, their use as molluscicides is contraindicated because they require special attention due to their possible toxicity against nontarget organisms.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eR. A. M. thanks Diretoria Executiva de Gest\u0026atilde;o da Pesquisa e da P\u0026oacute;s-gradua\u0026ccedil;\u0026atilde;o - DEGPP (Executive Board of Research and Postgraduate Management)/UNIPAR for scholarship. The authors thank Dr. Felipe A. La Porta (UTFPR) for assistance with the statistical analyses.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eD. B. H., Z. C. G. and A. L. J. contributed to the conception and design of the experiments. E. J.\u0026nbsp;contributed to the collection and identification of the botanical material\u003cem\u003e.\u0026nbsp;\u003c/em\u003eD. B. H., R. A. M., G. Z., I. A. S., and E. J. contributed to the data acquisition. D. B. H., R. A. M., and A. L. J. contributed to the analysis and interpretation of the data. A. L. J. supervised the project and wrote the manuscript with input from all the authors. All the authors read and agreed to the publication of this version of the article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis project was financially supported by Paranaense University (UNIPAR - Proj: 15950 and 18004).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no conflicts of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eDoes not apply.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eInformed consent\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eDoes not apply.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAlves, T. M. D., Silva, A. F., Brand\u0026atilde;o, M., Grandi, T. S. M., Sm\u0026acirc;nia, E. F. A., Sm\u0026acirc;nia, A., \u0026amp; Zani, C. L. (2000). 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(1999). Screening of Asteraceae (Compositae) plant extrats for molluscicidal activity. \u003cem\u003eMem\u0026oacute;rias do Instituto Oswaldo Cruz\u003c/em\u003e,\u003cem\u003e 94\u003c/em\u003e(3), 411-412. https://doi.org/10.1590/S0074-02761999000300023\u003c/li\u003e\n\u003cli\u003eMenon, M., Vithayathil, P. J., Raju, S. M, \u0026amp; Ramadoss, C. S. (2002). Isolation and characterization of proteolytic enzymes from the latex of \u003cem\u003eSynadenium grantii\u003c/em\u003e Hook, \u0026apos;f\u0026apos;. \u003cem\u003ePlant Science\u003c/em\u003e, \u003cem\u003e163\u003c/em\u003e(1), 131-139. https://doi.org/10.1016/S0168-9452(02)00085-7\u003c/li\u003e\n\u003cli\u003e\u003cem\u003eNath, \u003c/em\u003eL. K., \u0026amp; \u003cem\u003eDutta, \u003c/em\u003eS. K. (1991). Extraction and purification of curcain, a protease from the latex of \u003cem\u003eJatropha \u003c/em\u003e\u003cem\u003ecurcas\u003c/em\u003e Linn. \u003cem\u003eJournal of Pharmacy and Pharmacology\u003c/em\u003e, \u003cem\u003e43\u003c/em\u003e(2), 111-114. https://doi.org/10.1111/j.2042-7158.1991.tb06642.x\u003c/li\u003e\n\u003cli\u003eOliveira-Filho, E. C., \u0026amp; Paumgartten, F. J. R. (2000). Toxicity of \u003cem\u003eEuphorbia milli\u003c/em\u003e latex and Niclosamide to snails and non-target aquatic species. \u003cem\u003eEcotoxicology and Environmental Safety\u003c/em\u003e, \u003cem\u003e46\u003c/em\u003e(3), 342-350. https://doi.org/10.1006/eesa.2000.1924\u003c/li\u003e\n\u003cli\u003eOsoniyi, O., \u0026amp; Onajobi, F. (2003). Coagulant and anticoagulant activities in \u003cem\u003eJatropha curcas\u003c/em\u003e latex. \u003cem\u003eJournal of Ethnopharmacology\u003c/em\u003e,\u003cem\u003e 89\u003c/em\u003e(1), 101-105. https://doi.org/10.1016/s0378-8741(03)00263-0 \u003c/li\u003e\n\u003cli\u003eRajesh, R., Nataraju, A., Gowda, C. D. R., Frey, B. M., Frey, F. J., \u0026amp; Vishwanath, B. S. (2006). Purification and characterization of a 34-kDa, heat stable glycoprotein from \u003cem\u003eSynadenium grantii\u003c/em\u003e latex: action on human fibrinogen and fibrin clot. \u003cem\u003eBiochimie\u003c/em\u003e, \u003cem\u003e88\u003c/em\u003e(10), 1313-1322. https://doi.org/10.1016/j.biochi.2006.06.007\u003c/li\u003e\n\u003cli\u003eRug, M., \u0026amp; Ruppel, A. (2000). Toxic activities of the plant \u003cem\u003eJatropha curcas\u003c/em\u003e against intermediate snail hosts and larvae of schistosomes. \u003cem\u003eTropical Medicine and International Health\u003c/em\u003e,\u003cem\u003e 5\u003c/em\u003e(6), 423-430. https://doi.org/10.1046/j.1365-3156.2000.00573.x\u003c/li\u003e\n\u003cli\u003eSilva, N. F. S., Cogo, J., Wiepieski, C. C. P., \u0026amp; Laverde Junior, A. (2008). Bioensaio de atividade moluscicida adaptado \u0026agrave; avalia\u0026ccedil;\u0026atilde;o de extratos de plantas medicinais. \u003cem\u003eArquivos de Ci\u0026ecirc;ncias Veterin\u0026aacute;rias e Zool\u0026oacute;gicas da UNIPAR\u003c/em\u003e, \u003cem\u003e11\u003c/em\u003e(2), 179-181. Available at: https://revistas.unipar.br/index.php/veterinaria/article/view/2575 \u003c/li\u003e\n\u003cli\u003eUzabakiliho, B., Largeau, C., \u0026amp; Casadevall, E. (1987). 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Available at: https://apps.who.int/iris/handle/10665/262863\u003c/li\u003e\n\u003cli\u003eWorld Health Organization (WHO) (1983). Reports of the scientific working group on plant molluscicides. \u003cem\u003eBulletin of the World Health Organization\u003c/em\u003e, \u003cem\u003e61\u003c/em\u003e, 927. Available at: https://apps.who.int/iris/handle/10665/60086\u003c/li\u003e\n\u003cli\u003eWorld Health Organization (WHO) (2023, February 1). Fact sheet N\u0026ordm;115: Schistosomiasis, Geneva, World Health Organization. Retrieved February 15, 2024, from: http://www.who.int/mediacentre/factsheets/fs115/en/\u003c/li\u003e\n\u003cli\u003eZani, C. L., Marston, A., Hamburger, M., \u0026amp; Hostettmann, K. (1993). Molluscicidal milliamines from \u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003ehislopii\u003c/em\u003e. \u003cem\u003ePhytochemistry\u003c/em\u003e, \u003cem\u003e34\u003c/em\u003e(1), 89-95. https://doi.org/10.1016/S0031-9422(00)90788-X\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[{"identity":"c117b115-91e8-4faa-a226-282d35a14df8","identifier":"10.13039/501100004842","name":"Universidade Paranaense","awardNumber":"UNIPAR/DEGPP-15950 and UNIPAR/DEGPP-18004","order_by":0}],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Federal University of Technology – Paraná","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Christ's crown, ecotoxicity, Euphorbiaceae, helminthiasis, plant molluscicide, snail control","lastPublishedDoi":"10.21203/rs.3.rs-4048314/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4048314/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eSchistosomiasis is a millennial disease that affects approximately 235\u0026nbsp;million people. Considering that most of the population affected by schistosomiasis faces different conditions of poverty, finding compounds from plants that can help control or eradicate this disease would be worthwhile especially if the plants are readily available for those who need to use them the most. This study aimed to evaluate the effects of fresh latex from fourteen locally cultivated Euphorbiaceae species on the freshwater snail \u003cem\u003eBiomphalaria glabrata\u003c/em\u003e Say, 1818 (Gastropoda: Planorbidae), the principal vector of the \u003cem\u003eSchistosoma mansoni\u003c/em\u003e trematode on the American continent. Adult \u003cem\u003eB. glabrata\u003c/em\u003e snails were exposed to various concentrations of fresh latex from Euphorbiaceae species for 24 and 48 hours. Mortality data were analyzed using the probit method. To evaluate the latex ecotoxicity of the promising species against other aquatic organisms, lethality tests were performed on \u003cem\u003ePoecilia reticulata\u003c/em\u003e Peter, 1859 (Cyprinodontiformes: Poeciliidae) fish and \u003cem\u003eArtemia salina\u003c/em\u003e Leach (Anostraca: Artemiidae) nauplii. Three species presented significant molluscicide activity: \u003cem\u003eEuphorbia milii\u003c/em\u003e var. \u003cem\u003ebreonii\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.9 \u0026micro;g/mL), \u003cem\u003eE. milii\u003c/em\u003e var. \u003cem\u003emilii\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.1 \u0026micro;g/mL) and \u003cem\u003eE. umbellata\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;9.1 \u0026micro;g/mL). Only \u003cem\u003eE. milii\u003c/em\u003e (var. \u003cem\u003ebreonii\u003c/em\u003e and var. \u003cem\u003emilii\u003c/em\u003e) was not lethal to the evaluated fish (LC\u003csub\u003e50\u003c/sub\u003e of 9.9 \u0026micro;g/mL) and \u003cem\u003eA. salina\u003c/em\u003e (LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026gt;\u0026thinsp;100.0 \u0026micro;g/mL). These results qualify both the \u003cem\u003eE. milii\u003c/em\u003e variety of fresh latices as versatile (active at low concentrations, biodegradable, inexpensive, and obtained from renewable resources) and alternative molluscicidal agents against \u003cem\u003eB. glabrata\u003c/em\u003e snails.\u003c/p\u003e","manuscriptTitle":"Versatile natural molluscicides against the snail Biomphalaria glabrata Say, 1818 (Pulmonata: Planorbidae), the principal vector of Schistosoma mansoni on the American continent","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-03-12 06:15:03","doi":"10.21203/rs.3.rs-4048314/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"28831863-1f5d-4f39-b461-177796d046e0","owner":[],"postedDate":"March 12th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":29327471,"name":"Tropical Medicine"},{"id":29327472,"name":"Chemical Biology"},{"id":29327473,"name":"Health Policy"}],"tags":[],"updatedAt":"2024-03-12T06:15:03+00:00","versionOfRecord":[],"versionCreatedAt":"2024-03-12 06:15:03","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4048314","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4048314","identity":"rs-4048314","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}