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Control of the Glassy Clover Snails Monacha Cartusiana Using Zingiber Officinale Extract As an Ecofriendly Molluscicide

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Control of the Glassy Clover Snails Monacha Cartusiana Using Zingiber Officinale Extract As an Ecofriendly Molluscicide African J. Biol. Sci., 15 (1): 101-115 (2019) ISSN 1687-4870 e- ISSN 2314-5501 (online) www.aasd.byethost13.com e.mail: [email protected] Control of the glassy clover snails Monacha cartusiana using Zingiber officinale extract as an ecofriendly molluscicide Mahmoud Abd El-Atti1*, Elsheakh A. A.2 , Khalil A.M.1 and Wesam S. Elgohary2 1- Zoology Department, Faculty of Science, Zagazig University, Zagazig. Egypt 2- Plant Protection Research Institute, Agr. Res. Center. Doki, Giza, Egypt *Corresponding author email: [email protected] ABSTRACT High population densities of Monacha cartusiana were observed to cause perdition of various economic crops at Sharkia Governorate. The present study was carried out control this harmful pest using ethanolic Ginger extract as a natural and environmentally safe molluscicide.Toxicological tests revealed that low concentration (20%) of ethanolic Ginger extract caused 10 % mortalityof cloversnails after one day of exposure and 66.7% mortalityafter28 days. The highest mortality percentage (90 %) was recorded after 28 days of treatment with40% Ginger extract. Biochemical investigations showed thatLC25 (7.04%) Ginger extract elevated ALT, AST, α & β esterases andphenoloxidase levels of treated snails compared to control.Histological inspections of the digestive gland and ovotestis of snails exposed to LC25Ginger extract revealed that the digestive tubules showedvarious tubularanomalies, inflammatory hemocytes infiltration, excessive luminal secretionsand appearanceof necrotic areas. Acini of ovotestis weredeformedand comprised misshapen eggs and distorted sperms.TEM investigations showed cellular malformations in the digestive gland of treated snails including; severely disrupted microvilli, ruptured cell membranes, fractured RER and mitochondrial pyknosis. Key words: Ecofreindly- Antioxidant Enzymes- Ginger extract-Histopathology- Monacha cartusiana-TEM. INTRODUCTION 2019). High population densities were Land snails are considered as serious discoveredinfesting various economic crops agricultural pests Egypt. They are at Sharkia Governorate, Egypt (Lokma, herbivorous animals that devour mellow 2013). M. cartusiana reported to act also as parts of vegetables, fruit trees and an intermediate host for sheep lungworm ornamental plants (Godan, 1983) and ruin and lancet liver fluke (Godan, 1983). mushy leaves, fruits, seeds and tubers (El- Chemical control applications still the most Okda, 1984). Undesirable smell and savor commonly used for controlling land caused by mucous secretions left on plants snailsalthough; they extensively prejudice during their movements, averts human and non-target organisms including man (Gabr other animals from feedingon these et al., 2006). The high costs and resistance contaminated plants (El-Deeb et al., 1999). exerted against synthetic pesticides The glassy clover snails (Monach encourage the usage of naturally derived cartusiana) were recorded extensively at pest control strategies (Massoud and Habib vast geographic areas at the Mediterranean 2003). Recently, several countries enhanced regions and Southeastern Europe the use of plant extracts in pest control due (Pieńkowska, 2018: Pieńkowska et al., to their low mammalian toxicity, low costs 102 Mahmoud Abd El-Atti et al. and fast biodegradability (Singh et al., 3. Experimental design 2000). Ginger extracts possess anti- Clover snails were divided randomly into inflammatory, analgesic, antioxidants and three main groups; the first group used for antimicrobial activities (Sharma et al., 2013; toxicological studies was subdivided into five Nikoli et al., 2014; Amri and Touil- sub groups and treated with different Boukoffa, 2016). Few studies have been concentrations of Ginger extract for 28 days. carried out on the molluscicidal activities of Four replicates were applied for each Ginger (Ahmad et al., 2013; and Edwin and subgroup.The second groupfed with baits Jacob 2017). Physiological and containing LC25 of ginger extract for 14 days histopathological investigations give andused for biochemical, histopathological and excellent indication about the hazards occurs ultrastructural investigations.The third group left within the organism because of exposure to as control group. toxicants. The present study aimed to control the glassy clover snail Monacha 4. Biochemical measurements: cartusiana using Ginger extract as a natural Digestive glands were dissected out from and alternative solution instead of using both control and treated snailsthen homogenized harmful chemical molluscicides. in distilled water using a Teflon homogenizer. The homogenates were centrifuged at 8000 MATERIALS AND METHODS r.p.m. for 15 minutes at 5ºC inrefrigeratedcentrifuge. Deposits were 1. Snails collection and acclimation discarded and supernatants were kept in Adult snails of M. cartusiana were a deep freezer untiluse. Activities of AST collected from clover fields at Malames and ALT enzymes were determined according to village, Meniet El-kamh district, Sharkia the procedure of Reitman and Frankle (1957). Governorate during spring 2018. Snailswere Alpha and Beta esterases assayed according to transferred to the laboratory and kept in Van Asperen, (1962). Phenoloxidase glass cages (50 x 30 x 30 cm) contained activitymeasured using the method of Ishaaya moist soil and fed daily with fresh Lettuce (1971).All biochemical measurements were for two weeks. carried out in Plant Protection Research Institute, Agriculture Research Centre, Giza, Egypt. 2. Ginger extracts preparation Rhizomes of Ginger were purchased 5. Histopathological studies from the InternationalCompany (Cairo- For light microscopy, digestive glands of Egypt). Rhizomeswere ground intoa fine both control as well as treated with LC25 Ginger powder using a pestle and mortar. The extract for 14 days were dissected out and fixed powder (30 g) was soaked in ethanol (600 in Bouin's fluid. Specimens dehydrated in a ml) in a Sechelt apparatus for twodays. graded series ofethanol, cleared in Xylene for 20 Ethanol was evaporated under minutes, impregnated in paraffin wax (three reducedpressure until brown extract was changes) at 60oC for 2 hours and embedded in obtained (Bakry et al., 2013). Poisonous paraffin wax. Sections (4-5 µm thick) were cut baits were formulated by mixing ethanolic by microtome, mounted on glass slides Ginger extract with five grams of sugarcane andstained with hematoxylin and eosin. syrup, completed with wheat bran to 100 grams and moisten with small amounts of 6. Ultrastructural studies water. Dissected digestive glands of both control and Ginger treated snails were fixed in 2.5 % glutaraldehyde in 0.05 M cocodylate 103 Control of the glassy clover snails Monacha cartusiana using Zingiber officinale extract as an ecofriendly molluscicide buffer containing 0.15 sucrose at pH 7 for RESULTS two hours. Specimens were fixed in 1% 1. Toxicity tests Osmium tetroxide at 4oC for an hour, Figure (1) showed that snail dehydrated in an ascending series of ethanol mortalities increased with increasing of both and embedded in Araldite f Epon. Blocks Ginger extract concentrations and were cut with diamond ultramicrotome exposuretime. At the first day of exposure, knifes. Semithin sections stained with 1% all treatments caused low mortalities. Toluidine blue. Ultra-thin sections stained Ethanolic extract of Ginger (20%) causes with aqueous Urinyl acetate and Lead citrate 10, 50, 56.6, 66 and 66.7% mortalitiesafter and examined under JEOL 100 CX TEM at 1, 7, 14, 21 and 28 days of treatment the Electron Microscopy Unit, Faculty of respectively.Exposure to 30 % Ginger Agriculture, El-Mansoura University, Egypt. extract caused 6.6, 36.7, 70, 76.6 and 83.3% for the same days of 7. Statistical analysis: exposurerespectively.Finally, 40% Ginger Values of toxicological and extract resulted in 10, 46.6, 76.6, 83.3 and biochemical studies were expressed as a 90% respectively, at the same time intervals. mean ± SE. The obtained data were analyzed The highest percentage of mortalities (90%) statistically for the significance of differences occurred after treatment with40% Ginger using student’s t-test (Goldstein, 1964). extract for 28 days (Fig. 1). Fig.(1): Mortality percentages of M. cartusiana exposed to different concentrations of Ginger extract for 28 days. 2. Biochemical measurements their levels slightly decreased with 2.1. Activities of ALT and AST increasing time of exposure, although levels Levels of AST were significantly were much higher than control (Fig. 2). elevated (P <0.05) after exposure to LC25 (7.04%) of Z. officinale extract and the 2.2. Activities of α and β-esterases highest level was recorded after two weeks LC25 of Z. officinale extract caused a of exposure compared to control. Activities highly significant increase (P <0.05) of α of ALT were significantly increased(P < and β-esterases activities after the first day 0.05 ) after the first day of treatment, while of exposure. Values of both enzymes were 104 Mahmoud Abd El-Atti et al. decreased insignificantly (P>0.05) after the following the first day of treatment. 7th and 14th day of treatment (Fig. 2). Phenoloxidase levels decreased insignificantly (P>0.05) after one week of 2.3. Activity of Phenoloxidase treatment. The lowest level of phenoloxidase Highly significant increase in the was achieved after two weeks of exposure to activity of Phenoloxidase was detected Ginger extract (Fig. 2). Fig. (2): Enzymatic activities of M. cartusiana treated with LC25 Ginger extract for 14 days. 3. Microscopical Examination
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