A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

(1)Dr.Marwah Hashim Dawood ,(2) Dr.Sajeda Y. Shareef ,(3) Dr.Saly Ahmad Thiyab

(1) Dept. of Clinical Laboratory Science, College of Pharmacy , (2)(3) Dept. of Biology , College of Science (1)(2)(3) Mosul University

Abstract

Two intestinal cestodes were chosen to make a compares, considering some aspects of energy metabolism; Ophiotaenia bofonis habitat small intestine of the house frog Bufo viridis viridis, and Cotugnea columbae habitat small intestine of the domestic pigeon Columba livia domestica. The present study reviled that concentration of macrobiomolecules , the energy raw materials, in O. bofonis is higher than that of C. columbae. There were significant differences at p≤0.05 between the concentration of total carbohydrates, protiens and lipides: (240, 181 µg/ml and 196mg/100ml, respectevely in the pigeon cestode and 105, 153 µg/ml and 133mg/100ml, respectively in the frog cestode). High performance liquid chromatography technique (HPLC) analysis for fatty acid elucidated that each of the two helminthes contain six types of fatty acid. O. bofonis found to have three saturated fatty acids: stearic, palmatic and myristic, 2.72%, 2.29% and 0.89% respectively and three unsaturated fatty acid: linoleic, linolienic and oliec, 73.13%, 13.07% and 7.91%, respectively. On the other hands, C. columbae found to have four saturated fatty acid: stearic, palmatic, myristic and caproic, 51.43%, 45.89%, 2.38% and 0.19% respectively, and two unsaturated fatty acides: linoleic, linolienic, 0.09% and 0.06%, respectively. Enzymatic study pointed to the significant differences at p≤0.05 between the enzymes activity in frog cestode and that of pigeon cestode. Activity of G6PDH and LDH (cytosolic enzymes) and SDH and FR (Mitochondrial respiretory enzymes), were :7.514 U/L, 56 IU/L, 130 nm FAD/min/ mg protien and 198 nm NADH/min/mg protien in O. bofonis extract; and 7.514 U/L, 56 IU/L, 130 nm FAD/min/ mg protien and 198 nm NADH/min/mg protien in C. columbae extract. The prevous resultes may reffered to a relativly higher activity of the enzymes under study, those take a part in glycolisis and energy metabolism libration in O. bofonis than that in C. columbia, in spite of that the occumulated souces of energy were more aboundant in pigeon cestode.

Tikrit Medical Journal 2017;Vol( 23 ) No( 1 ) :113-127

A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

Key words: energy metabolism, Ophiotaenia bofonis, Cotugnea columbae, carbohydrates, lipids, fatty acids, G-6-PD, LDH, SDH, FR.

Introduction

Parasites are in need for unusually great formed via malate dismutation, a demand on energy generating fermentative pathway which is present mechanisms. So, too, does the need to in all types of parasitic cestodes and produce vast numbers of offspring. nematodes (Maule and Marks, 2006) . Protection against the immune system Thus,the energy metabolism of of the host is another costely energy parasitic is clearly and drain (Bryant and Behm, 1989). substencially different from that of their There is no doubt that the variance in hosts. Apparently, they have adapted to parasites demand for energy is closely exploid anaerobic niches and the related to host type, habitat (location in surplus substrate that are commonly the host body), in addition to the present in their enveronment. This echosystem in which the host enable them to use low-effeciency themselves lieve (Chapple, 1980). catabolic pathways, and lose several Intestinal cestodes are a group of anabolic pathways, such as those endoparasitic helminthes. Their involved in the biosynthesis of fatty elongated body enables them to live in acids and steroles. Obveously, parasitic the tubular habitat (Smyth and flatworms adapted to an apportunistic McManus, 2007). Cestodes miss way of life (Bryant and Behm, 1989) alimentary canal, thus derive their The choosen cestodes of the present nutrition from the host`s gut across it`s study were Ophiotaenia bofonis; class: highly specialized, metabolically active (Eucestoda); order: body cerface or tegument (Smyth and ; family: McManus, 2007). The metabolic and . Subfamily: in vitro studies suggest that there is a Proteocephalinae. Ophiotania bofonis complex nutritional relationship occurs habitat small intestine of house frog between the gastrointestinal cestodes ( Bufo viridis viridis). The other cestod and it`s host (Sonune, 2012). was Cotugnia columbae; class: cestoda; The most important variation in the order: cyclophyllidea; family: enveronment arround the parasite is the davaineidae. Cotugngnia columbae avaibility of food and oxygen. As for habitat small intestine of domestic parasitic flatworms, they are generally pigeon (Columba livia) do not use oxygen as the final electron (Yamaguti,1959 and Jones & Bray, acceptor, but use fermentative 1994). processes to obtain most of their ATP. Researches about energy metabolism in Most adult flatworms inside the final Ophiotaenia spp. and Cotugnia spp. are host produce end products for the scarce. Some studies were fermentative carbohydrate breakdown, epidimeological (Silva , et al. ,2006; such as succinate, acetate, propanioate Begum and sehrin, 2012 and Ribeir, et and lactate.These end products are al. 2014), otheres were taxonomic

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A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

( Chambrier, et al., 2012; shukla, et al, Biology Department, College of 2012; Gamil, 2012 and Sanap and Science, University of Mosul, Siddiqui, 2013), the otheres were depending on Yamaguti (1959) ; biochemical (Al-Naftachi, 2006; Schmidt (1986); Jones and Bray Waghmare and Chavan, 2010; and (1994); Scholz and Chambrier (2003). Biswal, et al.,2012). Biochemical studies: Therefore, this research aimed to I-Preparation of worms extract: The compare betweenthe some aspects of worms were suspended in a 0.05M energy metabolism of Ophiotaenia Tris-HCl buffer (7.4) at a concentration bofonis as an amphebian intestinal of 10% wet wight/volume; then the cestode and Cotugngnia columbae as suspension homogenized in a tissue avian intestinal cestodes; trying to give grinder. Cell membranes disrupted some concepts about the effects of host using ultrasonic disintegration (MSE), a ecosystem variation on the energy 12000 vibration/second for 30 second metabolism of their intestinal cestodes. in ice-bath. Four cycles of sonication were applied with the suspention. Ultracentrifugation at Materials and Methods 15000g/30minutes was done using MSE superspeed ultracentrifuge. The 1-Specimen collection: The adult supernatent fraction was choosen for cestodes Ophiotaenia bofonis were the accessive biochemical studies (Al- collected from the upper part of the Naftachi, 2006). small intestine of Bufo viridis viridis (the house frog) , which were obtained II- Preparation of mitochondrial from different areas of Nenivah extract: As for the mitochondrial Governarate, whereas the adult enzymes, succinate dehydrogenase and cestodes Cotugnia columbae were fumarate reductase, the worms were collected from small intestine of extracted utilizing method adopted by Columba livia domestica (domestic Pumpori and Srivastava (1987). A pigeon) which were obtained from local modified mitochondrial medium (pH markets in Mosule City. The collection 7.4) described by Scheibel, et al. were made during 2011- 2012. The (1968) was used. The medium collected then bringed and consisted of: 0.24M sucrose, 0.15% dissected in the research lab/Biology bovine serum albumine (BSA) and Department/College of Science/ 0.00M ethylendiaminetetraacetic acid University of Mosul. (EDTA). Worm`s tissue were granded The collected cestodes were washed in a concentration of 10% (W/V), the several times with NaCl buffered seline homogenate then cenrifuged at 1000g 0.85%, put in petridish, examined under for 20 minute, the supernatent then Hamilton dissecting microscope. wet recentrifuged at 15000g for 30 miute. wieght for each sample was fixed. Then The precepetate which contain kept in a deep freeze at -18ºC for mitochondrial pellet was washed twice subseqent studies. Some helminthes with modified mitochondrial medium. were squized and fixed in hot formalin Washed pellet were suspended finally 10%, then dehydrated with gradiant in 0.05 M Tris-HCl buffer pH 7.4. The ethanol concentrations, clarified by mitochondrial fraction was desrupted lactophenol and mounted by DPX. by four successive barests 30 second Classification was performed in (with 1 minute cooling intervals) using

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A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

ultrasonic disentegrator, all the above saperating funnel after mixing with operations wre carried out at 4ºC. diethyl ether. Rotary vacum evaporater III-Estimation of macromolecules: was used to extract fatty acid mixture. A-Estimation of total carbohydrates: B-HPLC analysis for fatty acids: Herbert, et al. method (1971) was High performance liquid applied to estimate cestodes chromatographic technique (HPLC)- carbohydrates in the worms extract. reversable type was used to analyze Absorbance was mesured at 488nm. fatty acid mixture. Carbohydrate concentration were 1ml of acetyl chloride was added to the estimated depending on carbohydrate crude fatty acids that prepared before standered curve. (Al-Kaisy, et al. 1991). Fatty cid esters B-Estimation of total protiens: and authentic fatty acids were analysed Lowry, et al. colorometric method using the following conditions: (1951) was used to estimate protien C18 colum (4.6×250)mm, mobile concentration in the worms extract. phace: (Acetonitrile 60% +water 40%), Foline reagent react with protein to flow rate: 0.4 ml/minute, wave length: produce blue complex in alkaline 254nm (adopted by Elliot, et al. 1989 medium. Absorbance was mesured at and Al-mowla, 2010). Analysis of 750nm. Concentartion of protien was worms fatty acids and authentic fatty estimated using protein standered acids was performed in Ibn-Sena curve. Company Labs/Bagdad University. C-Estimation of total lipids: Chabrol V-Enzymes studies: and Chardonnet colorometric method A-Assay system for Glucose-6- (1937) was used to evaluate whole phosphate dehydrogenase (G6PDH) lipids content (cited by Tietz, 1980). 20 activity:G6PDH (EC1.1.1.49) is a µL of worms extract was heated with cytosolic enzyme that catalyze the first concentrated sulphoric acid. step in the pentose phosphate shunt, Phosphovaniline indicator was added to oxidizing G-6-P to 6-P-gluconate and the mixture to produce purple-red color reduce NADP to NADPH. The activity complex. Absorbance were determined of this enzyme is mesured using Tietz at 540nm. Whole lipids in each sample spectrophotometric modified method were estimated depending on the rule: (1995). Formation rate of NADPH is proportional to the G-6-P

activity,which is mesured IV-Fatty acids analysis: spectroscopically as an increase in A-Fatty acids extraction: Worms absorbance at 340nm. Unit of enzyme extract was lyophilized using Christ- activity in the worms extract was BETA-Liophilizer/Germany. Fatty estimated according to the following acids were extracted according to Al- rule:

Kaisy, et al. modified method (1991). G-6-PD(IU/L) = ×492 200mg of dried extract wre dissolved in B-Assay system for Lactate 10ml of 7.5N NaOH (which prepared in dehydrogenase (LDH) activity: LDH 60% methnol). The mixture then heated (EC1.1.1.27) activity were evaluated to 105ºC for 90minutes, after cooling, depending on a method adopted by 12ml of distilled water added, the wolf (1989). LDH reduce pyruvate to mixture then acidified using 20% lactate in the presence of NADH (co- sulphoric acid to adjustified pH to 2. factor). Fatty acid mixture withdrawn with

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A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

A specific analysis set, provided by registered from zero-time for 10 Biomerierux/France was used. minutes. Unit of enzyme activity was Estimation of LDH activity depends on expresed as the ammount of enzyme the reaction betweenthe residual required for the oxidation of 1µM of pyruvate and 2,4-Dinitrophenyl NADH/minute of reaction, depending hydrazine, which produce a reddish on NADH extention coeffecient which brown compound, Pyruvate hydrazone, is 6.2×103 M-3cm-1. at basic media. Absorption measured at VI-Statistical analysis:Student test 365 nm. was used for the analysis of data to C-Assay system for succinate compare differences between any two dehydrogenase (SDH) activity: SDH treatments (Steel and Torri, 1980). (EC1.3.99.1) activity was assayed using Statistical package for sociaty (spss) a method adopted by Al-Hasany(2004). program, version 12 was sought to Flavine adenine dinucleotide (FAD) analyse the data by computer. All reduced to FADH2 by succenic acid results were significant with P≤0.05. (SA), giving bright yellow color. Formation of NADH is propertional to the succinate dehydrogenase activity, Results and Discussion which is mesured spectroscopically as Biochemical estimation and increase in absorbance at 375nm. comparisom for the energy Concentration of consumered FAD was occumulated macromolecules in estimated according to Lambert-Bear intestinal cestodes i.e. O. bofonis and C. rule: C=A/Ɛ. Unit of enzyme activity columbae were expresed in table. 1. expressed as nM FAD/min/mg protein. Generally, concentration of D- Assay system for fumarate carbohydrate were higher than that of reductase (FR) avtivity: FR, (EC lipides and protiens in the two 1.3.99.1) activity was assayed examined helminthes. On the other according to modified method hands, there were significant described by Denicola-Seoane et differences at p≤0.05 between the al.(1992), addopted by Chen, et al. concentration of carbohydrates, lipides (2001) and Al-Hammoshi (2006). The and protienes of the tow cestodes, method depends on absorbancy which were, 153 µg/ml , 105 mg/100ml decrease of the reduced form of and 113 µg/ml, respectively in O. NADH at 340nm, in the presence of bofonis, and 240 µg/ml , 196 mg/100ml fumarate (the substrate), and crude and 181 µg/ml , respectively in enzyme in the worm extract of the C.columbae, depending on the wet worms. The reaction was monitored weight. Eventually, concentrations of spectroscopically at 37 ºC. The the three macromolecules were higher decrease in absorbance of NADH was in the pigeon cestode.

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A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

Table(1): Concentration of total carbohydrtes, lipids and proteines in Ophiotaenia bofonis and Cotugnia columbae extract: Cestode Carbohydrates conc.µg/ml Lipids conc.mg/100ml Protiens conc.µg/ml Ophiotaenia bofonis 153**±11.31 105**±9.21 113**±8.48 Cotugnia columbae 240±13.56 196±6.83 181±9.17 *Each value represent mean of three replicates±SE. **Reffered to presence of significant differences between the values at P≤0.05, according to t-test, two sample analysis.

The present resultes some what digonopora cestode depends on consisted with that of Al-Naftachi anaerobic carbohydrate metabolism to (2006), who compared biochemicaly obtain the energy required ,a regular between intestinal cestodes of four supply of glycogen is necessary vertebrate hosts, Bothreiocephalus sp .Hence large quantities of (in Barbus luteus fish), Ophiotaenia polysaccharide are stored which can be europaea (in snake), Raillietina oxidized to yield ATP. echinobothrida (in gull bird), and On the other hands, The present Moniezia expansa (in sheep) .She resultes agree with that of Al-Naftachi revieled that carbohydrates had the (2006) who elucidated that hiest concentration than protienes and concentration of carbohydrate in gull lipides in the four examened bird intestinal cestode (Raillietina helminthes. Wereas lipids had the less echinobothrida) was higher than that of percentage of the cestode wet wight. snake cestode (Ophiotaenia europaea). She gathered this result with the role of But not agree with the same researcher carbohydrates in these unaerobic of that proteines and lipids intestinal parasites as main energy concentration in snake cestode were reservior. The same as for Biswal, et.al. higher than that of gull bird cestode. (2012), who stated that carbohydrates has the highest percentage of dray HPLC analysis of lipids revealed that weight (25.2%) in Cotugnia cuneata O. bofonis adult worm contain six types (the intestinal cestode of domestic of fatty acides depending on retention pigeon), followed by protien (21.3%), time of the authentic fatty acid esters and lipid (9.9%), he concluded that (figure.1 and Table.2&3). Three were glycogen was the most domenant form saturated :myristic C14:0(0.89%); of carbohydrates. Furthermore, Sonune palmatic C16:0 (2.29%); and stearic (2012) pointed out that percentage of acid C18:0 (2.72%). The others were lipids is higher in the gastrontestinal non-saturated acids : oliec cestode of Ovis bahral sheep, than C18:1(7.91%) ; linoliec C18:2 carbohydrates and protienes. He (73.12%) and lenolenic C18:3(13.07%). reffered to the importance of lipides in The unsaturated Linoleic and linolenic these helminth as energy source and in acids were the most domenant types. ova formation. Waghmare and Chavan Fatty acid analysis of C.columbae adult (2010) stated that Cotugnia worm also, expose six types of fatty

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acids (Figure.2 and table 2&3). Four unsaturated fatty acids: oliec fatty acides were saturated :capric C18:1(0.06%) and linoliec acid C10:0 (0.19%); myristic C14:0 (2.38); C18:2(0.09%). The saturated stearic palmatic C16:0 (45.86%) and stearic and palmatic acids were the most acid C16:0( 51.42%), in addition to two domenant types.

Table(2): Retention times (minutes) of authentic fatty acids esters Saturated fatty acids Non-Saturated fatty acids Fatty acid Ca My Pa St Ol Li Ln C10:0 C14:0 C16:0 C18:0 C18:1 C18:2 C18:3 Retention 6.106 6.649 7.712 8.401 10.151 10.925 12.210 time(minute) Ca: Capric acid My: Myristic acid Pa: Palmatic acid St: Stearic acid Ol: Oleic acid Li: Linoleic acid Ln: Linolenic acid. Table(3): Types and percentage% of fatty acids found in Ophiotaenia bofonis and Cotugnia columbae extract Cestodes Fatty acids% Ca My Pa St Ol Li Ln Ophiotaenia bofonis 0.8 2.2 2.7 7.9 73.1 13.0 0.0 9 9 2 1 2 7 Cotugnia 0.1 2.3 45.8 51.4 0.0 columba 0.09 0.0 9 8 6 2 6 e

Figure (1): Chromatogram of fatty acid esters, which were separated from the extract of frog cestod, Ophiotaenia bofonis.

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Figure(2): Chromatogram of fatty acid esters, which were separated from the extract of pigeon intestinal cestod, Cotugnia columbae.

Thus, C16 and C18 were the most synthesis in cestodes is restricted to the common types of fatty acides in the two chain lengthining of host derived fatty studied cestodes , they may represent acides by sequential addition of acetyl- the main absorbed fatty acids through CoA. Jacobsen and Fairbarin (1967) body wall, those converted to the other demonstrated that Hymenolepis type of fatty acids within the parasite diminuta can convert palmate and body. This is come with some stearate into unsaturated fatty acids as consistance with the results of Aisien long as 26 carbon. Elongation of C16, and Ogiji (1989), who analyze fatty C18, C18:1, C18:2, C18:3 to C20 and acides of th lizard intestinal cestode, C22 fatty acids was demonstrated also Oochoristica agamea.They adverted in Spirometra mansonoides cestode that total lipids of the helminth fresh (Myer, et al. 1966). weight were 8.4%, and that 16 and 18- Glucose, fatty acids and amino acids are carbon fatty acids were the most the three substrates which organisms abundant fatty acids. The same as for can use to maintain metabolic Mondal et al. (2009) who analyses fatty homeostasis. They are aquired for the acids composition of Raillietina generation of energy, but also as a echinobothrida, the intestinal cestode of building blocks for the biosynthesis of fowl (bird), they referred to presence of the macromolecules. 18 fatty acids, and that chaif components fatty acids were palmatic In the present work, the variant (C16) and stearic acids (C18). It is concentration of carbohydrates, reported that cestodes are able to absorb proteines, lipides and fatty acides in the both short and long –chain fatty acides extract of the two cestod O.bofonis and directly from host intestine, through a C. columbae may related to the mixture of diffution and mediated variation between the frog and pigeon transport (Smyth and McManus, 2007). considering nutritional habits and the Smyth (1994) stated that fatty acid physiological differences between the

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A comparative study for some aspects of energy metabolism in two samples of intestinal cestodes: in house frog and domestic pigeon

two host; domestic pigeon, a bird that is normally as energy source reserve. endothermal (warm-blooded) When glucose supply become limitted, who need excessive metabolic rates body organs can use fatty acides allover the seasons. ( White, etal 2007 ); directly to generate energy (Houten and and house frog, an amphebian animal Wanders, 2010). Fraga et al (2012) that is poikilothermal ( cold-blooded) found that treating mice artificially animal, who need variant basic infected with Taenia metabolic rates between hibernation crassiceps cysticerci, with low dose of season and activity season( Moore, albendazole, resulted in block of 1964). Smyth and McManus (2007) glucose uptake by the larvae, the event endicated that glycogen (the most that induced fatty acids oxidation as important energy reserve in cestodes) energy alternative source ie. lead to fluctuates over a wide range, due to metabolic alterations in the larvae. that factor’s such as season’s ,physiological reffered to a great host-parasite state of the host, the time of autopsy interaction. Additionally,the adult ,strain of the host, rate of the infection cestodes take large ammount of lipids to and stage in life cycle. obtain suffecient ammount of fatty acids In parasitic helminths, the relationship and fat-soluble vitamines and the access between diet and fatty acides lipid is stored and eventually excreated composition is particularly marked in (Smyth ,1994) . Stored lipides are the cestodes (Barrett, 1983). Smyth and source of energy indispensable for the McManus ( 2007) reffered also to the formation of the cyst tissue in high variation in lipid contents of the Hymenolipis dimenuta (Moczon, 2006). cestodes. And that the ability to Lipids play an important role in long- synthsize fatty acids de novo in cestodes term adaptation and completion of the has been very much restricted and thus life cycle during their endoparasitic stage (Sato, et al. ,2008). they have to depend largely on the fatty acids of the host to fullfill their Mitochondria is the main site for requirments. George, et.al. (1969) were oxidation of plasma free fatty acids or analyse fatty acids composition of five lipoprotein-associated triglycerides (see intestinal cestodes, obtained from 5 Wanders, et al. 2010). Mitochondrial of shark. They find out that fatty acid β-oxidation (FAO) pathway there was complete qualitative play a pivotal role in the energy correspondance between host and homeostasis, but it compete with parasite acids, including the distinctive glucose as a primary oxidative C20-C22 polyunsaturated acides. As for substrate. The mechanism behind this C16 and C18 acides, they observed so-called glucose-fatty acid cycle or significant qualitative differences. Randle cycle (Bartlet and Eaton, 2004). George, et.al. (1971) were examined It is interesting to know that β-oxidation lipid content for three species of of one molecule of stearic acid (C:18) cestodes in leopard shark and brown will give 120 ATP molecules, while smooth hound shark. They find out that oxidation of one glucose molecule (C:6) there were obvious relationship between will give 32 ATP molecules. Thus, the present fatty acids and the marine Fatty acids have a higher ATP food chain. yield/Carbon atom than carbohydrates. As for energy metabolism, parasitic Besides, ATP yeald of saturated fatty stages of helminths do not use lipids acid is higher than that of unsaturated

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fatty acids ( Houten, and Wanders, gastrointestinal cestodes (table.4). Unit 2010). of enzyme activity for G6PDH and On the other hands, the cestodes utilize LDH in cellular extract was higher in O. different degrees of protein for bofonis , 7.515 U/L and 65IU/L producing energy. Litruchers revealed respectively, than that of C. columbae that the parasites able to adopt ,4.856 U/L and 37 IU/L respectively. themselves to the parasitic mode of life, The same as for the activities of SDH only due to protein (Jadhav, 2008). and FR, which were higher in the frog cestode , 130 nM FAD/min/mg protein In the present work, enzymatic studies and 198 nMNADPH/min/mg protein, depending on colorometric methodes, respectively than that found in the generally pointed to the significant pigeon cestod, 69nM FAD/min/mg difference between activity of the four protein and 133nM NADPH/min/mg estimated enzymes (G6PDH, LDH, protein respectively. SDH and FR ) at p≤0.05 in the two

Table(4): activity of G6PD,LDH, SDH, FR in Ophiotaenia bofonis and Cotugnia columbae adults extracts Enzyme activity G6PDH SDH FR U/L LDH nM FAD/min/mg nM Cestodes IU/L protien NADH/min/mg protien Ophiotaenia 7.514**±1.38 65**±3.8 130**±11.13 198**±6.15 bofonis Cotugnia 4.856±0.62 37±5.5 69±5.14 133±7.4 columbae *Each value represin ent mean of three replicates±SE. **Reffered to presence of significant differences between the values at P≤0.05, according to t-test, two sample analysis.

It is appeared that the cytosolic steps for controlling the extent of enzyme, G6PDH activity were very glycolisis or glycogen synthesis, since low in the extract of the two studied that G6PDH is the very first enzyme cestodes (Table. 4). This result of pentose phosphate shunt (Tian, et al, consisted with those recorded by 1998) , a metabolic pathway that Pampori and Srivasta (1987) , who supplies reducing energy to cells by found that G6PDH was present in very maintaining the level of the co-enzyme low concentration in Cotugnia NADPH. G6PDH reduces digonopora, a fowl intestinal cestode. nicotinamide adenine dinucleotide In the present work, the low phosphate (NADP) to NADPH while concentration of G6PDH in the the two oxidizing glucose-6-phosphate. Von cestodes (Table. 4) may reffered to that Brand ( 1979) manifested the this enzyme involved in rate limiting exestance of functional

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pentose phosphate cycle in accumulate succinate like: Echinococcus granulosus, Ascaris Hymenolepis diminuta, Moniezia lumbricoides, Hymenolepis dimenuta expansa, Echinococcus granulosus and Fasciola hepatica. cyst, Taenia taeniaformis adult and Al-Naftachi revieled that G6PDH larvae (Saz, 1970). Pampori, et al. activity in O. europaea (snake (1984) stated that fumarate and cestode) were higher than that of succinate induced significant Railletina echinobothria (avian production of ATP in Cotugnea cestode), she combined this result with dignopora adult cestod, and that a net the inherited variation. This is agree phosphorylation of approximately 2 with the present study that G6PDH mol of ADP was observed for each activity were higher in frog cestode mol of CO2 liberated from malate or than that of pigeon cestode. succinate. Pampori and Srivasta (1987) endicated also that FR and SDH, the LDH activity in the two studied mitochondrial enzymes were found in helminthes were higher than that of a significant concentrations in C. G6PDH of the same helminthes dignopora. Waghmare and Chavan (Table. 4), this result some what (2010) reffered to that presence of high simmilar to those reported in other level of SDH activity in Cotugnia parasites (Von Brand, 1979 and digonopora suggesting the existence Pampori and Srivasta ,1987). LDH of CO2 fixation pathway or partial brings about reduction of pyruvate reverse of Kreb’s cycle. In the present and results in production of lactic study, the higher activity of SDH and acid and thereby supplying an ATP FR in O. bofonis than that in C. molecule. Waghmare and Chavan columbae reffered to that carbohydrate (2010) bind the relatively high activity dissimilation in frog cestode and so of LDH in C. dignopora adult with ATP yeild is more active than that in the role of this enzyme in the energy pigeon cestode. supply and in the maintenance of the cytoplasmic redox state. Waghmare This variance in enzymes activity in and Chavan (2010) mentioned the the two helminthes may reflect the important role of pyruvate in the variance in metabolic rates of their energy metabolism of cestode, host, specislly during activity and pyruvate may converted to lactate by proliferation seasons. the action of LDH which is excreted Conclusion out along with energy rich product which utilized by the host. 1-The occumulated sources of energy (carbohydrates, lipides, and protienes) SDH and FR were found in significant semed to be more available in concentrations in the tow studied Cotugnea columbia (the pigeon worms (Table.4), indicated that these cestode). Semeoltaneously, the cestodes are aerobic fermenters, in activity of energy releaving enzymes which FR activity provide succinate , (G6PDH, LDH, SDH and FR) were which in turen used by SDH in the higher in Ophiotaenia bofonis (the frog opposite direction , also fumarate cestod). These variance between the involving this enzyme system giving two cestodes may related to the rise to succinate, could act as terminal variance of enveronmental conditions electron accepter (Bryant and Behm, within there habitats. 1989). Several cestodes were proved to

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