Hittite Journal of Science and Engineering, 2014, 1 (1) 07-11 ISSN NUMBER: 2149-2123 DOI: 10.17350/HJSE19030000002

Investigation of Midgut’s Ultrastructure of viridis Decourt, 1909 and Notonecta maculata Fab., 1794 (: )

Menderes Suicmez and Rana Ozmen Hitit University, Department of Molecular Biology and Genetics, Corum, TURKEY

ABSTRACT Article History: Received: 2014/08/18 Accepted: 2014/11/19 Online: 2014/12/31 n this study, midgut’s ultrastructure of the Notonecta viridis and Notonecta maculata was

Iexamined under transmission electron microscope (TEM). It was observed that midgut Correspondence to: Menderes Suicmez, of the Notonecta viridis and Notonecta maculata are almost similar. Digestive canal is Hitit University, Faculty of Arts and divided into three parts as foregut, midgut and hindgut. Midgut is wider and longer part Science, Department of Molecular Biology and Genetics, Corum, Turkey than other parts of digestive canal. Midgut’s heamasoel side is covered with muscular Tel: +90 (364) 227-7000 (1669) tissue and connective tissue and lumen side is covered with epithelial tissue. Epithelial Fax: +90 (364) 227-7005 layer consists of three different cells: Endocrine cells, regenerative cells and enterocytes E-Mail: [email protected] cells. Endocrine cells possess secretory granules in the cytoplasm and they have the basement membrane folding in basal. Regenerative cells are small undifferentiated cells which are responsible for cellular regeneration. Enterocyte cells have many mitochondria and deep basal membrane folding in basal.

Key Words: Notonecta viridis; Notonecta maculata; Midgut; Ultrastructure

INTRODUCTION

pparently, since have so many species and Digestive canal of insects is divided into three parts Aperceived as harmful organisms, it has brought as front, middle and last intestine. Pre and post intestine to mind the idea of avoidance of these organisms and is ectodermic and coated with chitin-cuticle but midgut fighting with them. Insects which we accept both is endoderm origin and not coated [3]. Notonecta viridis harmful as well as beneficial even have important and Notonecta maculata (Hemiptera: Notonectida) are functions in maintaining the ecological balance capable of feeding predators and this situation impede [1]. body is divided into three parts as head, excessive reproduction of other insects in the aquatic thorax and abdomen. Head segments are completely environment. Because these species are fed everyday by fused and the boundaries of segments cannot be eating 24-28 larvae, they help to provide the distinguished in adults [2]. balance of food chain in the aquatic environment. So, by preventing excessive reproduction of mosquitoes, they In generally, insects get their nutrients by biting- have an important place as ecological for the reduction chewing or sucking but internal parasite larvae get of the demand for insecticides [1]. their nutrients through the body surface. Types of diet and food types of insects are as follows feeding with MATERIAL AND METHOD various parts of plants (herbivores), feeding with peat Specimens were placed in 3% glutaraldehyde in moss (saprophytes), feeding with small insects or other 0.1 M phosphate buffer at pH 7.2 for 3 h at room , feeding with fungi and bacteria, feeding with temperature. After rinsing in phosphate buffer, the feces, carrion and blood, and those feeding with the specimens were postfixed in 1% buffered osmium filtrate. M. Suicmez and R. Ozmen / Hittite J Sci Eng, 2014, 1 (1) 7–11 The digestive canal of canal digestive The X6000) Midgut epithelium consists of three different cell types types cell different ofthree consists epithelium Midgut RESULTS cells. It is seen that they are almost similar to each other of both species are seen similar. Midgut The abdomen. the in folds many has Hindgut canal. is surrounded abdomen the in found Midgut proventriculus. enlarged other like hindgut and midgut offoregut, consists examined using a Zeiss Libra120 electron microscope. electron Libra120 aZeiss using examined sections Thin araldite. in embedded and dehydrated were stained with uranyl acetate and lead citrate [4] and citrate lead and acetate uranyl with were stained Hemiptera. Foregut consists of eosophagus and partially partially and ofeosophagus consists Foregut Hemiptera. a layer of muscle and connective tissue from outside. outside. from tissue connective and ofmuscle a layer such as endocrine cells, enterocytes and regenerative regenerative and enterocytes cells, endocrine as such tetroxide at pH 7.2tetroxide for 3hat 4 the length of insects. Morphologically, digestive canals canals digestive Morphologically, ofinsects. length the than to2times 1.5 is canal digestive ofthe length total by a single layer of epithelial tissue from inside and by and inside from tissue ofepithelial layer by asingle is wider and longer from other parts of the digestive digestive ofthe parts other from longer and wider is Basal membrane folding, membranefolding, Basal Figure 3. Figure Figure 1. M: Mitochondria, Mitochondria, Regenerative cell intheof N. maculata midgut Enterocyte cell in the midgut of N. maculata N: Nucleus, Nucleus, No : Nucleolus, : Nucleolus, V: N. maculata Vacuol (TEM, x6000) (TEM, Vacuol Sg : Secretory granule (TEM, granule (TEM, : Secretory o C. They were then and N ; : Nucleus, Nucleus, : No: N. viridis Nucleolus, Nucleolus, Bmf :

08 The same nucleus type, in a similar manner, is seen at seen is manner, asimilar in type, nucleus same The viridis viridis viridis membrane folding, folding, membrane endocrine cells of the intestine of these two species feeding feeding species two ofthese intestine ofthe cells endocrine 4). 3and (Figures portion basal in cell the into extending foldings membrane basal that observed It is nucleus. in nucleoli obvious are there and dissemination ahomogeneous shows chromatin The cell. the within dense notvery are which mitochondria and reticulum endoplasmic or in groups in basal position round or oval-shaped cells 2). 1and (Figures other toeach equal of cells ofepithelial ultrastructure the to considered plasmic reticulum (TEM, x8 000) x8 (TEM, reticulum plasmic nucleus are seen in the enterocytes cells of cells enterocytes the in seen are nucleus and the density of intracellular mitochondria is almost almost is mitochondria ofintracellular density the and the in chromatin distribute non-homogenously and Nucleus that are responsible for cellular regeneration contain rough rough contain regeneration for cellular responsible are that inclusions are seen in the enterocytes cells of cells enterocytes the in seen are inclusions Figure 2. Figure 4 The most significant content is the secretory granule in in granule secretory the content is significant most The Regenerative cells exist in epithelial tissue separately separately tissue epithelial in exist cells Regenerative . However, more intense amounts of intracellular ofintracellular amounts . However, moreintense Sg: . and Regeneratif cell in midgut of Enterocyte cell in the midgut of N. viridis Secretory granules, granules, Secretory N. Maculata No: Nucleolus, Nucleolus, . A large and smooth nucleus nucleus smooth and . Alarge M: To Mitochondria, : Tracheol (TEM, X8000). (TEM, : Tracheol Notonecta viridis Notonecta RER: . No: . No: N. maculata Nucleolus, . Rough endo- Rough

N. viridis Bmf: Bmf: Basal Basal N: N. N. .

Figure 5. Endocrine cell in midgut of N. maculata. Sg: Secretory granule, Figure 6. Endocrine cell in midgut of N. maculata. M: Mitochondria, RER: Rough endoplasmic reticulum, M: Mitochondria, N: Nucleus (TEM, Sg: Secretory granüle, G: Golgi apparatus (TEM, X 16 000) X 8 000) DISCUSSION with foods. Furthermore, the cell is seen very rich in terms of mitochondria and rough endoplasmic reticulum The digestive canal of N. viridis and N. maculata contains (Figure 5 and 6). Vacuole-like structures and also Golgi similar structures as in many species of Hemiptera apparatus are observed in the cells. Mitochondria and the which are searched before. However, Barber et al. [5] density of rough endoplasmic reticulum are higher than reported that midgut consists of four parts including the other cell types. first, second, third and fourth ventriculus and there are Malpighian tubules at the junction of the midgut and last It is observed that N. viridis’ endocrine cells are intestine. much more intense endoplasmic reticulum (Figure 7). It is observed density of basal membrane foldings in basal The midgut epithelium consists of regenerative cells, portions of these cells and there are mitochondria between enterocyte cells and endocrine cells as reported by Demir these basal membrane foldings (Figure 8). and Suiçmez [6]. As stated in many studies [7-12], we have observed significantly larger nucleus, nucleolus and the The reason of both enterocytes and endocrine cells RER cisternae in the cytoplasm of the midgut cells of both contain secretory granules in greater amounts is probably species. duo to their tasks related to digestive and secretory. Both enterocytes and endocrine cells contain secretory granules Several authors [7, 8, 10, 14-18] reported the existence in greater amounts since probably making tasks are related of enterocytes cells responsible for absorption which were to digestive and secretory. These secretory granules are observed in the midgut of N. maculata and N. viridis. spread approximately the same amount on each side of the cell. / Hittite J Sci Eng, 2014, 7–11 1 (1) M. Suicmez and R. Ozmen M. Suicmez Figure 7. Endocrine cell in midgut of N. viridis. Sg: Secretory granule, Figure 8. Endocrine cell in midgut of N. viridis. M: Mitochondria, Bmf: RER: Rough endoplasmic reticulum, M: Mitochondria (TEM, x10 000) Basal membrane folding (TEM, x10 000)

09 M. Suicmez and R. Ozmen / Hittite J Sci Eng, 2014, 1 (1) 7–11 15, 19]. Hung et al. and Levy et al. supported that a large alarge that supported al. et Levy and 15, 19]. al. et Hung [8, 15]. Nucleus has a few nucleoli as stated byRost- stated as nucleoli [8, afew 15]. has Nucleus Levy et al. and Nardi et al. [11, 15, 19] were also observed in [11, in al. et observed 15,19] Nardi and were also al. et Levy contain well-developed rough endoplasmic reticulum and and reticulum endoplasmic rough well-developed contain in seen are foldings membrane basal obvious very that al. et byRost-Roszkowska supported It is secretion. tomake cells specialized are cells [6,12,14,21]. these that cells It suggests Urospherite and phosphate observe itcouldn’t of midgut, Buschinger, and byBillen were stated which vesicles electron [19]. al. et byNardi Transparent indicated as midgut of the in cells ofenterocyte cytoplasm of the cell as indicated by Fialho et al., Hung et al. al. et Hung al., et byFialho indicated as cell of the mitochondria [6, 21]. mitochondria other in than cells endocrine in granules more secretory Reticulum cisternae that were mentioned by Hung et al., al., et byHung were mentioned that cisternae Reticulum Endoplasmic Smooth and Reticulum Endoplasmic Rough [16, Rost-Roszkowska Undrul 17] and observed notbe could Roszkowska and Undrul [17] Undrul seen and Roszkowska are nucleolus and number of basal membrane infoldings, which are formed by by formed are which infoldings, membrane ofbasal number related to the synthesis and secretion functions since they they since functions secretion and synthesis tothe related [11,15]. mitochondria with associated are cells, regenerative granules and large lysosomal bodies weren’t detected either either weren’t detected bodies lysosomal large and granules and Demir and Suiçmez reported that these cells are directly directly are cells these that reported Suiçmez and Demir and [17]. byRost-Roszkowska Undrul and confirmed as acid region other any in clearly observed be couldn’t apparatus Golgi cells, [20]. enterocyte byTaha In al. et mentioned as able to mark which were observed 11, 15,17, [6-8, byauthors 12, reported as 19]. before [7, 8]. Ferritin are quite large and smooth and cover a large portion portion alarge cover and smooth and large quite are structures which contains carbonate chloride but non-uric but non-uric chloride carbonate contains which structures surrounded [14]. al. et byGül stated as nucleus the in bysignificantly mitochondria were observed in the the basal portion endocrine cells [21]. cells endocrine Rost-Roszkowska al. et portion basal the extensions nomicrovilli have and cells enterocyte the than epithelium [15]. differentiation the types During cell these of parts apical the in extensions nomicrovilli are there that indicated al. et Hung we examined. species ofboth cells the of study the in cells endocrine into differentiate can that both species we dealt, as stated by other authors [6, 7, authors byother 11, stated as 12, we dealt, species both in a very clear way in our study. study. our in way clear avery in in the apical parts as in enterocytes cells [13]. cells are There enterocytes in as parts apical the in of membrane basal ofthe cells the into made invaginations As Habibi et al. mentioned endocrine cells are shorter shorter are cells endocrine mentioned al. et Habibi As A large number of rough endoplasmic reticulum reticulum endoplasmic ofrough number A large We observed the digestive cells and regenerative cells cells regenerative and cells digestive the We observed The nucleus of nucleus The Glycogen granules and spherical crystals weren’t crystals spherical and granules Glycogen N. maculata N. maculata enterocyte cells cells enterocyte and N. viridis

10 ACKNOWLEDGEMENTS We are indebted to the Hitit Universty Scientific Research Research Scientific Universty Hitit tothe We indebted are This research was supported by the Hittite University University Hittite bythe supported was research This These insects which meet their food from aquatic aquatic from food their meet which insects These These chemicals are getting through to organisms in in toorganisms through getting are chemicals These 1. N. viridis Mustafa Duran, who identified the species. species. the identified who Duran, Mustafa 4. CONCLUSIONS Projects for financial support of this research. I also Ialso research. ofthis support for financial Projects Scientific Research Projects (Project No. FEF03.10.003). No.FEF03.10.003). (Project Projects Research Scientific various ways. It will be impossible to sustain their existence existence their tosustain impossible be It will ways. various express thanks to Mr. Mehmet Aydınkal for correcting for correcting Aydınkal toMr. Mehmet thanks express directly or indirectly affected from the adverse changes in that in balance in remain population their provide can organisms also important fulfill functions in maintaining over time as as over time chemicals many too are There live. they which environment maintain their presence in the aquatic environment. However, it’s an inevitable reality that they are either either are they that reality inevitable However, it’s an region. region. place to combat mosquitoes as an alternative way. Because 2. and this prevents their excessive proliferation and they they and proliferation excessive their prevents this and affected by these chemicals as in other organisms. other in as chemicals bythese affected species are also used as indicator organisms in the aquatic aquatic the in organisms indicator as used also are species the English of the manuscript. I specially thank Prof. Dr. Prof. thank Ispecially manuscript. ofthe English the their environment. When evaluated with this aspect these proliferation. excessive their prevent also and species the of population the control they predators, are species these these earlier; we mentioned As balance. ecological the insect species consume the mosquito larvae as nutrient nutrient as larvae mosquito the consume species insect in soil and water as a result of so many agricultural activities. activities. agricultural many so of result a as water and soil in 3. Thanks to these features, they have an important Yayıncılık, Ankara, 2011. Aldemir A, Boşgelmez A. A. Boşgelmez A, Aldemir Ankara, 2006. Ankara, (Cypriniformes:Cyprinidae) ve Ozban N, Özmutlu Ö. Mikropreparasyon Yöntemleri. İ.Ü.Fen Yöntemleri. Ö. Mikropreparasyon Özmutlu N, Ozban Salman S. Omurgasız Hayvanlar Biyolojisi, Palme Yayıncılık, Yayıncılık, Palme Biyolojisi, Hayvanlar Omurgasız S. Salman Fakültesi Yayınlarından, İstanbul, 1991. İstanbul, Yayınlarından, Fakültesi Palme Biyoloji. Genel E. Gündüz, and İ, Türkan A, Demirsoy 2003. Ankara. Beytepe, ABD, Ekoloji Bölümü, Biyoloji Fen Fakültesi, Üniversitesi Hacettepe Kars. Bölümü, Biyoloji Fakültesi, Fen-Edebiyat Üniversitesi Tezi, Kafkas Doktora Notonectidae)’in Sivrisinek Larvaları Üzerindeki Predasyonu. and N.viridis N. maculata REFERENCES and N. maculata are aquatic insects and and insects aquatic are Notonecta viridis are also negatively negatively also are Alburnus orontis (Hemiptera: (Hemiptera: 5. Barber Dan T,Cooksey Lynita M, Abell, Dawid W. A Study 14. Gül N, Sayar H, Özsoy N, Ayvalı C. 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