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1 The Mizo Post-Graduate Science Society was founded in 1995 and is chartered in Aizawl, India, as a non-profit, voluntary, scientific and educational organization to promote and make further development of science for the younger generation. Since 2001 the society publishes Science Vision, with financial assistance from the Mizoram Science, Technology & Environment Wing. Sci Vis is an interdisciplinary science journal, and is published quarterly in the months of March, June, September, and December. It aims to provide a forum for the reporting and discussion of news and issues concerning science, and to disseminate the information to the public. Though it is the official publication of the society, Sci Vis assumes no responsibility for opinions and conclusions drawn, or accuracy of data therein including those of editorials, research articles, news, commentaries and reviews which merely reflect the personal views of the authors and not official points of view of the MI- POGRASS or the institutions/organizations with which the authors are affiliated. Copyright 2009 by the MIPOGRASS. All rights reserved. Reproduction of the works in any material form, in whole or part, including the storing of it in any medium by electronic means other than private use is prohibited without written permission of the MIPOGRASS. Sci Vis is licensed under the terms of the Creative Commons Attribution License [Creative Commons Attribution- Noncommercial-No Derivative Works 2.5 India License], which permits the contents unrestricted use for noncommercial, educational purposes, and distribution, provided that the original author(s) and the source (viz, Sci Vis) are duly credited. Sci Vis authors are provided with the electronic copy of their articles, and they are at liberty to: (1) use their original figures or tables in their future works; (2) make copies of their papers for their own personal use, including classroom use, or for the personal use of colleagues; (3) include their papers as part of their dissertations; or (4) use all or part of their articles in printed compilations of their own works. However, citation of the original source must be included with due copyright notice of MI- POGRASS. Sci Vis is indexed in getcited. Cover: Raillietina echinobothrida from the intestine of local fowl, Gallus domesticus. Photo: K. Lalchhandama SCIENCE VISION ISSN CONTENTS 158 Editorial: Hmêl danglamna tlângah 159 Hazard effects of excess of zinc in diet K. B. Singh and K. Taneja 166 Laboratory evaluation of the pathogenicity of three entomopathogenic nematodes against larvae of cabbage butterf l y, P i e r i s b r a s s i c a e L i n n a e u s (Lepidoptera: Pieridae) Lalramliana and A. K. Yadav 174 On the structure of Raillietina echinobothrida, the tapeworm of domestic fowl K. Lalchhandama 183 Application of Mathieu potential to calculation of photocurrent from the surface of metals B. Zoliana, Z. Pachuau and R. K. Thapa 188 Traditional fishing methods in rivers and streams of Mizoram, north-east India H. Lalthanzara and P. B. Lalthanpuii science hmasâwnna langsârte 199 Ig Nobel Prize 2009 Volume 9, Number 4, October-December, 2009 Science Vision 2009 MIPOGRASS. All rights reserved

2 Original Research Sci Vis 9 (4), October-December, 2009 ISSN On the structure of Raillietina echinobothrida, the tapeworm of domestic fowl K. Lalchhandama Department of Zoology, Pachhunga University College, Mizoram University, Aizawl , India Received 12 November 2009 Revised 21 December 2009 Accepted 28 December 2009 ABSTRACT The structure of Raillietina echinobothrida, the gastrointestinal tapeworm of the domestic fowl, Gallus domesticus, was studied using light and scanning electron microscopy. There are already reports on the fine structure of the parasite; however, due to the choice of procedure, many of the resultant information are inadequate, contradicting, and occasionally, erroneous. A slightly modified technique in the microscopic preparations employed in the present study such as the use of formaldehyde as a fixative and tetramethylsilane prior to air drying for SEM, and successive treatments with xylene and clove oil during histological processing provided far more superior methods, and definitely, better results. Unlike in other studies, the scolex was unambiguously a round, distended and bulbous anterior end of the body. The suckers were protruding oval structures, while the apical rostellum was distinctly an invaginated, depressed and hollow structure. The central spaces of both the suckers and rostellum were covered with smooth tegument, made up of ciliary microtriches. The microtriches on the proglottids were arranged in smooth cascades, all directed toward the posterior, and giving the topography of the tegument a uniform velvety appearance. The body cavity was mostly occupied by uteri containing fertilized eggs in a gravid proglottid; and by testes, ovaries, and vitellarium dispersed among the parenchyma in a mature proglottid. The parenchyma notably filled up the remaining pseudocoel of the body. The distinctive characteristics of R. echinobothrida were established to be the double layered rostellum of pick mattock-shaped hooks, a thick short neck and a single egg in each egg capsule. Key words: Gallus domesticus; hooks; microscopy; microtriches; Raillietina echinobothrida; rostellum; suckers. INTRODUCTION The tapeworms belonging to the genus Raillietina Fuhrman, 1920 (phylum Platyhelminthes; class Cestoidea; subclass Eucestoda; order Corresponding author: K. Lalchhandama Tel (residence) chhandama@gmail.com Cyclophyllidea; family Davaineidae) are the second most prevalent avian helminth parasites, and particularly of domestic fowl, Gallus domesticus Linnaeus, 1758 (family Phasianidae); and R. echinobothrida Mégnin, 1880, is the most important species in terms of prevalence and pathogenicity. 1,2 The parasite inhabits the small intestine, from where it obtains nutrition 174 Science Vision 2009 MIPOGRASS. All rights reserved.

3 from the digested food of the host. The tapeworm is responsible for stunted growth of young chicken, emaciation of adult and decreased egg production of hen. In general the tapeworm does not cause gross pathological damages on well nourished chicken, but do compete for food when they grow to excessive number. In such situation, severe lesions on the intestinal walls and diarrhoea could arise, which ostensibly resulted in ill health. 3 Under heavy infestation, R. echinobothrida is listed as one of the most pathogenic tapeworms, causing conspicuous intestinal nodules in chicken, 4 with characteristic hyperplastic enteritis associated with the formation of granuloma. 5 The symptom is termed nodular tapeworm disease in poultry. Intestinal nodules often result in degeneration and necrosis of intestinal villi. 7 There are already a considerable number of reports on the structure of R. echinobothrida. But many, if not all, of them are imperfect, and some are utterly flawed apparently due to inappropriate methodologies employed, particularly while processing the parasites for microscopic studies. Some microscopic data also pose serious ambiguity as to the distinctive identifying features of R. echinobothrida in comparison with those of other Raillietina sp., especially of R. tetragona Molin, The identification is further compounded as the latest observations resolved that there really are no significant morphological differences between these species. 8 Recent investigations into the effects of anthelmintic agents on the structure of the parasite have clearly revealed that structural descriptions and morphological details in most of the available literature are not giving the precise picture This is discernibly due to R. echinobothrida and other soft-bodied helminths being rather delicate and easily distorted when they are subjected to certain chemicals during sophisticated microscopic processing. Therefore, it has become a veritable challenge to rectify the scientific information on the matter and to improve the protocol for better understanding the structure of R. echinobothrida that is the purpose of this study. MATERIALS AND METHODS Collection of tapeworms Live adult R. echinobothrida were collected upon autopsy from the small intestines of local fowl, G. domesticus, which were procured from the local abattoir at Aizawl, India. The tapeworm was identified at the Department of Zoology, North-Eastern Hill University, Shillong, India, as previously described The fresh worms were collected in and thoroughly washed with 9% phosphate buffered saline (PBS). The most robust specimens of the lot were chosen for microscopic processing. Chemicals and reagents All the chemicals and reagents used were standard analytical grades, obtained either from Merck or S.D. Fine Chemicals Limited, Mumbai, India, except alcohol, which was supplied by Bengal Chemicals, Kolkata, India. Scanning electron microscopy (SEM) Some of the tapeworms were fixed in 4% neutral phosphate-buffered formaldehyde at 4ºC at least for 24 h. After post fixation in 1% buffered osmium tetraoxide for 1h, the worms were washed with PBS. Subsequent dehydration was carried out through ascending concentration of acetone up to pure acetone. Following the standardized scanning electron microscopic methods developed by Dey et al. 12 and Roy and Tandon, 13 specific for helminth parasites, the specimens were treated with tetramethylsilane for 10 minutes and then allowed to dry at room temperature (~25ºC). The dried materials were placed on metal stubs and sputter-coated with gold in a fine-coat ion sputter, JFC-1100 (JEOL). The gold-coated specimens were ob- Science Vision 2009 MIPOGRASS. All rights reserved. 175

4 served under scanning electron microscope (LEO 435 VP) at an electron accelerating voltage of 20 kv. Photomicrography using light microscope Some tapeworms were fixed in aqueous Bouin s fluid overnight. The fixative was completely removed under running tap water, and the specimens were dehydrated through grades of alcohol up to pure ethanol. After cutting them into small pieces, they were treated with a mixture of xylene and clove oil until clear transparent specimens were obtained, and then cleared in pure xylene. After complete infiltration with liquid paraffin and molten wax at ~58ºC and embedding in solid paraffin, they were solidified under room temperature, and trimmed into rectangular blocks. Sections were cut at 8 μm thickness using Erma Japan type rotary microtome (Biocraft & Scientific Industries, India). The sections were then deparaffinized using pure xylene, were completely dehydrated, stained with eosin and haematoxylin, and finally mounted on glass slides using DPX mountant. Photomicrographs were prepared from Zeiss image analyzer HBO 50. RESULTS SEM observations of the morphology The cestode R. echinobothrida is a whitish, soft-bodied helminth. Scanning electron micrographs showed that the parasite possessed the classic tapeworm body contour composed of a succession of ribbon-like body segments, was extensively elongated, entirely covered with a tegument, and dorso-ventrally flattened, hence apt for the general name flatworm (Greek platys + helmins) (Fig. 1a). The body was broad at the posterior region, gradually narrowing toward the anterior end, and terminated at the anterior into a pin headsized scolex, the diameter of which was about μm (Fig. 1b). The scolex exhibited a globular design and bore two conspicuous structural components, namely suckers (= acetabula) and rostellum, which are the holdfasts of the worm to the mucosa of the host s intestine. There were four suckers which were rather evenly spaced from one another, thus located radially around the equatorial sphere of the scolex (Fig. 1b). One of the important observations was that each scolex was oval, about μm long along the longitudinal axis and μm broad along the transverse axis of the body, and lined with hundreds of finely pointed hooklets or spines along the rim. The spines were arranged in about 10 rows with each row running in diagonal direction but parallel to one another (Fig. 1c). Each spine was having a rod-like shaft and a pointed tip, and more or less uniform in length; μm long toward the outer rim, but relatively short, about 3-5 μm long toward the centre. All the spines were slightly curved and bent toward the centre of the sucker. The suckers surrounded at the centre of the scolex an apical mouth-like opening called rostellum, which had a diameter of about 40 μm, and slightly smaller than the suckers themselves. The rostellum was circularly lined with hundreds of pick mattock-shaped hooks, having an extended base and a pointed tip. These seemingly sharp tools are the device for grasping the host s intestinal wall during attachment. The hooks were arranged in two distinct layers, each of which in turn consisted of several rows of hooks (Fig. 1d). The outer layer consisted of 5-7 rows, whereas the inner layer was evidently made up of rows. Each hook in both the layers measured about μm in length. Within a layer, the hooks were aligned in such a meticulous pattern that all the rows overlapped with each other, and an individual hook of one row alternate with that of the next, displaying a regular zigzag design. The rostellum was not entirely hollow, instead densely filled up with microtriches at the centre. The scolex was followed by a body segment-forming portion termed the neck, which 176 Science Vision 2009 MIPOGRASS. All rights reserved.

5 a b c d Figure 1. Scanning electron micrographs of anterior portion of R. echinobothrida; a. A series of segments (proglottids) of the body proper (strobila) with terminal knob-like scolex, and a short neck in between; b. The scolex bearing apical mouth-like rostellum surrounded by four oval suckers (acetabula); c. The rim of the sucker showing a mass of fine microtriches in the upper region and rows of pointed spines in the lower region; d. The rostellar opening revealing two layers of mattock-shaped hooks along the rim, each layer consisting of rows of hooks, and the bottom left corner showing dense microtriches that fill up the hollow rostellum. was a short and unsegmented region, and a lavishly segmented body proper called the strobila. The strobila was highly elongated consisting of a long series body segments called the proglottids. The size of the proglottids steadily increased from the region succeeding the neck toward the posterior end of the strobila. The entire body covering of the body is named the tegument, and was densely covered with specialized hair-like microvilli called microtriches, all regularly directed toward the posterior, thereby giving the whole body surface a well groomed, velvety appearance (Fig. 2a). Each microtrich on the mature proglottid was filamentous, delicate and tubular, measuring about 5-7 μm in length, and with a rounded blunt end. The microtriches formed small periodic ridges and furrows all over the tegument, and were arranged as cascades of filaments so as to give the tegumental topography a smooth silky pattern (Fig. 2b). Science Vision 2009 MIPOGRASS. All rights reserved. 177

6 a b Figure 2. Scanning electron micrographs of a mature proglottid of R. echinobothrida; a. A smooth velvety surface of the tegument, forming ridges and furrows; b. Hairy microtriches covering the entire tegument in regular cascades, all directed toward the posterior. Observations of histological structure Photomicrographs of the transverse section of the proglottids of R. echinobothrida revealed that the body was entirely surrounded by a biologically active syncytial layer called the tegument (Fig. 3a,b,c). It is the outer epidermis of the thin tegument that gave off ciliary microtriches. The middle portion was a thick and homogenous subtegument. The innermost layer was a thin basal membrane and remained closely associated with a conspicuous muscular wall, made up of criss-crossing muscles, namely circular, dorso-ventral and longitudinal muscles. Body cavity was completely lacking with the internal region consisting of compact acoelomate layer, composed of spongy tissue called parenchyma, which is in turn made up of connective tissue and loose parenchymal cells. Several sac-like gland cells, nerve fibres, vitellarium or yolk gland, excretory and reproductive systems lie embedded in this parenchyma (Fig. 3c). The cestode is sexually monoeceous and contained a number of testes and two ovaries in each proglottid. Numerous testes were seen to be distributed on aboral side of each proglottid; while the ovaries and vitellarium were situ- ated at the centre of the body. However, the gravid proglottids toward the extreme posterior end of the body were almost entirely occupied by exceptionally prominent egg capsules, obscuring any other major organs (Fig. 3a,b). These egg capsules enclosed the developing embryos, referred to as oncospheres. Significantly, a single egg capsule contained only one oncosphere, which appears to be unique to R. echinobothrida (Fig. 3b). The gravid proglottids are ready for discharge to produce larvae in the external environment. DISCUSSION R. echinobothrida has recently emerged as one of the most widely studied helminth parasites, and as a commonly employed parasite model in the pharmacology of anthelmintic drugs, largely because of its world-wide prevalence, easy handling, and above all, elegant structure. The fine morphological intricacies and rather simplistic anatomy, owing to lack of complicated digestive and respiratory tracts, make the tapeworm an ideal subject of observation when treated with anthelmintics, since anthelmintics readily cause extensive structural damage. 178 Science Vision 2009 MIPOGRASS. All rights reserved.

7 B LC Tg LC LM Ec Ec Os P CM a b N Tt Ov V LM DVM CM Ec c Figure 3. Photomicrographs of histological sections of R. echinobothrida; a. Through a gravid proglottid showing the tegument (Tg), numerous egg capsules (Ec), prominent lateral excretory canals (LC) [x 50, bar = 100 μm]; b. Higher magnification showing distinct regions such as basal membrane (B), longitudinal (LC) and circular (CM) muscles, parenchyma tissue (P), egg capsules containing oncospheres (Os) [x 200, bar = 20 μm]; c. Though a mature proglottid showing dorso-ventral muscle (DVM), nerve cord (N), ovary (Ov), testes (Tt), and vitellarium (V) [x 200, bar = 20 μm]. However, it has come to light more than once that the tegument, particularly on the scolex, of R. echinobothrida is highly crinkled and irregularly folded, shrunken and corrugated with conspicuous creases around the suckers and rostellum. 1,8,14,15 This description appears to be not so. Such diagnoses are particularly elusive and misleading while identifying R. echinobothrida and R. tetragona. A series of observations have unequivocally revealed an otherwise description, which is in stark contrast with the said definitions Confirmed in this study, the tegument has a smooth surface, and the scolex is somewhat a prominent knob-like, spherically inflated, quadri-radially symmetric structure. The microtriches on the tegument were also layered in a velvety cascade throughout the strobila. The reason for these manifest differences is apparently due to the use of conventional protocol such as fixation with glutaraldehyde and critical point drying with CO 2 in the previous Science Vision 2009 MIPOGRASS. All rights reserved. 179

8 studies, which were demonstrated to cause severe deformation on the body of cestodes and trematodes, the soft-bodied helminths. 12 As persistently depicted, the general portrayal of suckers as rounded in R. echinobothrida, while oval in R. tetragona 3,4,16 is also crucially incongruous. Conforming to the microscopic observations of Isamu, 17 the present SEM study makes it definitive that the suckers are not essentially rounded, but impeccably oval. In fact SEM showed that the suckers of R. echinobothrida are almost perfectly identical to those of R. tetragona. 8 The presence of heavily armed, spiny suckers (hence the name echinobothrida) as the holdfast organs is the typical characteristic of the cyclophyllidean tapeworms. 1 The most obvious distinguishing feature of R. echinobothrida among the species of Raillietina is so far the rostellum, which bears double layers, not just rows, of hooks, while the closely identical species R. tetragona bears only a single layer, which is somewhat comparatively inconspicuous. The hooks of rostellum in R. echinobothrida as a matter of fact are uniquely arranged in two layers, with each layer consisting of several rows of hooks. Rostellum is a retractile holdfast, so, its appearance can be quite different when it is protracted 8,13,15 and retracted, as observed in this study. Moreover, the scientific literature is replete with the description of rostellar hooks of tapeworms as hammer-shaped, and intermittently reiterated in all standard textbooks. If rendering an analogy of tools is a compulsion, mattock-shaped would be the best choice of expression. Each hook utterly resembled a mattock the head of a pick mattock, to be precise in having a broad base, steadily tapering toward the top, and pointed at the apex. Another unique feature of R. echinobothrida was that each egg capsule contained a single egg, which developed into a single oncosphere as observed by Li et al. 8 Particularly in the gravid proglottid, each of the egg capsules was marked with a single prominent dark spot, the oncosphere, surrounded by yolk matters. The account of Isamu was probably mistaken for R. tetragona, 7 which described 3-8 eggs in a single egg capsule. Microtriches are documented to exhibit wide range of morphology, and serve as an identifying character among the members of Eucestoda. 25,26 They are also of special interest in pharmacology as they are the basic interface of the tapeworm with its surrounding, thus serve as the primary site of absorption of nutrients and the target site of anthelmintics The microtriches of R. echinobothrida were observed as fine cylindrical tubular filaments, with smooth rounded ends, and arranged in rows corresponding to the regular ridges of the tegument. Further, the common assertion that microtriches are presumably involved in the attachment of the parasite to the intestinal mucosa of the host, 14,31-32 may not be reasonable in case of R. echinobothrida, since the microtriches appeared too slender and delicate, with their blunt ends, to make strong holdfasts of any sort. A conclusion can be drawn from the present investigation that microscopy is the most powerful tool for the study of the structure of all range of organisms. However, different organisms are fashioned with a variety of structure and composition; therefore, conventional microscopic techniques can have critical drawbacks while specimens are being processed. For instance, the use of 4% formaldehyde as a fixative, in a duration of fixation for 24 h, air-drying with tetramethylsilane in SEM are reasonably superior to other protocols to study the structure of helminths like R. echinobothrida and the soft-bodied kind. ACKNOWLEDGEMENT Materials for scanning electron microscopy were processed by Dr. B. Roy, and image analyzer was used with permission of Prof. (Mrs.) V. Tandon, both in the Department of Zoology, 180 Science Vision 2009 MIPOGRASS. All rights reserved.

9 North-Eastern Hill University, Shillong, India. The research was in part supported by Higher and Technical Education, Government of Mizoram, in the form of the Mizoram Research Fellowship. REFERENCES 1. Cheng TC (1986). General Parasitology, 2 nd edn. Academic Press, Division of Hardcourt Brace & Company, San Diego, California, USA, pp Permin A & Hansen JW (2003). The Epidemiology, Diagnosis and Control of Poultry Parasites: An FAO Handbook. Food and Agriculture Organization of the United Nations, Rome, Italy, pp Small L (1996). Internal parasites (worms) of poultry. Agnote, 669, McDougald LR (2003). Cestodes and trematodes. In: Diseases of Poultry, 11 th edn (YM Saif, HJ Barnes, AM Fadly, JR Glisson, LR McDougald & DE Swayne, eds). Iowa State Press, Blackwell Publishing Company, Iowa, USA, pp Nadakal AM, Mohandas A, John KO & Muraleedharan K (1973). Contribution to the biology of the fowl cestode Raillietina echinobothrida with a note on its pathogenicity. Trans Amer Microsc Soc, 92, Kumar PR, Ravindran R, Lakshmanan B, Senthamil Selvan P, Subramanian H & Sreekumaran T (2007). Pathology of nodular tapeworm in backyard poultry. J Parasit Dis, 31, Isamu S (1954). Morphological studies on the chicken tapeworm, Raillietina (Raillietina) echinobothrida. Zool Mag [Doubutsugaku zasshi], 63, Li M, Hai-yun L & Bao-zuo Y (2009). Comparative study on morphology and development of two species of Raillietina from chicken. Chin J Parasitol Parasit Dis, 27, Roy B, Lalchhandama K & Dutta BK (2007). Anticestodal efficacy of Acacia oxyphylla on Raillietina echinobothrida: a light and electron microscopic studies. Pharmacologyonline, 1, Roy B, Lalchhandama K & Dutta BK (2008). Scanning electron microscopic observations on the in vitro anthelmintic effects of Millettia pachycarpa on Raillietina echinobothrida. Phcog Mag, 4, Lalchhandama K (2009). Cestocidal activity of Acacia caesia stem bark on Raillietina echinobothrida. Phcog Res, 1, Dey S, Basu Baul TS, Roy B & Dey D (1989). A new rapid method of air-drying for scanning electron microscopy using tetramethylsilane. J Microsc, 156, Roy B & Tandon V (1991). Usefulness of tetramethylsilane in the preparation of helminth parasites for scanning electron microscopy. Riv Parassitol, 8, Radha T, Satyaprema VA, Ramalingam K, Indumathi SP & Venkatesh C (2006). Ultrastructure of polymorphic microtriches in the tegument of Raillietina echinobothrida that infects Gallus domesticus (fowl). J Parasit Dis, 30, Tandon V, Pal P, Roy B, Rao HS P & Reddy KS (1997). In vitro anthelmintic activity of root-tuber extract of Flemingia vestita, an indigenous plant in Shillong, India. Parasitol Res, 83, Kaufmann J (1996). Parasitic Infections of Domestic Animals: a Diagnostic Manual. Birkhäuser Verlag, Basel, Switzerland, p Isamu S (1985). Raillietina (Raillietina) galli Yamaguti is a synonym of Raillietina (Raillietina) tetragona (Molin). Zool Mag [Doubutsugaku zasshi], 64, Chandler AC (1923). Observations on the life cycle of Davainea proglottina in the United States. Trans Amer Microsc Soc, 42, Erickson AB (1974). Helminth parasites of rabbits of the genus Sylvilagus. J Wildl Manage, 11, Bartel MH & Hansen MF (1964). Raillietina (Raillietina) loeweni sp. n. (Cestoda: Davaineidae) from the hare in Kansas, with notes on Raillietina of North American mammals. J Parasitol, 50, Mariaux J, Bona FV & Vaucher C (1992). A new genus of Metadilepididae (Cestoda: Cyclophyllidea) parasitic in Terpsiphone rufiventer (Aves: Muscicapidae) from the Ivory Coast. J Parasitol, 78, Jones A & Anderson TJC (1996). Raillietina melomyos n. sp. (Cestoda, Davaineidae) from mosaictailed rats in Papua New Guinea. Syst Parasitol, 33, Hoberg EP (1999). Phylogenetic analysis among the families of the Cyclophyllidea (Eucestoda) based on comparative morphology, with new hypotheses for coevolution in vertebrates. Syst Parasitol, 42, Nikolov PN & Georgiev BB (2008).Taxonomic revision and phylogenetic analysis of the cestode genus Paraprogynotaenia Rysavy, 1966 (Cyclophyllidea: Progynotaeniidae). Syst Parasitol, 71, Whittaker FH & Garvajal JG (1980). Scanning electron microscopy of scolices of some cestodes from elasmobranchs. Proc Helminthol Soc Wash, 47, Lumsden RD and Hildreth MB (1983). The fine structure of adult tapeworms. In: Biology of the Eucestoda Vol.1 (C Arme & PW Pappas, eds). Academic Press, New York, USA, pp Becker B, Mehlhorn H, Andrews P & Thomas H (1980). Scanning and transmission electron microscope studies on the efficacy of praziquantel on Hymenolepis nana (Cestoda) in vitro. Z Parasitenkd, 61, Science Vision 2009 MIPOGRASS. All rights reserved. 181

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