World Journal of and Marine Sciences 6 (4): 319-327, 2014 ISSN 2078-4589 © IDOSI Publications, 2014 DOI: 10.5829/idosi.wjfms.2014.06.04.84274

Diplectanum harid Sp.nov. And chlorostigma Sp.nov. (: ) from Scaridae and Hosts of the Red Sea

12Kareem Morsy, Hoda El Fayoumi, 2 Gamal Al Shahawy and 2 Mohamed Fahmy

1Department of Zoology, Faculty of Science, Cairo University, Giza, Egypt 2Department of Zoology, Faculty of Science, Beni-Suef University, Beni-Suef, Egypt

Abstract: harid sp. nov. and Pseudorhabdosynochus chlorostigma sp. nov. (Monogenea: Diplectanidae) were described from the gills of the parrot fish Scarus harid (F: Scaridae Forsskal, 1775) and Brown spotted reef cod chlorostigma (F: Serranidae Valenciennes, 1828) respectively. Fish were collected from boot landing sites and fishermen at different water locations at Hurghada City along the Red Sea, Egypt. The morphology and morphometric characterization of the isolated worms were described by means of light microscopy. Twenty five (62.5%) out 40 speciemens of Scarus harid were infected with a Diplectanum sp. and thirteen (54.2%) out 24 specimens of Epinephelus chlorostigma were infected with Pseudorhabdosynochus sp. most of the infected fish had very pale gills and showed symptoms of anemia. Morphologically, the adult worm of Diplectanum sp., possessed a body which was elongated with a total length 0.574-0.579 (0.576±0.02) mm and a maximum width 0.103-0.107 (0.105 ±0.02) mm at the level of . , broad, differentiated from the rest of the body, measured 0.17–0.21 (0.19±0.02) mm and provided with continuous rows of . Two pairs of lateral hamuli, three bars and 14 marginal hooklets were also observed. The copulatory organ was small and tubular, with no accessory piece and composed of anterior cone and straight posterior tube with total length 0.063-0.082 (0.069±0.002) mm. The body of the second Pseudorhabdosynochus chlorostigma sp.nov was wide with a length 0.54 –0.58 (0.56±0.02) mm and width 0.19–0.23 (0.21±0.02) mm. The anterior region was equipped by 3 pairs of head organs and 2 pairs of eye-spots. Copulatory organ was composed of a long coiled tube that terminated in a club-shaped structure which was characteristic for this species and measured 0.070–0.072 (0.071±0.002). Haptor differentiated from the rest of the body, with a width of 0.26–0.30 (0.28±0.02) mm and provided with similar squamodiscs, 2 pairs of lateral hamuli, 3 bars and 14 marginal hooklets. The two new species were compared with those described previously from the same , it was shown that there were significant morphological and morphometric differences especially for the copulatory organ, which was a strong criteria for the placement these monogenean parasites as new species with new and locality records in Egypt.

Key words: Diplectanum harid sp.nov Pseudorhabdosynochus chlorostigma sp.nov Monogenea Diplectanidae Red Sea Light microscopic study

INTRODUCTION severe destructions of the gills as well as severe losses too [2]. Gills of infested fish were congested or pale Fish are important members of aquatic ecosystems haemorrhagic with hyper secretion of mucus. These signs and an important source of human food. Increased may be due to severe irritation caused by movement, interest in fish culture has also increased awareness of feeding activity, fixation and attachment of monogenean and experience with parasites that affect fish health, worms. Also, the presence of thick mucus secretion leads growth and survival. The gills of fish represent one of the to respiratory failure and osmotic stress ending in the fish biotope mostly exploited by different fish ectoparasites death. Like all ectoparasites, monogeneans have well [1], among them, monogenetic trematodes that cause developed attachment structures, the anterior attachment

Corresponding Author: Kareem Morsy, Department of Zoology, Faculty of Science Cairo University, Giza, Egypt.

319 World J. Fish & Marine Sci., 6 (4): 319-327, 2014 organ and the posterior haptor which is associated with MATERIALS AND METHODS hard (sclerotized) structures in the form of hooks, anchors, clamps. The disease caused by monogenean A total of 40 specimens of Scarusharid (F: Scaridae parasites, causes serious problems in aquaculture [3-9] [20]) and 24 of Epinephelus chlorostigma (F: Serranidae) with an obvious pathogenicity. Immature worms of these (size range: 14-28 cm, mean 18.5±7.15cm; body weight parasites attach to the gill filaments of their hosts and 100-250 g, mean 205±20 g) were caught from the coasts of migrate to the buccal cavity wall for maturation, as the Hurghada City of the Red Sea in Egypt. Samples were worms ingest the blood from the gills of host fish, heavily obtained at irregular intervals in 2013. Captured fish were infected wild and cultured fish become anaemic [9,10]. kept alive in aquaria filled with the same water source and The genus Diplectanum [11] represents one of the examined within few hours. Skin surface, fins and gills genera of the family Diplectanidae which are gill parasites were firstly examined by naked eyes and with the help of of marine perciformes [12, 13]. Family Diplectanidae a dissecting microscope for any attached parasites, includes about 22 genera and more than 218 valid species. lesions or external changes. After removing opercula and The first species which detected as a member of exposing gill arches, each gill was removed carefully from Diplectanidae was proposed by [14] named as the fish, immersed in normal saline to remove any excess aequans. Later, Diesing [11] proposed the gill mucus. Monogenean parasites were recovered with a genus Diplectanum as a new genus when he transferred Pasteur pipette using a dissecting binocular microscope. Dactylogyrus aequans to Diplectanum aequans and Worms were fixed in 4% formalin and then washed with considered it as a type species of this new genus. distilled water to remove excess fixative. Worm Recently, Diesing [11] mentioned that the genus identification was confirmed by mounting specimens on Diplectanum is restricted to species that have male slides in drops of ammonium picrate glycerine under cover copulatory organ with nested tubes, accessory slips and examining hard parts using light microscopy. For copulatory organ, prostatic reservoir separated into three each monogenean parasite, the sclerotized parts of the zones, ventral and dorsal squamodiscs. haptor were measured using an ocular micrometer Yamaguti [15] established Pseudorhabdosynochus calibrated against a stage micrometer slide [21]. Ten for P. epinepheli from the gills of Epinephelus akaara specimens were measured for the range and the mean ± collected from the Inland Sea of Japan. The genus was standard deviation (SD). Prevalence, mean abundance and characterized by the presence of a sclerotised male measurements followed the guidelines of Bush et al. [22]. copulatory organ and a "quadriloculate organ" composed of four chambers and squamodiscs reduced to RESULTS membranous plaques with several curved transverse ridges. Twenty five (62.5%) out of 40 specimens of Scarus Species of Pseudorhabdosynochus parasitize marine harid were infected with a Diplectanum sp. and thirteen fish in warm seas. Santos et al. [16] listed the valid (54.2%) out 24 specimens of Epinephelus chlorostigma species with the addition of P. coioidesis Bu et al. [17] were infected with Pseudorhabdosynochus sp. most of from Epinephelus coioides and E. areolatus in Malaysia, the infected fish had very pale gills and showed Hong-Kong and Indonesia and P. chinensis Zhang et al. symptoms of anemia. [18] from E. tauvinain , Pseudorhabdosynochus Diplectanum harid sp. nov. (Figs. 1-8, 17I, Table 1) now comprises 24 species, of which 20 parasitize serranids Body of the adult worm was elongated with a total mainly of the genus Epinephelus. length 0.574-0.579 (0.576±0.02) mm and a maximum width Diplectanids often show strict host-specificity [19]; 0.103-0.107 (0.105 ±0.02) mm at the level of ovary. The thus, it is a safe prediction that many other species of anterior attachment organ had three pairs of cephalic Pseudorhabdosynochus are still undescribed. glands. Also, two pairs of eyes were present on the dorsal During a survey of gill monogenean parasites of body region anterior to the . The posterior pair of marine fish, the parrot fish Scarus harid (F: Scaridae, [20]) eyes was larger than the anterior one. The muscular and Brown spotted reef cod Epinephelus chlorostigma pharynx was sub spherical and was located behind the (F: Serranidae) from the Red Sea, Egypt were found margins of the posterior pair of eyes. The caeca was infested by two new species of diplectanids. Herein, simple and terminated blindly at the level of posterior descriptions of these new species were carried out using margin of the vitelline field. A quadriloculate organ with light microscopy. four chambers measured 0.066–0.070 (0.068±0.002) mm

320 World J. Fish & Marine Sci., 6 (4): 319-327, 2014

Figs. 1-8: Photomicrographs of the adult Diplectanum harid sp.nov.1 Whole mount preparation of the adult worm showing its anterior attachment organ (AT), two pairs of eyes (E), copulatory organ (CO), Quadriloculate organ (Q), squamodiscs (S) and the posterior haptor (H). 2-7 High magnifications of: 2haptor (H) consists of squamodiscs (S), two pairs of hamuli, one pair of dorsal hamuli (DH) and one pair of ventral hamuli (VH), a ventral bar (VB) and one pair of dorsal bars (DB). 3 Quadriloculate organ (Q). 4,5 Copulatory organ (CO). 6 Dorsal bar (DB) and the dorsal hamulus (DH). 7 Dorsal bar (DB), dorsal (DH) and ventral (VH) hamuli. 8 Ventral bar (VB)

321 World J. Fish & Marine Sci., 6 (4): 319-327, 2014

Fig. 17I: Schematic drawings of: a Haptor of the adult Diplectanum sp.nov., composed of ventral bar (VB) with two ventral hamuli (VH), dorsal bar (DB) with two lateral dorsal hamuli (DH) and squamodiscs (S). b Quadriloculate organ. c Copulatory organ

Table 1: Comparison between measurements of Diplectanum harid sp.nov. recorded from the current study and some of the previously described species Aspect D. calathus Edgar et al. 2008 D. parvus Jean-Lou, J., 2008 D. gatunense Edgar et al. 2008 Present study Total body length 0.625 (0.417-1.050) 0.320 (0.178-0.330) 0.575 (0.260-0.787) 0.576(0.574-0.579) Maximum body width 0.160 (0.115-0.250) 0.085 (0.065-0.145) 0.108 (0.087-0.130) 0.105 (0.103-0.107) Ventral humuli -Total length 0.032 (0.024-0.045) 0.040 (0.036-0.043) 0.040 (0.038-0.041) 0.027 (0.024-0.029) -Shaft length 0.030 (0.020-0.041) 0.032 (0.030-0.038) 0.020 (0.019-0.021) 0.012 (0.010-0.014) -Inner root length - - - 0.016 (0.015-0.018) -Outer root length - - - 0.007 (0.006-0.008) -Point length - - - 0.005 (0.004-0.006) Ventral bar -Total length 0.045 (0.028-0.051) 0.073 (0.072-0.099) 0.125 (0.114-0.136) 0.023 (0.021-0.024) -Width 0.012 (0.010-0.015) 0.008 (0.007-0.012) - 0.007 (0.006-0.008) Dorsal humuli -Total length 0.031 (0.027-0.039) 0.035 (0.030-0.038) 0.034 (0.030-0.036) 0.022 (0.020-0.024) -Shaft length 0.020 (0.017-0.027) 0.022 (0.019-0.026) 0.020(0.019-0.021) 0.015 (0.012-0.016) -Point length - - - 0.005 (0.004-0.006) Dorsal bar -Total length 0.046 (0.043-0.055) 0.058 (0.055-0.068) 0.053 (0.050-0.058) 0.054 (0.043-0.063) -Width 0.012 (0.011-0.018) 0.012 (0.009-0.015) - 0.006 (0.005-0.007) Copulatory organ Length - - 0.073 (0.064-0.078) 0.069 (0.063-0.082) in length and 0.030–0.034 (0.032±0.002) mm in width, the closed circles. The dorsal hamulus had indistinct roots posterior (fourth) chamber was prolonged by a sclerotized and measured 0.020-0.024(0.022±0.002) mm, the shaft tube. Haptor was broad, differentiated from the rest of the length was 0.012-0.016(0.015±0.002) mm, the point length body, measured 0.17–0.21 (0.19±0.02) mm and provided was 0.004-0.006(0.005±0.002) mm. The ventral hamulus with continuous rows of squamodiscs. Two pairs of with distinct roots, the total length of this hamulus lateral hamuli, three bars and 14 marginal hooklets were was 0.024-0.029(0.027±0.002) mm, shaft length was observed. Squamodiscs small with rodlets, the central 0.010-0.014 (0.012±0.002) mm, the point length was rodlets often disturbed not forming regular rows, the other 0.004-0.006 (0.005±0.002) mm, inner root length was rodlets forming regular rows, these rows never formed 0.015-0.018 (0.016±0.002) mm and the outer root length

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Figs. 9-16: Photomicrographs of the adult Pseudorhabdosynochus chlorostigma sp.nov. infecting the gills and skin of brawn spotted reef cod Epinephelus chlorostigma: 9 The anterior prohaptor area of the adult worm (PR), the two pairs of eyes (E) and pharynx region (PH). 10-16 High magnifications of: 10 Quadriloculateorgan (Q) and the copulatory organ (CO). 11 Opisthohaptor consisting of squamodiscs (S), two pairs of hamuli, one pair of dorsal hamuli (DH) and one pair of ventral hamuli (VH), ventral bar (VB) as one piece and one pair of dorsal bars (DB). 12 Dorsal bar (DB), dorsal hamulus (DH), Ventral bar (VB) and ventral hamulus (VH). 13 Dorsalhamulus (DH) and ventral hamulus (VH). 14 Dorsal bar (DB). 15 Ventral bar (VB) and the two dorsal bars (DB). 16A Copulatory organ (CO)

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Fig. 17II: Schematic drawing of: a Haptor of the adult of the adult Pseudorhabdosynochus sp. nov., composed of ventral bar (VB) with two ventral hamuli (VH), dorsal bars (DB) with two lateral dorsal hamuli (DH) and squamodiscs (S). b Quadriloculate organ. c Copulatory organ.

Table 2: Comparison between measurements of Pseudorhabdosynochus chlorostigma sp. nov. recorded from the current study and some of the previously described species P. manifestus P. malabaricu P. maternus Aspect Justine & Sigura 2007 Justine & Sigura 2007 Justine & Sigura 2007 Present study Total body length 0.66 (0.45 – 0.87) 0.54 (0.450 – 0.650) 0.56 (0.450 –0.690) 0.56 (0.54 – 0.58) Maximum body width 0.30 (0.17 – 0.41) 0.32 (0.240 – 0.410) 0.32 (0.290 – 0.380) 0.21 ( 0.19 – 0.23) Pharnyx length 0.05 (0.035 –0.076) 0.05 (0.043 – 0.075) 0.05 (0.040 – 0.060) 0.09 (0.07 – 0.11) Pharnyx width 0.04 (0.035 –0.058) 0.04 (0.042 – 0.060) 0.04 (0.036 – 0.045) 0.07 (0.05 – 0.09) Haptor width 0.02 (0.019 – 0.30) 0.33 ( 0.270 – 0.420) 0.29 (0.260 – 0.300) 0.28 (0.26 – 0.30) Ventral humuli -Total length 0.05 (0.053 –0.062) 0.04 (0.039 – 0.045) 0.05 (0.050 – 0.063) 0.04 ( 0.02 – 0.06) -Shaft length 0.04 (0.044 –0.053) 0.05 (0.053 – 0.063) 0.04 ( 0.041 – 0.051) 0.02 (0.01 – 0.03) -Inner root length – – – 0.013 (0.011 –0.014) -Outer root length – – – 0.017 ( 0.016 –0.018) -Point length – – – 0.008 ( 0.007 –0.009 Ventral bar -Total length 0.11 (0.108 –0.126) 0.13 ( 0.092 – 0.156) 0.14 (0.127 – 0.141) 0.90 (0.88 – 0.92) -Width 0.02 (0.016 –0.027) 0.04 (0.042 – 0.060) 0.02 (0.014 – 0.024) 0.11 ( 0.10 – 0.12) Dorsal humuli -Total length 0.048 (0.044 -0.052) 0.048 (0.045 –0.050) 0.047 (0.044 – 0.051) 0.035 ( 0.033 –0.037) -Shaft length 0.030 ( 0.027 0.033) 0.030 (0.027 –0.032) 0.029 (0.028 – 0.032) 0.021 ( 0.020 –0.022) -Point length – – – 0.006 (0.004 – 0.008) Dorsal bar -Total length 0.04 (0.074 –0.031) 0.11 ( 0.103 – 0.124) 0.10 (0.101 – 0.110) 0.38 (0.36 – 0.40) -Width 0.02 (0.020 –0.031) 0.02 (0.019 –0.031) 0.02 (0.021 – 0.028) 0.09 (0.07 – 0.11) Copulatory organ length – – – 00.071 ( 0.070 –0.072) Quadriloculate organ Length 0.050 (0.050 –0.060) 0.010 (0.015 – 0.18) 0.010 (0.017 – 0.018) 0.70 (0.50 – 0.90) was 0.006-0.008 (0.007±0.002) mm. Two dorsal bars 0.021-0.024 (0.023±0.002) mm and the maximum width were curved with flattened medial extremity and was 0.040-0.043 (0.042±0.002) mm. The copulatory cylindrical lateral extremity, its length was 0.01-0.03 organ was small, tubular, with no accessory piece (0.02±0.002) mm and 0.005-0.007 (0.006±0.002) mm as and composed of anterior cone and straight a maximum width. The ventral bar was flat, posterior tube with total length 0.063-0.082 (0.069 ±0.002) elongate with pointed ends, its total length was mm.

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Taxonomic summary anterior wall; fourth chamber ended in elongate Family: Diplectanidae Bychowsky [23] sclerotized cone, prolonged by sclerotized tube; end of Host: Parrot fish Scarus harid (F: Scaridae [20]) tube prolonged by thin unsclerotised filament of variable Infection site: Gills and skin. length. Inner length of quadriloculate organ was 0.39–0.35 Locality: Coasts of Hurghada City, Red Sea, Egypt. (0.37±0.02) mm. Material deposition: Slides were deposited in the Zoology Department museum, Faculty of Science, Cairo Taxonomic summary University, Egypt Family: Diplectanidae Bychowsky[23] Etymology: The specific name was derived from the Host: Brown spotted reef cod Epinephelus chlorostigma specific name of the fish from which the parasite was (F: Serranidae). discovered and described for the first time. Infection site: Gills and skin. Locality: Coasts of Hurghada City, Red Sea, Egypt Pseudorhabdosynochus chlorostigma (Figs. 9-16, Material deposition: Slides were deposited in the 17II, Table 2). Zoology Department museum, Faculty of Science, Cairo Body was wide with a length 0.54-0.58 (0.56±0.02) mm, University, Egypt width 0.19–0.23 (0.21±0.02) mm. Anterior region with 3 Etymology: The specific name was derived from the pairs of head organs and 2 pairs of eye-spots. Pharynx specific name of the fish from which the parasite was was subspherical with a diameter of 0.07–0.11 (0.09±0.02) discovered and described for the first time. mm in length and 0.05-0.09 (0.07±0.02) mm in width. Copulatory organ was composed of a long coiled DISCUSSION copulatory tube that terminated in a club-shaped structure which was a characteristic taxonomic structure for this Diplectanum harid sp. nov., (Table 1) species and measured 0.070–0.072 (0.071±0.002) mm. Intraspecific variations between the morphometric Haptor was differentiated from the rest of the body with data of the sclerotized parts of Diplectanum sp. recorded a width of 0.26–0.30 (0.28±0.02) mm and provided with in the present study and those registered before were similar squamodiscs, 2 pairs of lateral hamuli, 3 bars and shown in (Table. 1). The comparison was done with 14 marginal hooklets. Squamodiscs made up of rows of Diplectanum parvus from urodeta, New rodlets; central rows forming almost closed ovals; rodlets Caledonia [24], D. calathus from gerreid (Teleostei) became thinner from the centre to periphery; ventral and from Mexico and Panama [25] and Diplectanum dorsal squamodiscs were similar, but the ventral was gatunense from Eugerres brasilianus from Gatun Lake, slightly larger. Ventral hamulus with thick handle and Panama Canal Watershed [25]. The total body length distinct guard, total length was 0.02–0.06 (0.04±0.02) mm, coincides with those of D. gatunense and D. calathus but with outer root length was 0.016–0.018 (0.017±0.001) mm, larger than D. parvus. The maximum body width is smaller inner root length 0.011–0.014 (0.013±0.001) mm, point than D. calathus and coincides with all other reported length 0.007–0.009 (0.008±0.001) mm and shaft length was species. The total length of the first pair of haumli in the 0.01–0.03 (0.02±0.01) mm. Dorsal hamulus with indistinct present parasite coincides with those in D. calathusand guard, its total length was 0.033–0.037 (0.035±0.002) mm, smaller than that in the other species. Measurements of point length was 0.004-0.008 (0.006±0.002) mm and the the dorsal hamuli were smaller than those recorded in both shaft length was 0.020–0.022 (0.021±0.001) mm. Dorsal D. Parvus and D. gatunense. Meanwhile, the (lateral) bars straight, with flattened medial extremity and measurements of the ventral bar were smaller than those thick cylindrical lateral extremity, their length was of previously reported species. The maximum width 0.36–0.40 (0.38±0.02) mm, maximum width was 0.07–0.11 coincides with that in D. parvus. Furthermore, the (0.09±0.02) mm. The ventral bar was flat with slightly measurements of the dorsal bar in this Diplectanum sp. constricted median portion and blunt extremities, its were smaller than those of the previously recorded length was 0.88–0.92 (0.90±0.02) mm, maximum width was species. The morphology of the copulatory complex is 0.10–0.12 (0.11±0.001) mm. Caeca simple, terminate blindly completely different from the comparable species at level of posterior margin of vitelline field. A (small, tubular, no accessory piece, composed of anterior quadriloculate organ with four chambers, the fourth cone and straight posterior tube), morphometrically, the (posterior) one was slightly more sclerotized than the total length of the quadriloculate organ is smaller than other three anterior chambers; first chamber with very thin that in D. gatunense reported by [25]. So according to the

325 World J. Fish & Marine Sci., 6 (4): 319-327, 2014 general morphology of the worm and due to the REFERENCES significant morphological and morphometric differences of the copulatory organ, which is a strong criteria for the 1. Rhode, K., 1982. Ecology of Marine Parasites. placement the monogenean parasite of the fish S. harid University of Queensland Press, St. Lucia, within the genus Diplectanum as a new species and we Queensland, Australia, pp: 245. propose to name it Diplectanum harid sp.nov. with new 2. Morsy, K.S., H. Saady, F. Abdel-Ghaffar, host and locality records in Egypt. A.R. Bashtar, H. Mehlhorn, S.A. Quraishy and Pseudorhabdosynochus chlorostigma, (Table 2) A. Adel, 2012. A new species of the genus harbour a rich fauna of parasites, especially Heterobothrium (Monogenea: Diclidophoridae) a large number of monogeneans on their gills. Recently, parasitizing the gills of tiger puffer fish Tetraodon Justine [26] mentioned twelve species of monogeneans lineatus (Tetraodontidae). A light and scanning on the high fin , (Bloch) electron microscopic study. Parasitol. Res., off New Caledonia, including eight new species of the 110(3): 1119-1124. diplectanid Pseudorhabdosynochus sp. Four host 3. Okamoto, T., 1963. On the problems of monogenetic species previously recorded to harbor trematode infection of puffers from inland Sea of Pseudorhabdosynochus sp. on their gills; these are Japan. Suisan Zoshoku., 3: 17-29. Epinephelus merra, E. fasciatus, E. rivulatus and 4. Ogawa, K. and K. Inouye, 1997. Heterobothrium E. howlandi. In addition, E. merra, E. fasciatus and infection of cultured tiger puffer, Takifugu rubripes E. howlandi harbour an unrelated and rare species of (Teleostei: Tetraodontidae), a field study. Fish. Pseudorhabdosynochus. Intraspecific variations between Pathol., 32: 15-20. 5. Yoshinaga, T., T. Kamaish, I. Segawa, A. Kumagai, measurements of Pseudorhabdosynochus sp. in the C. Nakayasu, K. Yamano, T. Takeuchi and present work and those registered before were shown in M. Sorimachi, 2000. Hematology, histopathology and (Table 2). In the present study Pseudorhabdosynochus the monogenean Neoheterobothrium hirame sp. was characterized by the presence of a copulatory infection in anemic flounder). Fish. Pathol., 35: 131-6. organ terminated at a club shaped structure which was a 6. Yoshinaga, T., T. Kamaishi, I. Segawa, K. Yamano, characteristic structure for this species and does not H. Ikeda and M. Sorimachi, 2001. Anemia caused matched to any of the previously recorded species. by challenges with the monogenean The morphology of the sclerotized structures did not Neoheterobothrium hirame in the Japanese flounder. differ significantly from the comparable species reported Fish. Pathol., 36: 13-20. by [27]. Haptor was observed with squamodiscs, the 7. Yoshinaga, T., H.K. Tsutsumi and K. Ogawa, 2009. ventral and dorsal Squamodiscs was similar, but the Origin of the diclidophorid monogenean ventral one was slightly larger which agreed with previous Neoheterobothrium hirame Ogawa, 1999, the study of [27]. The total length and shaft length of the causative agent of anemia in olive flounder dorsal humuli in the present worm were smaller than that Paralichthys olivaceus. Fish. Sci., 75: 1167-76. reported by [27]. Meanwhile, the total length of the 8. Mushiake, K., K. Mori and M. Arimoto, 2001. ventral humuli (present study) was nearly equal to that Epizootiology of anemia in wild Japanese flounder. recorded by [27]. The total length of the ventral bar was Fish. Pathol., 36: 125-132. relatively larger in the present study and its width was 9. Nakayasu, C., T. Yoshinaga and A. Kumagai, 2002. nearly larger to that recorded by [27]. Moreover, Hematology of anemia ex-perimentally induced by measurements of the complex (dorsal) bar (the present repeated bleeding in Japanese flounder with parasite) were larger than those recorded previously. comments on the cause of flounder anemia recently The copulatroy organ morphology is different from those prevailing in Japan. Fish. Pathol., 37: 125-130. of the comparable species. Although most of the 10. Anshary, H., K. Ogawa, M. Higuchi and T. Fujii, 2001. morphological features of the present worm are similar to A study of long-term changes in summer infection the previously recorded species of levels of Japanese flounder Paralichthys olivaceus Pseudorhabdosynochus with more or less difference in with the monogenean Neoheterobothrium hirame in measurements, the completely different copulatory organ the central Sea of Japan, with an application of a is significant for the placement of this species as a new new technique for collecting small parasites from species with new host and locality records in Egypt. the gill filaments. Fish. Pathol., 36: 27-32.

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