1296 J. Parasitol., 93(6), 2007, pp. 1296–1302  American Society of Parasitologists 2007 ULTRASTRUCTURE OF THE SPERMATOZOON OF PARACHRISTIANELLA TRYGONIS DOLLFUS, 1946 (TRYPANORHYNCHA: EUTETRARHYNCHIDAE) Jordi Miquel, Zdzislaw S ´ widerski*†, Lassad Neifar‡, and Catarina Eira§ Laboratori de Parasitologia, Departament de Microbiologia i Parasitologia Sanita `ries, Facultat de Farma `cia, Universitat de Barcelona, Av. Joan XXIII, sn, E-08028 Barcelona, Spain. e-mail: jordimiquel@ub.edu ABSTRACT: The ultrastructural organization of the mature spermatozoon of the trypanorhynch cestode Parachristianella trygonis is described by transmission electron microscopy. The spermatozoon is a long and filiform cell, tapered at both ends, lacking both mitochondrion and crested bodies. Its cytoplasm contains 2 axonemes of the 9+‘1’ pattern of the Trepaxonemata longitu- dinally displaced, parallel cortical microtubules, the nucleus and glycogen in form of both -glycogen rosettes and -glycogen particles. The anterior extremity of the spermatozoon is characterized by the presence of an arclike row of up to 10 parallel cortical microtubules that partially surround the first axoneme. The present study emphasizes the ultrastructural similarity between mature spermatozoa of all 4 trypanorhynchs that have been studied to date. Nevertheless, several features, i.e., the characteristics of spermatozoa extremities, the absence of crested bodies, and the possible presence of an arclike layer of cortical microtubules, need a more thorough analysis or confirmation in some of these species. The Trypanorhyncha comprises species of cestodes infecting elasmobranchs as final hosts. This order has been considered one of the most chaotic and confusing tapeworm groups (see Wardle and McLeod, 1952; Campbell and Beveridge, 1994). Nevertheless, a new classification of the order has been recently developed by Palm (2004), in which the tentacular armature was considered to be of limited importance for the classification of higher taxa. According to this author, the order comprises 254 species, with 15 families distributed into 5 superfamilies (Tentacularioidea, Gymnorhynchoidea, Lacistorhynchoidea, Otobothrioidea, and Eutetrarhynchoidea). The morphology of the spermatozoon was not considered in this classification of trypanorhynchs. However, it could provide useful morphologi- cal characters for the phylogeny/classification of trypanorhynch cestodes, as has been demonstrated for other taxa (see Euzet et al., 1981; S ´ widerski, 1986; Justine, 1991, 1998, 2001; Ba ˆ and Marchand, 1995; Hoberg, Mariaux et al., 1997; Hoberg et al., 2001; Olson et al., 2001). Among the Eucestoda, there are ultrastructural studies on the spermatozoa of species belonging to 12 of the 14 orders of Eucestoda recognized by Khalil et al. (1994). These include the Caryophyllidea, Spathebothriidea, Pseudophyllidea, Haploboth- riidea, Diphyllidea, Trypanorhyncha, Tetraphyllidea, Lecani- cephalidea, Proteocephalidea, Nippotaeniidea, Tetrabothriidea, and Cyclophyllidea (see Justine, 2001; Ndiaye, 2003). With re- spect to the Trypanorhyncha, the available data on ultrastruc- tural aspects of the spermatozoon are limited to 3 species be- longing to 2 of the 5 superfamilies, the Lacistorhynchoidea and Eutetrarhynchoidea. These include the lacistorhynchids Lacis- torhynchus tenuis (S ´ widerski, 1976, 1994) and Grillotia eri- Received 16 January 2007; revised 25 April 2007; accepted 27 April 2007. * W. Stefan ´ski, Institute of Parasitology, Polish Academy of Sciences, 51/55 Twarda Street, 00-818 Warsaw, Poland. † Department of General Biology and Parasitology, Warsaw Medical University, Chalubin ´skiego 5, 02-004 Warsaw, Poland. ‡ De ´partement des Sciences de la Vie, Faculte ´ des Sciences de Sfax, BP802, 3018 Sfax, Tunisia. § SPVS-Estac ¸a ˜o de Campo de Quiaios, Apartado 16-EC Quiaios, 3081- 101 Figueira da Foz, Portugal.  Departamento de Biologia, Universidade do Minho, Campus de Gual- tar, 4710-057 Braga, Portugal. naceus (McKerr, 1985) and the eutetrarhynchid Dollfusiella spi- nulifera (Miquel and S ´ widerski, 2006). The aim of present study is to describe the ultrastructure of the mature spermatozoon of another trypanorhynch cestode, Parachristianella trygonis Dollfus, 1946, thus providing the first data on this genus and increasing the available data for the Eutetrarhynchidae. MATERIALS AND METHODS Adult specimens of Parachristianella trygonis Dollfus, 1946 were removed from the spiral valve of the common stingray, Dasyatis pas- tinaca (Linnaeus, 1758), collected at Sfax and Zarzis (southern coast of Tunisia). The live cestodes were placed in a 0.9% NaCl solution. After dissection, different portions of mature proglottides were routinely processed for transmission electron microscope (TEM) examination. Thus, they were fixed in cold (4 C) 2.5% glutaraldehyde in a 0.1 M sodium cacodylate buffer at pH 7.2 for a minimum of 2 hr, rinsed in a 0.1 M sodium cacodylate buffer at pH 7.2, postfixed in cold (4 C) 1% osmium tetroxide in the same buffer for 1 hr, rinsed in a 0.1 M sodium cacodylate buffer at pH 7.2, dehydrated in an ethanol series and pro- pylene oxide, and finally embedded in Spurr’s resin. Ultrathin sections were obtained using a Reichert-Jung Ultracut E ultramicrotome, placed on copper grids, and double stained with uranyl acetate and lead citrate according to Reynolds (1963). Ultrathin sections were examined using a JEOL 1010 TEM operated at an accelerating voltage of 80 kV. The Thie ´ry (1967) technique was used to emphasize the presence of glycogen particles. Gold grids were treated in periodic acid, thiocar- bohydrazide, and silver proteinate (PA-TCH-SP) as follows: 30 min in 10% PA, rinsed in distilled water, 24 hr in TCH, rinsed in acetic solu- tions and distilled water, 30 min in 1% SP in the dark, and rinsed in distilled water. RESULTS The ultrastructure of the mature spermatozoon of P. trygonis is illustrated in Figures 1–16. The observation of several lon- gitudinal and cross-sections by means of TEM has enabled to establish 5 (I–V) different regions in the mature spermatozoon. The male gamete is a long filiform cell, tapered at both ends, which lacks mitochondria. The cytoplasm contains (1) 2 lon- gitudinally displaced trepaxonematan axonemes of different lengths, (2) 2 short rows of parallel cortical microtubules, (3) the nucleus, and (4) glycogen in form of both -glycogen ro- settes and -glycogen particles. Region I (Figs. 1, 5, 16I) corresponds to the anterior extrem- ity of the spermatozoon. Its anterior tip contains a small number