Tissue and Cell 42 (2010) 137–150 Contents lists available at ScienceDirect Tissue and Cell journal homepage: www.elsevier.com/locate/tice Spermatogenesis in the blue swimming crab, Portunus pelagicus, and evidence for histones in mature sperm nuclei Michael J. Stewart a,∗ , Praphaporn Stewart c , Nantawan Soonklang c , Vichai Linthong d , Peter J. Hanna b,d , Wei Duan a , Prasert Sobhon d,∗∗ a School of Medicine, Deakin University, Geelong, Victoria 3217, Australia b Faculty of Science and Technology, Deakin University, Geelong, Victoria 3217, Australia c Department of Preclinical Science, Faculty of Medicine, Thammasat University, Pathumthani 12121, Thailand d Department of Anatomy, Faculty of Science, Mahidol University, Bangkok 10400, Thailand article info Article history: Received 9 February 2010 Received in revised form 17 March 2010 Accepted 19 March 2010 Available online 21 April 2010 Keywords: Portunus pelagicus Spermatogenesis Spermatozoa Ultrastructure Basic nuclear proteins abstract Spermatogenesis in the blue swimming crab, Portunus pelagicus, is described by light and electron microscopy. The testis is composed of anterior (AT) and posterior (PT) lobes, that are partitioned into lobules by connective tissue trabecula, and further divided into zones (germinal, transformation and evacuation), each with various stages of cellular differentiation. The vas deferens is classified into three distinct regions: anterior (AVD), median (MVD), and posterior (PVD), on the presence of spermatophores and two secretions, termed substance I and II. Based on the degree and patterns of heterochromatin, sper- matogenesis is classified into 13 stages: two spermatogonia (SgA and SgB), six primary spermatocytes (leptotene, zygotene, pachytene, diplotene, diakinesis, and metaphase), a secondary spermatocyte (SSc), three spermatids (St 1–3), and a mature spermatozoon. Spermatid stages are differentiated by chromatin decondensation and the formation of an acrosomal complex, which is unique to brachyurans. Mature spermatozoa are aflagellated, and have a nuclear projection and a spherical acrosome. AUT-PAGE and Western blots show that, during chromatin decondensation, there is a reduction of most histones, with only small amounts of H2B and H3 remaining in mature spermatozoa. © 2010 Elsevier Ltd. All rights reserved. 1. Introduction The morphology of the testis and vas deferens, including sperm and spermatophore formation, has been described in several crustacean species using light microscopy. These species include Callenectus sapidus (Cronin, 1947) Panulirus versicolor (Talbot and Summers, 1978), Scylla serrata (Uma and Subramoniam, 1979), Scyllarus chacei (Hinsch and Mcnight, 1988), Chiononectes. opilio (Beninger et al., 1988), Thenu orientalis (Burton, 1995), Lithodes maja (Tudge et al., 1998), Aristaeomprha foliacea (Desantis et al., 2003), Goniopis cruenata (Garcia and Silva, 2006), and Maja brachydactyla (Simeó et al., 2009). Sperm (spermatozoa) are produced through spermatogenesis in a manner similar to that which occurs in vertebrates. Spermato- genesis begins with proliferation of spermatogonia and formation of primary spermatocytes. These then pass through meiosis into secondary spermatocytes, which differentiate into spermatids ∗ Corresponding author. ∗∗ Corresponding author. Tel.: +66 2 2015406; fax: +66 2 3547168. E-mail addresses: mjstewa@deakin.edu.au (M.J. Stewart), scpso@mahidol.ac.th (P. Sobhon). and through spermiogenesis, finally mature sperm (Fasten, 1918; Shigekawa and Clark, 1986; Nagao and Munehara, 2003; Okumura and Hara, 2004; Garcia and Silva, 2006). Mature sperm are then transported to the vas deferens, and become encapsulated into spermatophores (Hinsch, 1988; Diesel, 1989; Chiba et al., 1992; Simeó et al., 2009). There have been numerous ultrastructural studies in a variety of decapod species (Langreth, 1969; Lynn and Clark, 1983; Pearson and Walker, 1975; Hinsch, 1988; Chiba et al., 1992; Medina and Rodríguez, 1992; Jamieson, 1994; Jamieson et al., 1997; Jamieson et al., 1998; Jamieson and Tudge, 2000; Kang et al., 2008; Kurtz et al., 2009). Early observations indicated that sperm are in a non-compact organization, with chromatin at different degrees of decondensation, and suspended in the nucleoplasm of a nucleus that is not appreciably reduced in volume (Hinsch, 1986; Tudge et al., 2001). In contrast, more recent studies imply that the final sperm nucleus is reduced in volume and possesses very compact chromatin. Arguably this is partly due to an increasing amount of sperm basic nuclear proteins (SNBP’s), particularly histones, pro- tamines and protamine-like proteins (Manochantr et al., 2003; Kurtz et al., 2008, 2009; Tudge, 2009). Nevertheless, other inves- tigations report that the sperm of most decapods have a nucleus that is generally cup-shaped and extends into radial arms or projec- 0040-8166/$ – see front matter © 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.tice.2010.03.002