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Spermatological characteristics are described for the first time in the genus Neomultitestis Machida, 1982, belonging to the family Lepocreadiidae Odhner, 1905. Adult Neomultitestis aspidogastriformis were collected from the digestive tract of Platax teira, caught off New Caledonia. The mature spermatozoon of N. aspidogastriformis is described by transmission electron microscopy (TEM). It includes two axonemes with 9 + “1” pattern of Trepaxonemata, an antero-lateral electron-dense material, attachment zones, two mitochondria, a nucleus, two bundles of parallel cortical microtubules, external ornamentation of the plasma membrane, spine-like bodies, and granules of glycogen. Certain ultrastructural characteristics of the spermatozoon, especially the antero-lateral electron-dense material, are shared by N. aspidogastriformis and all species of the superfamily Lepocreadioidea studied so far. The antero-lateral electron-dense material, present in different families of lepocreadioids, supports the monophyletic status of the superfamily. We also list several ultrastructural characteristics which characterize the mature spermatozoon of the Lepocreadioidea.

One of the main characters used in acoel taxonomy is the male copulatory organ. Despite this, ul- trastructural studies of this structure are scarce. We studied the ultrastructure of the copulatory organ in eight species of acoels belonging to the taxon Childia. Members of Childia possess a well-developed conical or cylindrical stylet-like structure composed of needles. Immunogold cytochemistry of tubulin was used to deter- mine the composition of the needles. Stylet-like struc- tures of Childia species at the ultrastructural level are basically similar. Stylet needles show intracellular differ- entiations. As shown both by ultrastructural and immu- nocytochemical methods, the stylet needles, in all species studied, are composed of long, parallel microtubules, ei- ther tightly packed or polymerized. We report unusual polymerization of microtubules, resulting in formation of a honeycomb-like structure in cross section. Variations of ultrastructure among Childia species include numbers and arrangement of stylet needles, shape of needles, nee- dle compactness, microtubule polymerization, direction of stylet growth, and presence/absence of different types of granules. The stylet-like structures are homologous within Childia, but are likely to prove nonhomologous with the other needle-like structures found in acoel copu- latory organs. Stylets in Platyhelminthes are not homolo- gous with stylet-like structures in acoels. J. Morphol.

Spermiogenesis and the sperma- tozoon were studied in Convoluta saliens, an acoel platyhelminth, by transmission electron microscopy, labelling of nuclei and immunocytochemistry of tubulin with various antibodies. Spermiogenesis involves forma- tion of a long spermatid shaft containing two axonemes. It is established that the nucleus, after a stage of elongation, does not migrate up to the distal extremity of the spermatid, and that the centriolar derivatives are located at the distal extremity of the shaft. This contrasts with the parasitic Platyhelminthes. The ma- ture spermatozoon, 180 mm in length, comprises a nuclear region, 50 mm in length, and a cytoplasmic region, with a short region of overlap. The cytoplasmic region contains two lateral axonemes with a 9 1 2 pattern of microtubules, granules of two different sizes, and two rows of longitudinal microtubules in the center. Each row consists of 5-6 singlet microtubules, with links between them. Whereas the two axonemes are labelled by antibodies against alpha, acetylated-alpha, and beta tubulin, the microtubule rows are labelled only by the anti-beta-tubulin antibody. This suggests that acetylation does not occur in this part of the cytoskel- eton, and that the epitope recognized by the anti-alpha- tubulin antibody (DM1A) is different in these units. Mol. Reprod. Dev. 52:74-85, 1999. r 1999 Wiley-Liss, Inc.

Using transmission electron microscopy and immunologic approaches with various antibodies against general tubulin and posttranslationally modified tubulin, we investigated microtubule organization during spermatogenesis in Heligmosomoides polygyrus, a species in which a conspicuous but transient microtubular system exists in several forms: a cytoplasmic network in the spermatocyte, the meiotic spindle, a perinuclear network and a longitudinal bundle of microtubules in the spermatid. This pattern differs from most nematodes including Caenorhabditis elegans, in which spermatids have not microtubules. In the spermatozoon of H. polygyrus, immunocytochemistry does not detect tubulin, but electron microscopy reveals two centrioles with a unique structure of 10 singlets. In male germ cells, microtubules are probably involved in cell shaping and positioning of organelles but not in cell motility. In all transient tubulin structures described in spermatocytes and spermatids of H. polygyrus, detyrosination, tyrosination, and polyglutamylation were detected, but acetylation and polyglycylation were not. The presence/absence of these posttranslational modifications is apparently not stage dependent. This is the first study of posttranslationally modified tubulin in nematode spermatogenesis.

The distribution of glycylated tubulin has been analyzed in different populations of stable microtubules in a digenean flatworm, Echinostoma caproni (Platyhelminthes). Two cellular types, spermatozoa and ciliated excretory cells, have been analyzed by means of immunofluorescence, immunogold, and immunoblotting techniques using two monoclonal antibodies (mAbs), AXO 49, and TAP 952, specifically directed against differently glycylated isoforms of tubulin. The presence of glycylated tubulin in the two cell types was shown. However, the differential reactivities of TAP 952 and AXO 49 mAbs with the two axoneme types suggest a difference in their glycylation level. In addition, within a single cell, the spermatozoon, cortical microtubules underlying the flagellar membrane, and axonemal microtubules were shown to comprise different tubulin isoforms, the latter ones only being labelled with one of the antiglycylated tubulin mAbs, TAP 952. Similarly, the antiacetylated (6-11B-1) and polyglutamylated (GT335) tubulin mAbs decorated the two types of axonemal microtubules, but not the cortical ones. From these data, a subcellular sorting of posttranslationally modified tubulin isoforms within spermatozoa, on the one hand, and a cellular sorting of glycylated isoforms inside the whole organism, on the other hand, is demonstrated in the flatworm E. caproni. Last, a sequential occurrence of tubulin posttranslational modifications was observed in the course of spermiogenesis. Acetylation appears first, followed shortly by glutamylation; glycylation takes place at the extreme end of spermiogenesis and, specifically, in a proximo-distal process. Thus in agreement

Spermiogenesis and sperm structure of the primitive acoel Paratomella rubra from the Ligurian Sea, Italy, were investigated by several methods. During spermiogenesis, after flagellar incorporation by formation of two longitudinal lateral grooves, spermatid elongation is characterized in Paratomellaby the presence of four membranes encircling each axoneme plus two membranes encircling both the axonemes and the nucleus. These structures were interpreted as being three cytoplasmic canals situated one inside the other. The filiform spermatozoon has two incorporated axonemes of 9+2 type, a nucleus almost as long as the sperm cell itself, a single elongate mitochondrion, and two types of membrane-bound granules, respectively, small and gastrula-shaped, and large. Organelles are highly ordered, the sperm is bilaterally symmetrical with a single long mitochondrion on the ventral side and a regular row of large granules, for some length embedded in the nucleus, on the dorsal side. Immunocytochemical studies and the use of fluorescent nuclear dyes reveal the spatial relationships of the axonemes with the nucleus. The granules were shown by Thiery, PTA and enzyme digestion tests to contain glycoproteins and/or polysaccharides and very little protein. Glycogen particles were detected in the cytoplasm. Cells containing coiled spermatozoa undergoing resorption were found in the parenchyma. New apomorphies of the taxon Paratomellabased on sperm structure are proposed: a very long nucleus, a highly bilaterally symmetrical pattern of organelles, a single long mitochondrion. These characters are not found in other Acoela, and particularly in Hesiolicium, sometimes considered closely related to Paratomella. © 1997 Published by Elsevier Science Ltd on behalf of The Royal Swedish Academy of Sciences.

Nematode spermatozoa are amoeboid cells. In Caenorhabditis elegans and Ascaris suum, previous studies have reported that sperm motility does not involve actin, but, instead, requires a specific cytoskeletal protein, namely major-sperm-protein (MSP). In Heligmosomoides polygyrus, a species with large and elongate spermatids and spermatozoa, cell organelles are easily identified even with light microscopy. Electrophoresis of Heligmosomoides sperm proteins indicates that the main protein band has a molecular weight of about 15 kDa, as MSP in other nematodes, and is specifically labelled by an anti-MSP antibody raised against C. elegans MSP. A minor band at 43 kDa was specifically labelled by an anti-actin antibody. Reaction of anti-actin and anti-MSP antibodies is specific to, and restricted to, their respective targets. Actin and MSP localisation, studied by indirect immunofluorescence in male germ cells of Heligmosomoides polygyrus, are similar: spermatids show rows of dots, corresponding to the fibrous bodies, around an unlabelled central longitudinal core; spermatozoa are labelled strictly in an anterior crescent-shaped cap, at the opposite pole to the nucleus, which contains fibres of the MSP cytoskeleton. Phalloidin labelling shows that F-actin is present in spermatids, but absent in spermatozoa. Tropomyosin shows a distinct pattern in spermatids, but is located in the MSP and actin-containing cap in spermatozoa. It is hypothesized that actin plays a role in the shaping of the cell and in the arrangement of its organelles during nematode spermiogenesis, when MSP is present, in an inactive state, in the fibrous bodies. The concentration of actin and tropomyosin in the anterior cap is not compatible with previous theories about the MSP cytoskeleton, which is supposed to act in the absence of actin.

The mature spermatozoa of four species of European decapod crabs (Clibanarius erythropus, Maja squinado, Cancer pagurusand Potamon fluviatile)have been investigated using indirect immuno‐fluorescence techniques for the presence of the cytoskeletal proteins actin and tubulin. Indirect immunofluorescence labelling with monoclonal anti‐actin antibody and three different monoclonal anti‐tubulin antibodies indicate that actin is present in the spermatozoa of all four species, but tubulins are restricted to the two species with microtubular arms, Clibanariusand Maja.The pattern of actin fluorescence varies between the spermatozoa of the four species, with Majaand Cancershowing intense fluorescence in the acrosome vesicle and in elements of the sperm cell involved in the acrosome reaction. The spermatozoon of each species is described ultrastructurally using transmission electron microscopy and correlations made between observed patterns of fluorescence and the cellular components described. No obvious filamentous actin (F‐actin) is visible in the electron micrographs of the spermatozoa of any of the species. In most cases the fluorescence is sufficiently specific to indicate in which region of the mature sperm cell the actin and tubulin occurs. Actin is acrosomal in Maja, Cancerand Potamonbut appears to be cytoplasmic in Clibanarius, while the tubulins appear only to be present in the cytoplasm of Clibanarius, Majaand Cancer. 1994 The Royal Swedish Academy of Sciences

Spermiogenesis and spermatozoa were studied by transmission and scanning electron microscopy in Troglocaridicolasp., a scutariellid epizoic on a cavernicolous freshwater shrimp. Spermiogenesis involves elongation of the spermatid in which the nucleus elongates, but remains close to the common cytoplasmic mass. Flagella first grow in opposite direction and at a right angle to the cytoplasmic shaft, and centrioles show associate structures. Later, the two centrioles rotate and the flagella emerge parallel, but still perpendicular to the shaft. An apical process elongates at the extremity of the spermatid shaft. The spermatozoon shows active flagellar beating and undulations of the sperm body. The spermatozoon comprises an anterior ‘corkscrew’ region, the flagellar insertion region, a cytoplasmic region and a posterior nuclear region. The corkscrew contains an electron dense structure, not membrane-bound, originating from the apical process of the spermatid. The flagella show the 9+‘1’ pattern, usual in Platyhelminthes. The cytoplasmic and nuclear regions show a cortical row of about 50 twisted longitudinal microtubules surrounding a row of electron dense, and not membrane-bound, 25-nm granules. These granules are original structures and seem to be known only in a few Platyhelminthes species in which a non-flagellar movement of the spermatozoon occurs. Thus, it is hypothesised that the 25-nm granules play a role in cellular motility. Sperm ultrastructure in Troglocaridicolashows major differences to that in the temnocephalids. It is therefore concluded that the phylogenetic position of the scutariellids within the Temnocephalidea should be reinvestigated.

The cytoskeletal protein actin was identified in the mature spermatozoon of the European edible crab, Cancer pagurus Linnaeus, by indirect immunofluoresce with monoclonal and polyclonal anti‐actin antibodies, fluorescent phalloidin, and DNAase I. The actin was localized in two distinct concentric rings within the acrosome vesicle of the spermatozoon and appeared to correlate with the internal zonation of the vehicle. Modifications of the fluorescent pattern for actin were observed in sperm cells which were undergoing changes associated with the acrosome reaction. In these cases, fluorescent staining was observed in the nucleocytoplasm immediately subjacent to the perforatorial column and sometimes in the perforatorial column within the acrosome vesicle. Equally intense fluorescence was observed in an apical perforatorial projection. SDS‐PAGE of C. pagurus sperm confirmed the presence of actin in the cells. A single band of actin (approximately 43 kDa) comigrated with rabbit muscle actin when immunoblotted onto nitrocellulose with mouse monoclonal anti‐actin. The actin‐associated cytoskeletal proteins α‐actinin, tropomyosin, and spectrin were also identified within the spermatozoon of C. pagurus using specific polyclonal antibodies, but their presence was not confirmed by SDS‐PAGE and immunoblotting. © 1994 Wiley‐Liss, Inc.

Justine, J.-L., Afzelius, B. A., Malmberg, G. & Mattei, X. 1993. Ultrastructure of spermiogenesis in Acanthocotyle and Myxinidocotyle (Platyhelminthes, Monogenea, Acanthocotylidae).Acta Zoologica (Stockholm) 74: 119-126. Spermiogenesis was studied by transmission electron microscopy in the acanthocotylid monogeneans Myxinidocotyle californica (from Eptatretus stoutii) and Acanthocotyle lobianchi (from Raja clavata). In Myxinidocotyle and Acanthocotyle, the zone of differentiation shows two 9+'1' axonemes, the elongating nucleus and mitochondrion, and a single cortical cytoplasmic microtubule. This single microtubule is found in the mature spermatozoon of both species and was also noted in capsalids. This requires a modified definition of 'pattern 2' of spermatozoa which becomes: 'spermatozoa with two axonemes and no cortical microtubules, except one single element much shorter than the spermatozoon'. A very unusual structure was found in Myxinidocotyle, but not in Acanthocotyle: the centriolar derivative of one of the 9+'1' axonemes is made up of 18 diverging singlets of unequal length associated with electron-dense cytoplasm. This seems to be the first case of a centriolar derivative without nine-fold symmetry associated with an axoneme with nine-fold symmetry. Jean-Lou Justine, Laboratoire de Biologie parasitaire-Protistologie-Helminthologie, URA CNRS 114, Museum national d'histoire naturelle, 61 rue Buffon, F-75231 Paris cedex 05, France.

Atriaster (Atriaster) sp., a gill parasite in the fish Diplodus cervinus caught near Dakar, Senegal, was studied by transmission electron microscopy. Spermiogenesis is similar to the general scheme in the Cercomeridae and particularly the polyopisthocotylean monogeneans, but shows two remarkable characteristics: (a) the zone of differentiation for each spermatid is separated from the common spermatid cytoplasmic mass by a region without longitudinal microtubules but with many mitochondria, herein termed the pre-zone of differentiation; (b) transverse sections of nuclei in late spermatids are polygonal with concave sides. These two characteristics are provisionally interpreted as autapomorphies, since observations in other species are lacking. The outer layer of the tegument is similar to that of other polyopisthocotylean monogeneans, and has microvilli and three kinds of cytoplasmic inclusions: granules with fibrous contents, granules with homogeneous contents, and elongate electron-dense bodies.

Rohde, K., Noury-Sraki, N., Watson, N., Justine, J.-L. & Euzet, L. 1990. Ultrastructure of the flame bulbs of Urastoma cyprinae (Platyhelminthes, 'Prolecithophora'. Urastomidae).Acta zool.. Stockh. 71: 211-216. The ultrastructure of the flame bulbs of the turbellarian Urastoma cyprinae from Mytilm galloprovincialis in the Mediterranean is described. The nucleus of the terminal cell is located some distance basal to the rootlets of the cilia forming the flame; the cytoplasm contains numerous tubules approximately 54-66 nm in diameter, and vesicles. Thick walled, densely packed rod-like structures coil around each other with a tendency towards longitudinal orientation close to the flame. The rod-like structures tightly surround the basal part of the flame and the distal cytoplasmic tube in the apical part of the flame. Some of them, including the inner predominantly longitudinally directed ones, are continuous with the cytoplasm of the terminal cell, others are continuous with the cytoplasm of the distal cytoplasmic tube. Internal leptotriches arise from the cytoplasm of the terminal cell and intrude between the basal parts of the cilia of the flame. The distal cytoplasmic tube possesses a septate junction. The flame bulb of Urastoma differs distinctly from those known from other Platyhelminthes; implications for the phylogeny of Platyhelminthes are discussed.

Comparative ultrastructural observations were carried out on the spermiogenesis of the capsalid Caballerocotyla manteri Price and the dionchid Dionchus remorae MacCallum. At the beginning of spermiogenesis the zones of differentiation (ZD) jut out in all directions. A large mitochondrion shaped like a perforated bead, and through which the elongated nucleus passes, is found facing each ZD. Later the ZD become parallel and are embedded within the common cytoplasmic mass. Cortical longitudinal microtubules are present in the ZD at the outset of spermiogenesis, but they later disappear. The spermatozoon is long and filiform. It shows two parallel axonemes of the 9 +“1” flatworm pattern, the nucleus and mitochondrion, and no cortical microtubule. The ultrastructure of spermiogenesis and spermatozoon is remarkably similar in the two species studied, as in other capsalids previously described. Two characteristics, the perforated bead shape of the spermatid mitochondrion and the progressive disappearance of the microtubules of the ZD, may be considered as synapomorphies which indicate close phylogenetic relationships between the families Dionchidae and Capsalidae. This interpretation coincides with Llewellyn's (1971) scheme of the evolution of the monogeneans.

Justine, J.-L., Ponce de Ledn, R., Mattei, X. 1987. Ultrastructural observations on the spermatozoa of two temnocephalids (Platyhelminthes). (MusCum National #Histoire Naturelle, Paris, France, Universidad de la Republica, Montevideo, Uruguay, and FacultC des Sciences, Dakar, SCnCga1.)-Acta 2001. (Stockh.) 68, 1-7. The spermatozoa of two Temnocephalidae collected in Uruguay, Temnocephala iheringi Haswell, 1893 (Host: Pomacea canaliculata) and Ternnocephala axenos Monticelli, 1899 (Host: Parastacus varicosus), were studied with a transmission electron microscope. In both species the spermatozoon is made up of a long sperm body which bears at one extremity two free flagella of the 9+'1' flatworm pattern. The sperm body contains the nucleus, mitochondria, dense bodies and parallel, cortical, longitudinal singlet microtubules. Along a part of the sperm body the palissade of the microtubules displays a spiral pattern in transverse sections. A part of the perimeter of the cell is thus lined by two overlapping rows of microtubules. This spiral pattern of the singlets is considered as a synapomorphy of the family Temnocephalidae. The singlet microtubules are interconnected by two kinds of links: tangential links between neighbouring singlets in the same row and radial links between singlets belonging to two rows. The presence of these links suggests that this structure could be a motile system of singlets.

Observations were performed in the uterus of a female Gonapodasmius sp., a gonochoristic didymozoid Trematode. The oocyte is a round cell 6 μm in diameter, which shows a ‘nucleolus-like cytoplasmic body’ and cortical granules. The spermatozoon is filiform, mobile and about 50 μm long. There is no acrosome. The anterior tip of the spermatozoon contains two centrioles made up of singlets and cortical microtubules with associated glycocalyx. The centrioles are continued as two axonemes of the classical 9 + ‘1’ pattern of flatworms, accompanied by a mitochondrion and a short row of cortical longitudinal microtubules. It is the posterior part of the sperm cell which contains the nucleus. At the outset of fertilization, the anterior part of the spermatozoon coils around the oocyte and penetrates it by lateral fusion. The posterior region of the spermatozoon, with the nucleus, is the last part to enter the oocyte, after passing through a perforation in the forming eggshell. The whole spermatozoon thus penetrates the female cell.

Spermiogenesis in Urastoma cyprinae (Graff. 1882) involvcs a progressive lengthening of the spermatid. Free flagella are only transitory. The mature spermatozoon is fusibrm. 45μm in length and 2.5 μm in width. It contains two incorporated axonermes of the flatworm 9+“1” pattern, two elongated mitochondria. an elongated nucleus and a row or cortical longitudinal microtubules. Observations on oogenesis concerncd only the immuturc ovary. lmmature oocytes contain few dense granules and accssory cells were not ohserved. Phylogenetic implications of a biflagellate spermatozoon in a Prolecithophora are important. The presence of two 9 +“1” axonemes confirms that Urastoma (and the Prolecithophora) belongs to the taxon Trepaxonemata Ehlers. Previous electron microscope studies on spermitozoa of Prolecithophora (four genera) only dealt with aflagellate spermatozoa. On this basis, Ehlers (1985) proposed two autapomorphics for the taxon Prolecithophora: aflagellate spermatozoon and spermatozoal mitochondrial derivatives with abundant membranes. The present observations on Urastoma contradict these two autapomorphics. The taxon Prolecithophora cannot be defined by autapomorphies of the spermatozoon.

Ultrastructural observations are presented for some of the stages occurring during fertilization in Dionchus remorae (a gill parasite of Echeneis naucrates) and are believed to be the first published concerning a monogenean. Fertilized female germ cells were found in the ovary. Several loops of the spermatozoon were present within the oocyte cytoplasm; the sperm nucleus became electron lucent and the parallel peripheral doublets of the axonemes became increasingly divergent. The cortical granules in the oocyte were not released immediately after penetration by the spermatozoon. The homogeneity apparently found in the oocyte ultrastructure and process of fertilization in the monogeneans and digeneans contrasts with the variety that exists in their sperm ultrastructure.

Three species of the genus Paravortex were studied. Spermiogenesis involves progressive lengthening of the spermatid. No centriole nor axoneme were found. Thc mature spermatozoon is threadlike, 30μm long and aflagellate. It contains a nucleus, two longitudinal parallel mitochondria, numerous dense bodies limited by a membrane, a row of cortical microtubules and granules. The spatial arrangement of the microtubules wah deduced from a statistical analysis of their number in transverse section. The microtubules are continuous along the length of the spermatozoon and are arranged along a long cone. The granules, not limited by a membrane, are about 25 nm in diameter. They are arranged in a semicircle and represent a new structure for sperm of Platyhclminthes. A statistical comparison demonstrated that two species may be distinguished by the number of their cortical sperm microtubules. This is the tirst ultrastructural study of an aflagellate spermatozoon in the Rhabdocoela. A rickettsia was found in P. cardii.

In Loimosina, during spermiogenesis, the zone of differentiation of the spermatid contains two centrioles continued as two 9 +‘1’axonemes. One of the axonemes lengthens and will become the principal axoneme of the spermatozoon. The other axoneme is as long as the first one at the beginning of spermiogenesis, but is shorter in the mature sperm cell. The spermatozoon consists of several regions: (a) at the anterior end, the centriolar derivative of the principal axoneme; (b) a short region which shows cortical microtubules coinciding with external ornamentations; (c) a long uniflagellate region, with mitochondrion; (d) a biflagellate region containing the anterior slender part of the nucleus; (e) the posterior part of the nucleus, with no accompanying cytoplasmic organelles. Spermiogenesis and sperm structure in Loimosina differ from what is known in all other described monogeneans, excepting the monocotylid Heterocofyle to which they show close resemblances. However, the alteration of the second axoneme is more complete in Heterocofyle than in Loimosina. Comparative study of spermiogenesis and sperm ultrastructure thus shows an interesting coincidence with classical phylogenies of the monogeneans, in which the families Loimoidae and Monocotylidae are closely related. Chez Loimosina, pendant la spermiogenese, la zone de differenciation de la spermatide contient deux centrioles prolonges par deux axonemes de type 9 +‘1′. L'un des axonemes s'allonge et deviendra l'axoneme principal du spermatozoide mur. Le deuxieme axontme est aussi long que le premier au debut de la spermiogenese, mais il est plus court dans le spermatozoide mur. Le spermatozoide comprend: (a) a l'avant, le derive centriolaire de l'axoneme principal; (b) une courte region contenant quelques microtubules corticaw longitudinaux coincidant avec des omementations extramembranaires; (c) une longue region miflagellee avec mitochondrie; (d) une region biflagellee contenant La partie anterieure effilee du noyau; (e) la region posterieure du noyau, sans organites cytoplasmiques accompagnateurs. La spermiogenese et la structure du spermatozoide de Loimosina sont differentes de ce qui est connu chez tous les autres Monogenes decrits, excepte le Monocotylidae Heterocofyle auquel elles ressemblent beaucoup. Toutefois, l'alteration du deuxieme axoneme est moins complete chez Loimosina que chez Heterocotyle. L'etude comparee des spermatozoides et des spermiogeneses montre une bonne coincidence avec les phylogeneses classiques, dans lesquelles les familles Loimoidae et Monocotylidae sont proches.

The spermatozoon of Toxorhynchites brevipalpis conradti Gruenberg is long and filiform. It contains an anterior nucleus, an axoneme of the 9 + 9 +“1” pattern and two mitochondrial derivatives. Comparison of our observations with bibliographical data shows that spermatozoon ultrastructure is homogeneous in the family Culicidae (Culicinae, Anophelinae and Toxorhyn-chitinae). The 9 + 9 +“1” pattern of the sperm axoneme may be considered a synapomorphy of the Culicidae.