Ultrastructure and histochemistry of the male reproductive system of the genus Callinectes Stimpson, 1860 (Brachyura: Portunidae).

We described the ultrastructure and histochemistry of the reproductive system of five Callinectes species, and evaluate the seasonal variation in weight of the reproductive system and hepatopancreas by comparing annual changes of somatic indices. The somatic indices changed little throughout the year. In Callinectes, spermatogenesis occurs inside the lobular testes and, within each lobule, the cells are at the same developmental stage. Spermatogenesis and spermiogenesis follow the same development pattern in all Callinectes studied. Mature spermatozoa are released into the seminiferous ducts through the collecting ducts. Cells of the vas deferens are secretory as evidenced by rough endoplasmic reticulum, Golgi complex, and secretory vesicles that produce the seminal fluid. The anterior vas deferens shows two portions: proximal and distal. In proximal portion (AVDp), spermatozoa are clustered and embedded in an electron-dense, basophilic glycoproteinaceous secretion Type I. In the distal portion (AVDd), the spermatophore wall is formed by incorporation of a less electron-dense glycoproteinaceous secretion Type II. The secretion Type I change to an acid polysaccharide-rich matrix that separates the spermatophores from each other. The median vas deferens (MVD) stores the spermatophores and produces the granular glycoproteinaceous seminal fluid. The posterior vas deferens (PVD) has few spermatophores. Its epithelium has many mitochondria and the PVD seminal fluid changes into a liquid and homogeneous glycoprotein. Many outpocketings in the PVD and MVD help to increase the fluid production. Overall, the reproductive pattern of Callinectes is similar to other species that produce sperm plugs. The secretions of AVD, MVD, and PVD are responsible for the polymerization that forms the solid, waxy plug in the seminal receptacle. The traits identified here are common to all Portunidae species studied so far.

Accessory glands of male reproductive system in Pseudochorthippus parallelus parallelus (Zetterstedt, 1821) (Orthoptera: Acrididae): A light and electron microscopic study.

The accessory glands of male reproductive system in insects play a significant role in the reproduction process by protecting sperm in spermatheca, preventing female to accept other males after mating and stimulating oviposition. The number, structure, and arrangement of the tubules of accessory glands can change from species to species. In this study, the accessory glands belonging the male reproductive system in Pseudochorthippus parallelus parallelus (Zetterstedt, 1821) (Orthoptera, Acrididae) were examined with stereomicroscope, light microscope, scanning (SEM), and transmission (TEM) electron microscopes at Gazi University, Faculty of Science in 2017-2019. P. parallelus parallelus is a widespread species that is located at the extending areas from Italy to the Northern Europe and also in Turkey. The accessory glands of P. parallelus parallelus' male reproductive system are composed of about 10 tubules. The tubules can be classified into two groups according to the thickness of their muscle tissues. Both groups have single layered epithelial cells with mitochondria, well-developed endoplasmic reticulum, spherical nucleus with electron dense chromatin, secretory vesicles and multivesicular bodies in their cytoplasm. In addition, apocrine type secretion is seen in epithelial cells.

Hox13 is essential for formation of a sensory organ at the terminal end of the sperm duct in Ciona.

Species-specific traits are thought to have been acquired by natural selection. Transcription factors play central roles in the evolution of species-specific traits. Hox genes encode a set of conserved transcription factors essential for establishing the anterior-posterior body axis of animals. Changes in the expression or function of Hox genes can lead to the diversification of animal-body plans. The tunicate ascidian Ciona intestinalis Type A has an orange-colored structure at the sperm duct terminus. This orange-pigmented organ (OPO) is the characteristic that can distinguish this ascidian from other closely related species. The OPO is formed by the accumulation of orange-pigmented cells (OPCs) that are present throughout the adult body. We show that Hox13 is essential for formation of the OPO. Hox13 is expressed in the epithelium of the sperm duct and neurons surrounding the terminal openings for sperm ejection, while OPCs themselves do not express this gene. OPCs are mobile cells that can move through the body vasculature by pseudopodia, suggesting that the OPO is formed by the accumulation of OPCs guided by Hox13-positive cells. Another ascidian species, Ciona savignyi, does not have an OPO. Like Hox13 of C. intestinalis, Hox13 of C. savignyi is expressed at the terminus of its sperm duct; however, its expression domain is limited to the circular area around the openings. The genetic changes responsible for the acquisition or loss of OPO are likely to occur in the expression pattern of Hox13.

Binucleation of male accessory gland cells in the common bed bug Cimex lectularius.

The insect male accessory gland (MAG) is an internal reproductive organ responsible for the synthesis and secretion of seminal fluid components, which play a pivotal role in the male reproductive strategy. In many species of insects, the effective ejaculation of the MAG products is essential for male reproduction. For this purpose, the fruit fly Drosophila has evolved binucleation in the MAG cells, which causes high plasticity of the glandular epithelium, leading to an increase in the volume of seminal fluid that is ejaculated. However, such a binucleation strategy has only been sporadically observed in Dipteran insects, including fruit flies. Here, we report the discovery of binucleation in the MAG of the common bed bug, Cimex lectularius, which belongs to hemimetabolous Hemiptera phylogenetically distant from holometabolous Diptera. In Cimex, the cell morphology and timing of synchrony during binucleation are quite different from those of Drosophila. Additionally, in Drosophila, the position of the two nuclei in the adult stage changes as a result of the mating history or the nutrient conditions; however, it remains stable in Cimex. These differences suggest that binucleation in the Cimex MAG plays a unique role in the male reproductive system that is distinct from that of Drosophila.

Rab-mediated trafficking in the secondary cells of Drosophila male accessory glands and its role in fecundity.

The male seminal fluid contains factors that affect female post-mating behavior and physiology. In Drosophila, most of these factors are secreted by the two epithelial cell types that make up the male accessory gland: the main and secondary cells. Although secondary cells represent only ~4% of the cells of the accessory gland, their contribution to the male seminal fluid is essential for sustaining the female post-mating response. To better understand the function of the secondary cells, we investigated their molecular organization, particularly with respect to the intracellular membrane transport machinery. We determined that large vacuole-like structures found in the secondary cells are trafficking hubs labeled by Rab6, 7, 11 and 19. Furthermore, these organelles require Rab6 for their formation and many are essential in the process of creating the long-term postmating behavior of females. In order to better serve the intracellular membrane and protein trafficking communities, we have created a searchable, online, open-access imaging resource to display our complete findings regarding Rab localization in the accessory gland.

Evaluation of an in vitro mouse testis organ culture system for assessing male reproductive toxicity.

BACKGROUND: Development of an in vitro system capable of producing mature sperm remains a challenging goal, with only few successes reported. Such a system, could be used to test agents for potential toxicity to the male reproductive system; to explore this, we exposed immature mouse testis fragments in culture to ethinylestradiol (EE), a well-known testicular toxicant in vivo. METHODS: Testis fragments from postnatal day 5 mice were cultured in Albumax I medium. After 24 hr of culture, fragments were treated with 0.01, 0.1 or 1 nM EE, then harvested after 20 days in culture and examined for histology or gene expression measures by quantitative PCR. RESULTS: There was substantial variability between fragments in the degree of spermatogenesis observed. The percentage of seminiferous tubules containing any dead germ cells increased as a result of EE exposure in a dose dependent fashion. This was accompanied with a decreased percentage of tubules with round spermatids. Expression of estrogen receptor 1, cytochrome P450, family 11, subfamily a, and polypeptide 1 also was reduced, depending on the dose. CONCLUSION: These gene expression changes in the testis fragments are similar to those seen after animals have been exposed to EE. Gene expression changes in testis fragments are encouraging, but the variability across samples will need to be reduced for this in vitro system to become a generally applicable method for assessing testicular toxicants.

Comparative morphology of male genitalic structures in the minute litter bugs Dipsocoromorpha (Insecta: Hemiptera: Heteroptera).

Insect male genitalia show an evolutionarily variable morphology that has proven to be valuable for both, species identifications and phylogenetic analyses at higher taxonomic levels. Accurate usage of genitalic characters in taxonomic descriptions and phylogenetic analyses depends on consistency of terminology and validity of homology hypotheses. Both areas are underdeveloped in many insect groups. We here document the morphology and advance homology hypotheses of male genitalic features for the hemipteran infraorder Dipsocoromorpha, the minute litter bugs. Genitalic structures and the pregenital abdomen in Dipsocoromorpha are strikingly modified and diverse compared to other Heteroptera. In addition to variation in the shape of phallic structures (parameres and aedeagus), minute litter bug genitalia vary in the direction and degree of asymmetry and feature a plethora of processes derived from various abdominal segments with significant variation at low taxonomic levels. Here, male genitalic structures for an extensive taxonomic sample (32 genera and 71 specimens) are documented using scanning electron and confocal microscopy, and a universal terminology for genitalic structures across minute litter bugs is established that will facilitate species discovery and evolutionary research. We conclude by proposing primary homology hypotheses across the infraorder that now can be tested in a phylogenetic framework.

Expression of ß-hexosaminidase in the male reproductive system of the lizard, Eutropis carinata. (Reptilia, Squamata) (Schneider, 1801).

Beta-hexosaminidase (Hex) is the major lysosomal enzyme associated with the event of fertilization. In this study, we have analyzed the distribution of Hex in the testis and the epididymis of the lizard, Eutropis carinata by a polyclonal antibody of ß-hexosaminidase isoform (Hex A). Presence of Hex in the epididymis was performed by Western blotting. The result reveals that Hex A is present in the epididymal epithelium, lumen as well as spermatozoa. The anatomical distribution of Hex was studied by immunohistochemical localization. The study reveals that Hex is intensely stained in the epithelium of anterior and middle regions of the epididymis, whereas, posterior epididymal epithelium shows moderate staining. In addition, seminiferous epithelium of the testis shows staining for Hex. But lumen of the testis did not show any reaction for Hex. Further, immunohistochemical localization of Hex on the spermatozoa from the testis and different regions of the epididymis revealed that the Hex from the testis did not show any staining; the epididymal epithelium is moderately localized in the spermatozoa of the anterior region and gradually increases in the intensity in the spermatozoa of the posterior region of the epididymis. This indicates that the Hex is released from the epididymal epithelium and binds to the spermatozoa, and in the lumen, it gradually increases from anterior to the posterior region of the epididymis. The result also suggests that Hex A bound to the epididymal spermatozoa originates from the epididymis and not from the testis. The regional difference in the expression of Hex in the epididymis of the lizard, E. carinata, indicates the possible site of secretion of this enzyme.

Morphological and histological structure of the male reproductive system of the water strider Gerris lacustris (Linnaeus 1758) (Gerridae, Heteroptera).

This work presents the male reproductive system morphology and histology of the water strider Gerris lacustris (Linnaeus 1758) (Gerridae, Heteroptera) using light microscopy and scanning electron microscopy. The male reproductive system of G. lacustris comprise of a pair of testes, two vasa deferentia, two seminal vesicles, an ejaculatory duct. There is no bulbus ejaculatorius and the long vas deferantia uniting to form a simple ductus ejaculatorius which is connected to the aedeagus. The testes are white colored and this cylindiric-shaped structure lies along genital abdominal segment. The testicular follicles have three different development zones (growth zone, maturation zone, differentiation zone). Each testis has two follicles, which are not lined by a common peritoneal sheath and involving many cysts arranged in a progressive order of maturation from the distal to the proximal region; spermiogenesis occurs in mature males, finishing with the organization of sperm bundles. The testes are connected to the seminal vesicles, specialized sperm storage places, by the vas deferentia.

Expression Profile of Human Fc Receptors in Mucosal Tissue: Implications for Antibody-Dependent Cellular Effector Functions Targeting HIV-1 Transmission.

The majority of new Human Immunodeficiency Virus (HIV)-1 infections are acquired via sexual transmission at mucosal surfaces. Partial efficacy (31.2%) of the Thai RV144 HIV-1 vaccine trial has been correlated with Antibody-dependent Cellular Cytotoxicity (ADCC) mediated by non-neutralizing antibodies targeting the V1V2 region of the HIV-1 envelope. This has led to speculation that ADCC and other antibody-dependent cellular effector functions might provide an important defense against mucosal acquisition of HIV-1 infection. However, the ability of antibody-dependent cellular effector mechanisms to impact on early mucosal transmission events will depend on a variety of parameters including effector cell type, frequency, the class of Fc-Receptor (FcR) expressed, the number of FcR per cell and the glycoslyation pattern of the induced antibodies. In this study, we characterize and compare the frequency and phenotype of IgG (CD16 [FcγRIII], CD32 [FcγRII] and CD64 [FcγRI]) and IgA (CD89 [FcαR]) receptor expression on effector cells within male and female genital mucosal tissue, colorectal tissue and red blood cell-lysed whole blood. The frequency of FcR expression on CD14+ monocytic cells, myeloid dendritic cells and natural killer cells were similar across the three mucosal tissue compartments, but significantly lower when compared to the FcR expression profile of effector cells isolated from whole blood, with many cells negative for all FcRs. Of the three tissues tested, penile tissue had the highest percentage of FcR positive effector cells. Immunofluorescent staining was used to determine the location of CD14+, CD11c+ and CD56+ cells within the three mucosal tissues. We show that the majority of effector cells across the different mucosal locations reside within the subepithelial lamina propria. The potential implication of the observed FcR expression patterns on the effectiveness of FcR-dependent cellular effector functions to impact on the initial events in mucosal transmission and dissemination warrants further mechanistic studies.

Probe-based confocal laser endomicroscopy (pCLE): a preclinical investigation of the male genital tract.

The aim of this study was to assess the potential of probe-based confocal laser endomicroscopy (pCLE) as a new diagnostic imaging technique for the male genital tract. For this purpose, testes, epididymides, and vasa deferentia were obtained during transsexual surgery of healthy patients (n = 10, 26-52 years). Prior to this, testes of rats (n = 10, Sprague-Dawley) and mice (n = 8, wild-type) were examined. Ex vivo tissues were investigated by pCLE after topical fluorescence staining. Images and pCLE real-time video sequences were compared to images acquired by confocal laser scanning microscopy (CLSM); this allowed the identifying of corresponding microstructures. Interestingly, the seminiferous tubules of transsexual humans contained mainly spermatogonia due to long-term estrogen treatment, whereas the seminiferous tubules of the murine and rat spermatogenesis-related cell types were differentiated. Mosaicking improved the inspection potential by wide-angle views. Similarly, the microarchitecture of the epididymis and the vas deferens was successfully visualized in situ and on a cellular level by pCLE. In summary, pCLE allows for real-time identification of relevant microstructures responsible for spermatogenesis under ex vivo conditions. Additionally, pCLE enabled to localize vital spermatozoa in the testis thus opening up new ways to improve sperm retrieval rates during assisted reproduction. Both clinically relevant experiences hold promise to introduce this diagnostic method into a clinical study, and to investigate its potential as a clinical diagnostic procedure to expedite and improve the medical situation.

Ex vivo investigations on the potential of optical coherence tomography (OCT) as a diagnostic tool for reproductive medicine in a bovine model.

Routine infertility investigations in the male and female include imaging techniques such as ultrasonography and endoscopy (fertiloscopy). However, these techniques lack the resolution to localize vital sperm or to reveal detailed morphological analysis of the oviduct which is often the cause of infertility in females. Therefore we set out to evaluate the efficiency of optical coherence tomography (OCT) as a diagnostic imaging tool for micron-scale visualization of the male and female genital tract. Using the bovine as a model, the optical features of the Telesto(TM) , Ganymede(TM) (both Thorlabs) and Niris(TM) (Imalux) OCT imaging systems were compared.

First evidence of neurons in the male copulatory organ of a spider (Arachnida, Araneae).

Spider males have evolved a remarkable way of transferring sperm by using a modified part of their pedipalps, the so-called palpal organ. The palpal organ is ontogenetically derived from tarsal claws; however, no nerves, sensory organs or muscles have been detected in the palpal bulb so far, suggesting that the spider male copulatory organ is numb and sensorily blind. Here, we document the presence of neurons and a nerve inside the male palpal organ of a spider for the first time. Several neurons that are located in the embolus are attached to the surrounding cuticle where stresses and strains lead to a deformation (stretching) of the palpal cuticle on a local scale, suggesting a putative proprioreceptive function. Consequently, the male copulatory organ of this species is not just a numb structure but likely able to directly perceive sensory input during sperm transfer. In addition, we identified two glands in the palpal organ, one of which is located in the embolus (embolus gland). The embolus gland appears to be directly innervated, which could allow for rapid modulation of secretory activity. Thus, we hypothesize that the transferred seminal fluid can be modulated to influence female processes.

Equine sperm-neutrophil binding.

When mares are inseminated repeatedly, protein molecules from the seminal plasma (SP) prevent sperm-neutrophil binding and reduced fertility. The molecule(s) responsible for sperm-neutrophil binding is not known and the identification of beneficial SP proteins is complicated by their large numbers and abundant variation. We examined several important aspects of sperm-neutrophil binding to ultimately facilitate the identification and isolation of the molecule(s) responsible. First, we raised anti-equine P-selectin antibodies to determine the involvement of this adhesion molecule in sperm-neutrophil binding. While these antibodies identified equine P-selectin, they did not inhibit sperm-neutrophil binding. However, acrosome-reacted equine sperm expressed a molecule similar to the ligand recognition unit of P-selectin. Second, we attempted to characterize SP protein binding to equine sperm and gauge their affinity. Biotinylated SP proteins were incubated with fresh sperm, washed over a viscous medium, electrophoresed, and probed with avidin. Several SP proteins bound to sperm with a strong affinity to withstand these treatments. This finding may prove valuable for future attempts to identify and characterize specific SP molecules. Lastly, we compared the secretions from male sex organs/glands on sperm motility, sperm-neutrophil binding, and their protein profile. We expected fewer proteins from individual organs/glands, which would facilitate isolation and identification of target molecules. While each secretion had a varying effect on motility and sperm-neutrophil binding, the protein profile was as complex as that seen in whole SP, indicating that collection of proteins from individual sources will not facilitate this work. Together, these experiments answer several important questions related to sperm-neutrophil binding, sperm-SP proteins interaction, and the complexity of the SP proteome.

Mitochondrial prohibitin and its ubiquitination during crayfish Procambarus clarkii spermiogenesis.

Prohibitin (PHB), an evolutionarily conserved mitochondrial membrane protein, is associated with spermatogenesis and sperm quality control in mammals. It is identified as a substrate of ubiquitin and thus may function via a mitochondrial ubiquitin-proteasome pathway. In this study, we examined the localization of PHB during spermiogenesis of the macrura crustacean Procambarus clarkii. We traced phb mRNA's temporal and spatial expression pattern in spermiogenesis, and found its localization highly coherent with acrosome formation and nuclear shaping, two key events during crustacean spermiogenesis. We further detected the associations of PHB with mitochondria and ubiquitin using immunofluorescent staining. PHB was co-localized with mitochondria through spermiogenesis. PHB as well as mitochondria were co-localized with ubiquitin from the late stage of spermiogenesis, and the co-signals reached their peak in the mature sperm. The results raise the hypothesis that PHB is likely to function in nuclear shaping and acrosome formation in the spermiogenesis of P. clarkii. In addition, it might possess a more profound role in mediating mitochondrial ubiquitination. For the first time this study uncovers the role of PHB in the spermiogenesis of macrura crustacean species.

Isoform-specific functions of Mud/NuMA mediate binucleation of Drosophila male accessory gland cells.

BACKGROUND: In standard cell division, the cells undergo karyokinesis and then cytokinesis. Some cells, however, such as cardiomyocytes and hepatocytes, can produce binucleate cells by going through mitosis without cytokinesis. This cytokinesis skipping is thought to be due to the inhibition of cytokinesis machinery such as the central spindle or the contractile ring, but the mechanisms regulating it are unclear. We investigated them by characterizing the binucleation event during development of the Drosophila male accessory gland, in which all cells are binucleate. RESULTS: The accessory gland cells arrested the cell cycle at 50 hours after puparium formation (APF) and in the middle of the pupal stage stopped proliferating for 5 hours. They then restarted the cell cycle and at 55 hours APF entered the M-phase synchronously. At this stage, accessory gland cells binucleated by mitosis without cytokinesis. Binucleating cells displayed the standard karyokinesis progression but also showed unusual features such as a non-round shape, spindle orientation along the apico-basal axis, and poor assembly of the central spindle. Mud, a Drosophila homolog of NuMA, regulated the processes responsible for these three features, the classical isoform Mud(PBD) and the two newly characterized isoforms Mud(L) and Mud(S) regulated them differently: Mud(L) repressed cell rounding, Mud(PBD) and Mud(S) oriented the spindle along the apico-basal axis, and Mud(S) and Mud(L) repressed central spindle assembly. Importantly, overexpression of Mud(S) induced binucleation even in standard proliferating cells such as those in imaginal discs. CONCLUSIONS: We characterized the binucleation in the Drosophila male accessory gland and examined mechanisms that regulated unusual morphologies of binucleating cells. We demonstrated that Mud, a microtubule binding protein regulating spindle orientation, was involved in this binucleation. We suggest that atypical functions exerted by three structurally different isoforms of Mud regulate cell rounding, spindle orientation and central spindle assembly in binucleation. We also propose that Mud(S) is a key regulator triggering cytokinesis skipping in binucleation processes.

Evolutionary morphology of the male reproductive system, spermatozoa and seminal fluid of spiders (Araneae, Arachnida)--current knowledge and future directions.

The male reproductive system and spermatozoa of spiders are known for their high structural diversity. Spider spermatozoa are flagellate and males transfer them to females in a coiled and encapsulated state using their modified pedipalps. Here, we provide a detailed overview of the present state of knowledge of the primary male reproductive system, sperm morphology and the structural diversity of seminal fluids with a focus on functional and evolutionary implications. Secondly, we conceptualized characters for the male genital system, spermiogenesis and spermatozoa for the first time based on published and new data. In total, we scored 40 characters for 129 species from 56 families representing all main spider clades. We obtained synapomorphies for several taxa including Opisthothelae, Araneomorphae, Dysderoidea, Scytodoidea, Telemidae, Linyphioidea, Mimetidae, Synotaxidae and the Divided Cribellum Clade. Furthermore, we recovered synspermia as a synapomorphy for ecribellate Haplogynae and thus propose Synspermiata as new name for this clade. We hope that these data will not only contribute to future phylogenetic studies but will also stimulate much needed evolutionary studies of reproductive systems in spiders.

Comparative morphology of spermatozoa and reproductive systems of zorapteran species from different world regions (Insecta, Zoraptera).

The male and female reproductive apparatus of Zorotypus magnicaudelli (Malaysia), Zorotypus huxleyi (Ecuador) and Zorotypus weidneri (Brazil) were examined and documented in detail. The genital apparatus and sperm of the three species show only minor differences. The testes are larger in Z. magnicaudelli. Z. huxleyi lacks the helical appendage in the accessory glands. A long cuticular flagellum is present in Z. magnicaudelli and in the previously studied Zorotypus caudelli like in several other species, whereas it is absent in Z. weidneri, Z. huxleyi, Zorotypus hubbardi, Zorotypus impolitus and Zorotypus guineensis. Characteristic features of the very similar sperm are the presence of: a) two dense arches above the axoneme; b) a 9 + 9+2 axoneme with detached subtubules A and B of doublets 1 and 6; c) the axonemal end degenerating with enlarging accessory tubules; d) accessory tubules with 17 protofilaments; e) three accessory bodies beneath the axoneme; and f) two mitochondrial derivatives of equal shape. The first characteristic (a) is unknown outside of Zoraptera and possibly autapomorphic. The sperm structure differs distinctly in Z. impolitus and Z. hubbardi, which produce giant sperm and possess a huge spermatheca. The presence of the same sperm type in species either provided with a sclerotized coiled flagellum in males or lacking this structure indicates that a different organization of the genital apparatus does not necessarily affect the sperm structure. The flagellum and its pouch has probably evolved within Zoraptera, but it cannot be excluded that it is a groundplan feature and was reduced several times. The fossil evidence and our findings suggest that distinct modifications in the genital apparatus occurred before the fragmentation of the Gondwanan landmass in the middle Cretaceous.

Dynamics of cell and tissue genesis in the male reproductive system of ticks (Acari: Ixodidae) Amblyomma cajennense [corrected] (Fabricius, 1787) and Amblyomma aureolatum (Pallas, 1772): a comparative analysis.

Ticks are classified into three families: Argasidae, Ixodidae, and Nutalliellidae. The taxonomy and phylogeny within Ixodidae are still discussed by the specialists, thus requiring further studies. Amblyomma cajennese and Amblyomma aureolatum (Brazil) belong to two species complexes known as "cajennese" and "ovale", respectively, and are directly related to the transmission of the Brazilian spotted fever. This confirms the medical and veterinary significance of these species, as well as the need for further morphological studies that will bring a better understanding of their taxonomy, phylogeny, and control. In this context, the present study aimed to characterize the morphology of the male reproductive system of A. cajennese and A. aureolatum when unfed and after 4 days of feeding, thereby seeking to: (a) distinguish the two species or "complexes", and (b) study an internal system which has the potential to be targeted by acaricides. Therefore, males from both species (unfed and after 4 days of feeding) were cold-anesthetized, dissected, and had their reproductive systems removed for histological analysis. The results showed that the morphology of the male reproductive system is generally similar between both species, like in other Ixodidae ticks, exhibiting a multilobed accessory gland complex related to seminal fluid secretion, a pair of vasa deferentia and a pair of testes housing germ cells (spermatocytes) in different stages. The main differences were found in the development of the accessory gland complex cells and germ cells, showing that the maturation of the male reproductive system starts later in A. aureolatum, when compared to A. cajennese. However, during the blood meal, A. aureolatum development is increased, thus making germ cell maturation and gland complex activity higher than in A. cajennese. This study shows the differences in the development of the male reproductive systems of both species, while providing information that can assist in the establishment of new control methods.

Androgen receptor function links human sexual dimorphism to DNA methylation.

Sex differences are well known to be determinants of development, health and disease. Epigenetic mechanisms are also known to differ between men and women through X-inactivation in females. We hypothesized that epigenetic sex differences may also result from sex hormone functions, in particular from long-lasting androgen programming. We aimed at investigating whether inactivation of the androgen receptor, the key regulator of normal male sex development, is associated with differences of the patterns of DNA methylation marks in genital tissues. To this end, we performed large scale array-based analysis of gene methylation profiles on genomic DNA from labioscrotal skin fibroblasts of 8 males and 26 individuals with androgen insensitivity syndrome (AIS) due to inactivating androgen receptor gene mutations. By this approach we identified differential methylation of 167 CpG loci representing 162 unique human genes. These were significantly enriched for androgen target genes and low CpG content promoter genes. Additional 75 genes showed a significant increase of heterogeneity of methylation in AIS compared to a high homogeneity in normal male controls. Our data show that normal and aberrant androgen receptor function is associated with distinct patterns of DNA-methylation marks in genital tissues. These findings support the concept that transcription factor binding to the DNA has an impact on the shape of the DNA methylome. These data which derived from a rare human model suggest that androgen programming of methylation marks contributes to sexual dimorphism in the human which might have considerable impact on the manifestation of sex-associated phenotypes and diseases.