Vitellogenic Oocytes Research Articles

The first evidence of intersexuality in Atka mackerel Pleurogrammus monopterygius (Hexagrammidae).

Histological examination of Atka mackerel ovotestes reveals the possibility of intersexuality. Individuals with bisexual gonads have been caught in the North Pacific near the south-east coast of the Kamchatka peninsula. While intersex appeared to be normal females with developing ovaries, histological analysis showed the presence of both female and male tissue in the same gonad. Specifically, primary growth, cortical alveolar and primary vitellogenic oocytes were located among spermatogonia cysts. The prevalence of intersexuality in the population was less than 0.1%.

Gonadal cycle of the blue crab Portunus segnis (Forskål, 1775) (Decapoda, Portunoidea) in the Mediterranean Sea off Alexandria, Egypt

Abstract Adults of Portunus segnis were captured from Ras el Tin beach on the Mediterranean Sea at Alexandria, Egypt, from January to December 2017. Male crabs are present in higher density in the sea during fall and winter, while female crabs are more common during spring and summer. Sex ratios were calculated as follows: number of males or females/total number of crabs. The calculated sex ratios were subjected to a Chi-square test according to the equation: , where O represents the observed frequency and E the expected frequency. One-way analysis of variance shows differences in the carapace size (length and width) and the weight of males and females during the four seasons. The hepatosomatic index of male crabs ranges from 9.1-9.25 in winter months, 8.7-7.9 in fall, being lower, 3.12-2.25, in spring and 5.96-4.13 in summer. The hepatosomatic index of female crabs is 8.9-3.9 in spring, 9.2-7.9 in summer, shifts lower to 5.1-3.2 in fall and to 4.2-1.5 in winter. The gonadosomatic index of female crabs is 0.12-8.2 in spring, 0.20-13.9 in summer and decreases to 0.66-7.8 in fall and 0.18-4.7 in winter. Ovaries and testes were isolated, fixed, and prepared for histological examination. The testicular lobes are surrounded by simple epithelium with haemolymph filling the interspaces. Seminiferous ducts are inserted in the testicular lobes. Spermatogenesis and spermiogenesis are described. The vas deferens is usually filled with spermatophores. Spermatophore formation begins in the testis near the vas deferens. Near the base of the penes there is glandular tissue. This study proposes the classification of the ovarian maturation into five stages: Stage I, Immature ovary with primary and secondary oogonia; Stage II, early maturing ovary with germ cells in different maturation phases; Stage III, mature ovary with vitellogenic oocytes and germ cells; Stage IV, maturing ovary with oocytes I, II and fully mature oocytes; and Stage V, ovary in regeneration. The wall of the spermatheca consists of a stratified squamous epithelium.

Gonadal development, reproductive investment and fecundity of Aegla castro Schimitt, 1942 (Crustacea, Anomura)

ABSTRACT Aegla castro, is an endemic anomuran crab from Brazil found in headwater streams of the Atlantic Forest on the Paranapanema River basin. This study aims to evaluate the gonadal maturity of A. castro based on macroscopic and microscopic analyses. Crabs were collected in Mauá da Serra (23º57’ S; 51º06ʹ W), Paraná, Brazil in the winter and spring of 2015, and summer, autumn, and spring of 2016. Individuals had their gonads analysed and categorized into stages of development. Stage1, ovaries are macroscopically undetectable through abdominal transparency, while ovaries of stages 2 and 3 could be detected through abdominal transparency and have characteristic colour and length in relation to the abdominal somites. Ovaries have pre-vitellogenic and vitellogenic oocytes in all stages of development, with germ cells and structural epithelium characteristic of these gonadal maturation phases. Spermatogenesis in males begins in spring, with the production of spermatozoa in autumn and spermiation in winter. Aegla castro has seasonal reproduction. The macroscopic stages of gonadal development correspond to the histological gonadal maturation, indicating that the macroscopic observation of the gonads of live animals is feasible in studies of the reproductive cycle of aeglids.

Scanning Electron Microscopy of Vitellogenic Oocytes and Spawned Eggs of the Portunid Crab Charybdis hellerii (Crustacea-Brachyura) (Milne Edwards, 1867).

Background:The portunid crab Charybdis hellerii is an economically critical aquatic species in the Mediterranean region. Several investigators have reported scanning electron microscopy (SEM) observations on some crustacean's eggs' morphology. Going through the previous studies, knowledge regarding the morphology of C. hellerii. vitellogenic oocytes and spawned egg membranes are not available.Aims and Objectives:In the present study, an attempt has been made to describe the morphology and the structure of the membranes of vitellogenic oocytes and the newly spawned eggs to provide necessary information for further studies on the reproductive and evolutionary biology of the crab C. hellerii.Materials and Methods:Samples of ripe pinkish orange ovaries of non-ovigarous females and the spawned incubated eggs of ovigerous females with orange and grey spawns were processed for scanning electron microscopy. The prepared samples were examined in a Zeiss DSM 940 scanning electron microscope.Results:The present SEM study revealed that, vitellogenic oocytes are highly packed with yolk inclusions, which appear to be embedded in a definite acellular matrix and surrounded by a distinct chorion, which is pierced by several pores. The follicle cells appear polygonal in shape and interconnected through thin lateral projections and strongly associated with vitellogenic oocytes. The brooded fertilized eggs are attached through a marked stalk (funiculus) and surrounded by three distinct envelopes, which showed specific ornamentations and variations in their surface topography. The outer envelope coarsely wrinkled, while the middle envelope showed finely wrinkled ornamentation, and the inner envelope appeared with its characteristic spongy, porous appearance.Conclusions:This study denotes a significant difference between mature vitellogenic oocytes inside the ripe ovary and the spawned ova. The differences have been shown in the structure and external ornamentation of their surrounding membranes. Unlike the vitellogenic oocytes, the spawned ova were surrounded by three distinct layers, which are differ in their surface architecture. Such membrane architecture is species specific characteristic and has been thought to be an adaptive feature for brooded fertilized eggs to survive from stressful environmental conditions.

Synchrony between Reproductive Phase and Flood Period: A Dispersion Mechanism for the Freshwater Clam Corbicula fluminea (Bivalvia: Corbiculidae) in a Brazilian Neotropical Floodplain.

This study evaluates the gonadal histology of Corbicula fluminea present in the upper Paraná River floodplain and samples of limnological variables to understand its reproductive cycle. Corbicula fluminea was monitored monthly from December 2013 to February 2015. Spermatogonia, primary and secondary spermatocytes, spermatids and spermatozoa were identified in the male follicles of the hermaphrodites. Oogonia, oogonial nests, previtellogenic oocytes, early vitellogenic oocytes, middle vitellogenic oocytes and full-grown vitellogenic oocytes were identified in the female follicles of the hermaphrodites and females. The reproductive phases were described as developing, active spawning/sperm releasing, regression and regeneration. Higher values of temperature, dissolved oxygen, total nitrogen and total phosphorous were identified during flood periods, while higher values of pH and conductivity were obtained during dry periods. The species either does not reproduce or reduces the intensity of reproduction in cold months, with the sex ratio not differing significantly between hermaphrodites and females with regard to month and reproductive phase. Thus, reproduction is synchronized with the flood period and its limnological characteristics and when the increase in connectivity between floodplain environments facilitates the larval dispersion of this non-native species into other environments.

Molecular cloning, characterisation and expression analysis of the vitellogenin genes vtgAo1 and vtgC during ovarian development in Chinese hook snout carp Opsariichthys bidens.

Vitellogenesis is essential for oocyte maturation. Vitellogenin (Vtg), a yolk precursor protein, plays an important role in oogenesis and vitellogenesis. Chinese hook snout carp Opsariichthys bidens is an economically important freshwater fish in China whose reproductive and developmental biology are not well understood. In this study, we undertook histological analysis to examine ovary development and oogenesis in O. bidens. The ovaries were divided into Stages II-V and oocytes were divided into perinuclear oocytes, cortical alveoli oocytes, vitellogenic oocytes and mature oocytes. Full-length cDNA sequences were cloned of two vtg genes from the liver of O. bidens, namely Ob-vtgAo1 and Ob-vtgC. Ob-vtgAo1 and Ob-vtgC cDNA are made up of 4136 and 4392 bases respectively and encode proteins containing 1335 and 1250 amino acids respectively. Ob-vtgAo1 contains three yolk protein domains: lipovitellin heavy chain (LvH), phosvitin (Pv) and lipovitellin light chain (LvL), whereas Ob-VtgC contains LvH and LvL, which are incomplete Vtgs. Ob-vtgAo1 and Ob-vtgC mRNA expression was significantly higher in the liver of O. bidens than in all other tissues. In oocytes of Stage II-III ovaries, yolk granules are almost absent and ovarian and hepatic Ob-vtgAo1 and Ob-vtgC expression is low. At Stage IV, the oocyte is filled with yolk granules and ovarian and hepatic Ob-vtgAo1 and Ob-vtgC expression is significantly increased. Collectively, these findings help us better understand vitellogenesis in O. bidens.

Histological Evidence of Multiple Spawning in Wild Female Japanese Eel Anguilla japonica.

The post ovulatory follicle (POF) is an important and reliable tissue structure used to investigate the spawning history in teleost fish. Fresh POFs shortly after spawning are comprised of cellular (follicular cells) and acellular (basement membrane and fibrils such as elastic fibers) components. The cellular components are quickly disintegrated by means of apoptosis, while the acellular components persist for a longer period. Since cellular components are well visualized by conventional hematoxylin-eosin (HE) staining but acellular components are not stained well, old POFs that have lost cellular components are difficult to identify. In this study, periodic acid-Schiff and Victoria blue staining, which can distinctly visualize acellular POF components, were applied to the ovarian tissues of Japanese eel (Anguilla japonica) (n = 9) captured from June to August of 2008, 2009, and 2013 at the southern West Mariana Ridge, a spawning area for Japanese eels. Only new POFs were observed in seven females caught in June, and these females had ovaries with early-to mid-vitellogenic stage oocytes. Both fresh and old POFs were observed in a female caught in July, and only mid-vitellogenic stage oocytes were observed. Only old POFs and no vitellogenic stage oocyte were observed in a female caught in August. A progressive decrease in muscle lipid content, gonad somatic index, and condition factors was observed from June to August. Thus, the female Japanese eel can spawn at least twice or three times at most during spawning season, depending on energy reserve.

GROWTH PERFORMANCE AND GONAD MATURATION OF AMUR CATFISH, Silurus asotus IN CAPTIVITY

This study was aimed to determine the growth performance and gonad maturation of Amur catfish, Silurus asotus reared in captivity. A total of 600 juvenile Amur catfish were reared in a 20-tonne aerated fibreglass tank for 3 months. The changes to body weight (mean ± SD) of Amur catfish at day 14 was 22.59 ± 3.59 g and attained a final average body weight of 110.95 ± 19.72 g. The average body weight gains (%) were 213.50% and gradually decreased to 109.95%. The specific growth rate recorded at 14-day intervals was 5.42%/day and fluctuated to 3.32%/day, 0.39%/day, 0.51%/day, with the corresponding measurements of 15.36 ± 1.07 cm and increments to 24.22 ± 1.34 cm. The final total length the fishes attained was an average of 25.58 ± 1.95 cm. The total length gain recorded was 124.61% and followed by 109.95%. The survival of Amur catfish within the three months was 29.17%. Length-weight relationship of the Amur catfish showed a strong correlation at log W = -2.6797 + 2.3102 log L and the value of the regression coefficient (b) equalled to 2.3102. Three stages of the oocyte (PO = primary oocyte, PVO = previtellogenic oocyte and VO = vitellogenic oocyte) determined histologically indicated that all-female gonad samples were in the developing stage. Matured gonad and spermatozoa were also observed in males. This study concludes that Amur catfish attained good growth performance and fast gonad maturation in captivity and can be a suitable species for aquaculture.

Insulin-like androgenic gland hormone 1 (IAG1) regulates sexual differentiation in a hermaphrodite shrimp through feedback to neuroendocrine factors

Insulin-like androgenic gland hormone (IAG) is regarded as a key sexual differentiation regulator in gonochoristic crustaceans. However, until now the knowledge concerning its functions in hermaphroditic crustaceans is scanty. Herein, we investigated the function of IAG (Lvit-IAG1) in peppermint shrimp Lysmata vittata, a species that possesses protandric simultaneous hermaphroditism (PSH) reproductive system, which is rare among crustaceans. Lvit-IAG1 was exclusively expressed in the androgenic gland. The qRT-PCR demonstrated that its mRNA expression level was relatively high at the functional male phase but decreased sharply in the subsequent euhermaphrodite phase. Both the short-term and long-term silencing experiments showed that Lvit-IAG1 negatively regulated both the gonad-inhibiting hormone (Lvit-GIH) and crustacean female sex hormone (Lvit-CFSH) expressions in the eyestalk ganglion. Besides, Lvit-IAG1 gene knockdown induced a retarded development of the appendices masculinae (AM) and male gonopores while suppressing the germ cells at the primary spermatocyte stage. Also, Lvit-IAG1 gene silencing hindered ovarian development. This in turn led to small vitellogenic oocytes and decreased expression of vitellogenin and vitellogenin receptor genes in hepatopancreas and ovarian region, respectively. Generally, this study’s findings imply that Lvit-IAG1 modulated the male sexual differentiation in PSH species L. vittata, and exhibited negative feedback on Lvit-GIH and Lvit-CFSH genes expression in the species’ eyestalk ganglion.

Effects of waterborne exposure to environmentally relevant concentrations of selenite on reproductive function of female zebrafish: A life cycle assessment.

Recently, bioaccumulation of dietary organic selenium (Se) in the ovaries and inhibition of reproduction in female aquatic animals have been reported. However, there is limited data on the subtle reproductive impacts of waterborne exposure to inorganic Se in fish. Here, zebrafish embryos (2h post-fertilization) were exposed to solutions with environmentally relevant levels of Na2SeO3 with concentrations of 0 (control), 7.98±0.31, 25.14±0.15, and 79.60±0.81μg Se/L for 120d until they reached sexual maturity. Female zebrafish were selected for reproductive toxicity assessment. In the early embryonic stage, whole-mount in situ hybridization of zebrafish embryos showed that waterborne Na2SeO3 exposure did not affect the observed location of vasa expression in primordial germ cells at 24, 48, and 72h post-fertilization. Life-cycle exposure to 25.14±0.15 and 79.60±0.81μg Se/L Na2SeO3 did not change the testosterone and 17β-estradiol contents in female zebrafish at the endpoint of exposure, but significantly reduced the proportion of early vitellogenic oocytes and mature oocytes. Follicle maturity retardation was accompanied by changes in transcriptional levels of the genes related to the hypothalamus-pituitary-gonad-liver (HPGL) axis. Transcriptional levels of cyp19a and lhr in the ovary were down-regulated, while the transcriptional level of fshr in the ovaries was up-regulated. In the 21-day cumulative spawning experiment, Na2SeO3 (25.14±0.15 and 79.60±0.81μg Se/L) caused fewer eggs to be produced. Additionally, the malformation of zebrafish offspring significantly increased in the group exposed to 79.60±0.81μg Se/L. In conclusion, for the first time, this study shows that life-cycle exposure to environmentally relevant concentrations of waterborne Na2SeO3 significantly delays ovarian maturation and reduces the fertility of the female zebrafish.

Loss of the glucocorticoid receptor causes accelerated ovarian ageing in zebrafish.

Reproductive decline in mid-adult females is an established phenotype of the ageing process. Stress and the rise in glucocorticoids (GCs) accelerate reproductive ageing, but little is known about the mechanisms involved. During stress, GCs activate the glucocorticoid receptor (GR), a ubiquitously expressed, ligand-bound transcription factor, to elicit physiological changes for restoring homeostasis. Here, we tested the hypothesis that GC-GR signalling is essential for accelerating reproductive ageing. To test this, we used a ubiquitous GR knockout (GRKO) zebrafish, which is inherently hypercortisolemic, to delineate the role of high cortisol and GR signalling on reproductive ageing. The loss of GR led to premature ovarian ageing, including high frequency of typical and atypical follicular atresia in vitellogenic oocytes, yolk liquefaction and large inflammatory infiltrates. The reduction in oocyte quality was also associated with a decline in ovarian tert expression in the adult GRKO fish compared to the early adult GRKO and adult wild-type zebrafish. Accelerated ovarian ageing also impacted the progeny, including lower breeding success, fecundity, egg fertilization rate and delayed somitogenesis and embryo survival in the adult GRKO fish. We adduce that GR signalling is essential for prolonging the reproductive lifespan and improving the egg quality and embryo viability in zebrafish.

Identification of a novel germ cell marker MnTdrd from the oriental river prawn Macrobrachium nipponense.

Germ cell-specific genes play an important role in establishing the reproductive system in sexual organisms and have been used as valuable markers for studying gametogenesis and sex differentiation. Previously, we isolated a vasa transcript as a germ cell marker to trace the origin and migration of germ cells in the oriental river prawn Macrobrachium nipponense. Here, we identified a new germ cell-specific marker MnTdrd RNA and assessed its temporal and spatial expression during oogenesis and embryogenesis. MnTdrd transcripts were expressed in high abundance in unfertilized eggs and embryos at cleavage stage and then dropped significantly during late embryogenesis, suggesting that MnTdrd mRNA is maternally inherited. In situ hybridization of ovarian tissue showed that MnTdrd mRNA was initially present in the cytoplasm of previtellogenic oocyte and localized to the perinuclear region as the accumulation of yolk in vitellogenic oocyte. Whole-mount in situ hybridization of embryos showed that MnTdrd-positive signals were only localized in one blastomere until 16-cell stage. In the blastula, there were approximately 16 MnTdrd-positive blastomeres. During embryonized-zoea stage, the MnTdrd-positive cells aggregated as a cluster and migrated to the genital rudiment which would develop into primordial germ cells (PGCs). The localized expression pattern of MnTdrd transcripts resembled that of the previously identified germ cell marker vasa, supporting the preformation mode of germ cell specification. Therefore, we concluded that MnTdrd, together with vasa, is a component of the germ plasm and might have critical roles in germ cell formation and differentiation in the prawn. Thus, MnTdrd can be used as a novel germ cell marker to trace the origin and migration of germ cells.

Ultra‐diluted bioactive compounds enhance energy storage and oocyte quality during gonad conditioning of Pacific calico scallop Argopecten ventricosus (Sowerby II, 1842)

Ultra-diluted bioactive compounds have emerged as an alternative to enhance marine organism performance and to improve broodstock gonad maturation. This study evaluated for the first time the effect of three homeopathically ultra-diluted bioactive compounds (HUBC’s) authorized by the health authority as medicines for humans, on the general reproductive physiology of Argopecten ventricosus, a commercially important scallop species. The experiment included three HUBC’s treatments at centesimal dilution assessed in triplicate under laboratory conditions, T1: PaV-30C + PhA-30C, T2: PaV-30C + SiT-30C, and T3: ViT-30C, and a non-treated field reference group (FRG) maintained in aquaculture facilities as reference. The bioenergetics of scallop reproduction and oocyte quality increased with the three HUBC treatments, but particularly in those treated with T2. The organisms treated with T3 showed not only the highest proliferation of vitellogenic and post-vitellogenic oocytes but also the greatest oxidative damage in tissues and oocytes. In contrast, the scallops from T1 and T2 showed less oxidative damage. The results suggest that ultra-diluted formula T2 is a potential alternative to optimize A. ventricosus gonad ripening, food-nutrient management and oocyte quality because although T3 allowed accelerating the gametogenesis process, it increased oxidative damage. This study could help to improve gonad broodstock condition and larval yield of this species in hatcheries.

Exposure to Boscalid Induces Reproductive Toxicity of Zebrafish by Gender-Specific Alterations in Steroidogenesis.

Boscalid is a succinate dehydrogenase inhibitor fungicide and is frequently detected in surface water. Due to the frequent detection of boscalid, we evaluated its impact on the reproduction of adult zebrafish following a 21 d exposure to 0, 0.01, 0.1, and 1.0 mg/L. Following exposure to boscalid, the fertility of female zebrafish and fertilization rate of spawning eggs were reduced in a concentration-dependent manner up to a respective 87% and 20% in the highest concentration. A significant 16% reduction in the percentage of late vitellogenic oocytes was noted in ovaries, and a significant 74% reduction in the percentage of spermatids in testis was also observed after treatment with 1.0 mg/L. 17β-Estradiol (E2) concentrations decreased significantly in females (34% decrease) but significantly increased in males (15% increase) following 1.0 mg/L boscalid treatment. The expression of genes (such as era, er2b, cyp19a, and cyp19b) related to the hypothalamus-pituitary-gonad-liver (HPGL) axis was significantly altered and positively correlated with E2 concentrations in female and male zebrafish (p < 0.05). Molecular docking results revealed that the binding modes between boscalid and target proteins (ER and CYP19) of zebrafish were similar to that of the reference compounds and the target proteins. The binding energies indicate that boscalid may have a weak estrogen-like binding effect or CYP19 inhibition, potentially altering the HPGL axis, thereby reducing E2 concentrations and fecundity in females. In contrast, boscalid caused significant induction of E2 steroidogenesis and subsequent feminization of gonads in males, indicating gender-specific adverse outcome pathways.

The IAG-Switch and Further Transcriptomic Insights Into Sexual Differentiation of a Protandric Shrimp

The insulin-like androgenic gland hormone (IAG) secreted from the androgenic gland (AG) is a unique endocrine controller (the IAG-switch) of crustacean sexual differentiation. However, while previous studies of the IAG-switch focused mainly on sexual differentiation at early developmental stages of gonochoristic species, this mechanism is yet to be deciphered during naturally occurring sexual shifts in hermaphrodite species. The Northern spot shrimp, Pandalus platyceros, is a protandric hermaphrodite species, native to the North Pacific Ocean. We collected four stages of spot shrimp in Southeast Alaska including: juveniles, adult males, transitionals and adult females. The AG was dissected from each stage and characterized histologically. Additionally, the IAG mRNA was sequenced. The function of the IAG-switch during the life history of this protandric species was demonstrated through monitoring IAG gene expression. Transcript levels were highest at the juvenile stage, then decreased significantly in mature males and became negligible in the transitional and female stages. Moreover, manipulating the IAG-switch via IAG loss of function in males through RNAi, induced the expected masculine to feminine sexual transformation that naturally occurs in this species. This included reduction in IAG transcript levels in males, elevation of vitellogenin gene expression in hepatopancreas and transformation of the gonad from an ovotestis containing both ovarian and testicular tissue to a true ovary with vitellogenic oocytes. Furthermore, a transcriptomic library from tissues associated with the endocrine axis upstream and downstream the IAG-switch yielded 1,801, 1,707, 1,946 and 182 differentially expressed genes between males, transitionals and females in the AG, eyestalk, gonad and hepatopancreas, respectively. Among these genes, the transcriptional pattern of six of them (all of them in the AG), between males, transitionals and females, had similar or inverted transcriptional pattern to that of IAG in P. platyceros. Five of these putatively IAG-switch associated genes are downregulated and one is upregulated throughout the P. platyceros shift from maleness to femaleness. Homolog protein sequences for the above novel genes were found in 17 other decapod species suggesting that they might represent conserved factors associated with the IAG-switch and involved in universal crustacean sexual differentiation mechanisms.

Potential role of Methoprene-tolerant (Met) in methyl farnesoate-mediated vitellogenesis in the Chinese mitten crab (Eriocheir sinensis).

Methoprene-tolerant (Met) belongs to the basic helix-loop-helix (bHLH)-Per-Arnt-Sim (PAS) family of nuclear transcriptional regulators and is a leading candidate receptor for juvenile hormone (JH III) in insects. Methyl farnesoate (MF) is a de-epoxide form of JH III that regulates many developmental processes in crustaceans, including reproduction, molting, and morphogenesis, much like JH III in insects. In this study, the full-length cDNA for Met was cloned from the Chinese mitten crab (Eriocheir sinensis) (EsMet). The amino acid sequence of EsMet contains three conserved domains (bHLH, PAS-A, and PASB) characteristic of the bHLH-PAS family, having six conserved amino acid residues specifically responsible for JH or MF binding. Tissue distribution analysis revealed that EsMet mRNA is highly expressed in the hepatopancreas. In addition, EsMet and EsVg expression in the hepatopancreas were found to be significantly increased in early endogenous vitellogenic oocytes (stage II) during ovarian development, and the hemolymph MF titer was significantly increased in late exogenous vitellogenic oocytes (stage III), indicating that EsMet is involved in vitellogenesis regulation. In vitro, MF addition markedly upregulated EsMet and EsVg expression in hepatopancreatic tissue, but only EsVg was induced in ovarian tissue. In vivo, EsMet and EsVg expression in the hepatopancreas were both significantly and synchronously increased after MF injection, but not in the ovaries. In addition, EsMet and EsVg expression were upregulated in the hepatopancreas after eyestalk ablation, while only EsVg expression was induced in the ovaries. Thus, our results indicate that Met may act as a receptor for MF in MF-mediated vitellogenesis in crustaceans.

Microorganization of ovaries and oogenesis of Haplotaxis sp. (Clitellata: Haplotaxidae).

Ovaries of Haplotaxis sp. were studied in active and nonactive states, that is, in a sexually mature specimen and in specimens outside of the reproductive period. Two pairs of ovaries were found in segments XI and XII. Especially in the nonactive state, they were in close contact with copulatory glands. Each ovary was composed of germ cells interconnected with syncytial cysts, which were enveloped by a layer of somatic cells. Within cysts each germ cell had one ring canal connecting it to the common anuclear cytoplasmic mass called a cytophore. During oogenesis clustering germ cells differentiated into nurse cells and oocytes; thus, the oogenesis was recognized as meroistic. Vitellogenic oocytes were detached from the ovaries and continued yolk absorption within the body cavity. Because recent studies have shown the variety of ovaries and germ line cyst organization in clitellates and suggest their evolutionary conservatism at the family or subfamily level, the data presented here can be valid in understanding the phylogenetic relationships among Clitellata. In this context, ovaries found in Haplotaxis sp. resembled those of the "Tubifex" type. "Tubifex" ovaries are characteristic for numerous microdrile taxa (tubificines, limnodriloidines, propappids, lumbriculids, and leech-like branchiobdellids) and can be regarded as the primary character for these Clitellata in which germ-line cysts are formed during early oogenesis. As the family Haplotaxidae is currently considered to be paraphyletic and the species studied here belongs to Haplotaxidae sensu stricto, our results support the close relationship of Haplotaxidae sensu stricto to the clade consisting of Lumbriculidae, Branchiobdellida, and Hirudinida, in which lumbriculids are sister to the latter two.

Changes induced by atrazine in Clarias gariepinus provide insight into alterations in ovarian histoarchitecture and direct effects on oogenesis

Clarias gariepinus juveniles were exposed to environmentally relevant concentrations of 0 (control), 2.5, 25, 250 and 500 μg L−1 atrazine in a quality-controlled 28-day laboratory procedure. Findings revealed a significant decrease in the levels of follicle-stimulating hormone, luteinizing hormone and prolactin relative to control (p < 0.05). Atrazine reduced the levels of testosterone while increasing the concentration of progesterone. Histologically, the control and treatments presented three stages of oocyte maturation: the chromatin nucleolar oocyte stage, early perinucleolar oocyte stage and the vitellogenic oocyte stage. However, in the ovaries of the treatment group with the lowest treatment concentration (2.5 μg L−1), atretic oocytes with broken membranes invaded many of the dead ova and empty spaces. In other treatments (25, 250 and 500 μg L−1), interfollicular spaces, vacuolation in oocyte formation, and dissolution of oocyte walls were observed. Disruption of the yolk vesicle and clumping of the cytoplasm in maturing oocytes was observed only at the highest atrazine concentration (500 μg L−1). Gross alterations in ovarian histoarchitecture and reproductive hormone levels observed in this study showed interference with oogenesis which may result in reduced egg viability and fecundity in fish with ecological implications in water bodies exposed to atrazine even at reduced concentrations.

An ultrastructural study of the ovary cord organization and oogenesis in the amphibian leech Batracobdella algira (Annelida, Clitellata, Hirudinida)

This study reveals the ovary micromorphology and the course of oogenesis in the leech Batracobdella algira (Glossiphoniidae). Using light, fluorescence, and electron microscopies, the paired ovaries were analyzed. At the beginning of the breeding season, the ovaries were small, but as oogenesis progressed, they increased in size significantly, broadened, and elongated. A single convoluted ovary cord was located inside each ovary. The ovary cord was composed of numerous germ cells gathered into syncytial groups, which are called germ-line cysts. During oogenesis, the clustering germ cells differentiated into two functional categories, i.e., nurse cells and oocytes, and therefore, this oogenesis was recognized as being meroistic. As a rule, each clustering germ cell had one connection in the form of a broad cytoplasmic channel (intercellular bridge) that connected it to the cytophore. There was a synchrony in the development of the clustering germ cells in the whole ovary cord. In the immature leeches, the ovary cords contained undifferentiated germ cells exclusively, from which, previtellogenic oocytes and nurse cells differentiated as the breeding season progressed. Only the oocytes grew considerably, gathered nutritive material, and protruded at the ovary cord surface. The vitellogenic oocytes subsequently detached from the cord and filled tightly the ovary sac, while the nurse cells and the cytophore degenerated. Ripe eggs were finally deposited into the cocoons. A comparison of the ovary structure and oogenesis revealed that almost all of the features that are described in the studied species were similar to those that are known from other representatives of Glossiphoniidae, which indicates their evolutionary conservatism within this family.

Gonadal development and intersex condition of marbled lungfish, Protopterus aethiopicus (Heckel, 1851), in contaminated sites in Lake Victoria, Uganda

The first evidence of the impact of environmental contamination on populations of Protopterus aethiopicus (Heckel, 1851) in terms of body indices, cellular development (histology) and intersex condition in Lake Victoria, Uganda is provided. Specimens were obtained from the more polluted swampy areas receiving wastes from residential and industrial settings (Ggaba, n = 11; Port Bell, n = 10; and Jinja, n = 7), and rural, less polluted areas (Bukakata, n = 3; Kasensero, n = 4; and Bale, n = 2), and gonadosomatic indices, growth condition factor and the morphometric body characteristics were determined. Fish specimens in less polluted sites had better condition and shapes, regardless of sex, compared with those in more polluted sites of the lake. Most lungfish collected had a total length of 45–55 cm. Irrespective of sex of fish and level of pollution at collection sites, samples (n = 26) collected inshore (1 063.42 g ± 109.32 SE) were heavier than those collected (n = 11) offshore (680.09 g ± 108.85 SE), F (1, 35) = 6.288, p = 0.019. Mean values of gonadosomatic indices were significantly higher in less polluted sites, compared with chemically contaminated urban sites (F (5, 31) = 2.783, p = 0.034), suggesting better growth performance in the former. Spermatogenic cell development progressed as spermatogonia, spermatocytes, spermatids and spermatozoa among males. For oogenic cells, development was through chromatin nucleolar, perinucleolar, cortical alveolar, early vitellogenic and late vitellogenic oocytes. Histological examinations revealed group asynchronous gonadal development and intersex condition among P. aethiopicus populations in Lake Victoria.