Structure of the male reproductive system of the blue swimmer crab Portunus pelagicus (Decapoda: Portunidae)

An attempt was made here to study the structure of the male reproductive system of Portunus pelagicus, which would improve the knowledge base on the reproductive biology of the species and also help in the maintenance of broodstock under controlled conditions. Male P. pelagicus of different sizes were collected from the Palk Bay off Mandapam (9°17′ N, 79°9′ E) and maintained under controlled conditions for the study. Tissues from testis, anterior vas deferens (AVD), median vas deferens (MVD), posterior vas deferens (PVD), ejaculatory duct and penis were fixed in Bouin's fluid and 2.5% buffered glutaraldehyde separately and processed for light and electron microscopic studies, respectively. The reproductive system consisted of testis, commissure, vas deferens, ejaculatory duct and penis. The vas deferens was divided based on the morphology and/or histology into AVD, MVD and PVD. The AVD was further divided based on histology into proximal and distal regions, and the MVD, based on diameter into major and minor coils. The testicular lobe had several lobules with a central seminiferous tubule, which continued till the penis. The seminiferous tubule was lined by a layer of cuboidal or columnar epithelium. The lining of the central tubule of the vas deferens formed several ‘folds’, which at times formed ‘pouches’. High incidence of cell organelles in the columnar epithelial cells, aggregations of vesicles and occurrence of blebs at the luminal periphery and the projection of numerous microvilli containing electron‐dense materials into the lumen from the cell lining denoted high secretory activity of the epithelial cells.     

Fatty acid compositional changes during the embryonic development of the swimming crab,Portunus pelagicus (Portunidae: Decapoda)

Abstract

The fatty acid composition, moisture, and total lipid of the eggs from the swimming crab, Portunus pelagicus, at three different embryonic stages (within 24 h, during the eye placode stage and the final heart beat stage), were measured. Results showed that the moisture and lipid content significantly increased and decreased (p < 0.05), respectively, as the stages progressed. The most prevalent fatty acids that were initially deposited included C16:0, C18:1n-9, and C18:0, while the most consumed fatty acids were C22:5n-6, C22:5n-3, and C20:1n-7. Among the major fatty acid groups, polyunsaturated fatty acids (PUFA) and long-chain PUFA (LC-PUFA) were consumed more than saturated fatty acids and significantly more (p < 0.05) than monounsaturated fatty acids (p < 0.05). Meanwhile, n-3 PUFA was deposited in significantly higher amounts (p < 0.05) than n-6 PUFA, but both were consumed at similar amounts at 43.4% and 41.3%, respectively. The relatively low amount of C20:5n-3 and C22:6n-3 consumption may indicate these fatty acids were conserved, while the essential fatty acids C18:3n-3 and C18:3n-6 were consumed at high amounts. These findings may have implications for broodstock nutrition in order to formulate a well-balanced diet.     

Isolation and characterization of microsatellite loci in Portunus pelagicus (Crustacea: Portunidae)

Seven dinucleotide and one tetranucleotide microsatellite loci were isolated and characterized, using routine protocols, from the blue swimmer crab, Portunus pelagicus. These loci will be useful for investigating the population structure of this broadly distributed and economically important portunid. In particular, all eight loci were polymorphic in P. pelagicus collected from throughout Australia. In addition, the genotype frequencies at each locus in each population sample were usually not significantly different from those expected under Hardy–Weinberg equilibrium conditions. Six of the eight loci also showed moderate to high levels of polymorphism in an undescribed species of Portunus from northern Australia.     

Induced thermal stress on serotonin levels in the blue swimmer crab,Portunus pelagicus

The temperature of habitat water has a drastic influence on the behavioral, physiological and biochemical mechanisms of crustaceans. Hyperglycemia is a typical response of many aquatic animals to harmful physical and chemical environmental changes. In crustaceans increased circulating crustacean hyperglycemic hormone (CHH) and hyperglycemia are reported to occur following exposure to several environmental stress. The biogenic amine, serotonin has been found to modulate the CHH levels and oxidation of serotonin into its metabolites is catalysed by monoamine oxidase. The blue swimmer crab, Portunus pelagicus is a dominant intertidal species utilized throughout the indo-pacific region and is a particularly important species of Palk bay. It has high nutritional value and delicious taste and hence their requirements of capture and cultivation of this species are constantly increasing. This species experiences varying and increasing temperature levels as it resides in an higher intertidal zone of Thondi coast. The present study examines the effect of thermal stress on the levels of serotonin and crustacean hyperglycemic hormone in the hemolymph of P. pelagicus and analyzes the effect of the monoamine oxidase inhibitor, pargyline on serotonin and CHH level after thermal stress. The results showed increased levels of glucose, CHH and serotonin on exposure to 26 °C in control animals. Pargyline injected crabs showed highly significant increase in the levels of CHH and serotonin on every 2 °C increase or decrease in temperature. A greater CHH level of 268.86±2.87 fmol/ml and a greater serotonin level of 177.69±10.10 ng/ml was observed at 24 °C. This could be due to the effect of in maintaining the level of serotonin in the hemolymph and preventing its oxidation, which in turn induces hyperglycemia by releasing CHH into hemolymph. Thus, the study demonstrates the effect of thermal stress on the hemolymph metabolites studied and the role of pargyline in elevating the levels of serotonin and CHH on thermal stress in the blue swimmer crab, P. pelagicus.     

Ovarian maturation of Penaeus subtilis (Decapoda: Penaeidae): A new insight to describe oocyte development and somatic structures

We observed the presence of follicular cells (FC) in the ovaries of Penaeus subtilis (n = 1198), which led us to classify the development of germ cells into six phases: oogonia, previtellogenic oocytes, primary and secondary vitellogenic oocytes, mature oocytes and atretic oocytes. The FC changes their shape according to the development of germ cells and showed a different distribution along the ovary, which allowed differentiating vitellogenic oocytes into primary and secondary. We also observed that the postovulatory follicles (POF) are composed of follicular cells. The presence of POF in penaeids ovaries is rarely reported, but allows the differentiation between spent and resting stages, commonly grouped in reproductive biology research. Furthermore, observation of ovarian lining was useful to differentiate immature females from females that had spawned at least once. Thus, ovarian development was classified into six stages: immature, early developing, advanced developing, ripe, spent and resting. The distribution and shape variations of FC, ovarian lining features and presence of POF were considered crucial for the classification of ovarian maturation stages. The methods developed here may improve estimates of their reproductive cycle, size at first maturity and spawning season, which are important variables in future studies of the reproductive dynamics.     

Reproductive biology of the shore crab Carcinus maenas (Decapoda,Portunidae): a macroscopic and histological view

A histological study of the reproductive cycle of male and female shore crabs, Carcinus maenas (Linnaeus), was performed monthly on the South West coast of Ireland (from December 2006 to July 2008). The calculated sex ratio deviated from equality, 1:0.53, revealing a strong male bias. A system was devised, based on screening of tissue sections, to describe and stage gametogenic development. Histological examinations revealed that ovarian development occurred biannually, with a primary winter cycle in which the larger crabs reproduced and a secondary summer cycle, when smaller crabs reproduced. An association was observed where more of the larger specimens were caught in the summer months and the smaller specimens in the winter months, which inversely correlated with the segregated breeding cycles. There was strong evidence that mature male crabs could potentially copulate year round since all mature specimens, collected throughout the year, contained viable spermatozoa. Developmental stages of oogenesis and spermatogenesis were described to develop a practical gonadal index for this portunid crab, providing information on the biology of this species, which will be of benefit for fisheries management.     

Ovarian Structure in Milnesium tardigradum (Tardigrada, Milnesiidae) during Early Vitellogenesis

The ultrastructure of the ovary of Milnesium tardigradum during early vitellogenesis is described. Within the ovary, there were large multinuclear cells surrounded by many mononuclear oocytes. Observation of serial sections revealed four multinuclear cells that were connected to each other by cytoplasmic bridges. Each peripheral oocyte was connected to the multinuclear cell. An enormous ER-like structure was conspicuous in the centre of the multinuclear cell. The presence of large numbers of lipid droplets and yolk granules in both multinuclear cells and many mononuclear oocytes suggested a role as nurse cells. A small number of these oocytes grow to be eggs. The structural features of the multinuclear nurse cell were compared with other known examples.     

霍氏啮小蜂的卵巢发育和卵子发生

霍氏啮小蜂是一些鳞翅目和双翅目昆虫蛹的寄生蜂。本文描述该寄生蜂从亚洲玉米螟羽化后0,6 12,18,24,30,36,42,48,54,60,66和72 h的卵巢发育和卵子发生过程。霍氏啮小蜂雌性生殖系统为典型的膜翅目姬小蜂类型,包括2个卵巢,每个卵巢通常有8-9根卵巢管,另有1对输卵管附腺,1个受精囊和1个受精囊腺等。卵黄和卵子形成于羽化后12 h,刚羽化的雌蜂不含成熟卵(成熟卵指数OI=0),成熟卵出现在羽化后24 h,成熟卵数量随后不断增加至数量较为稳定。卵巢管内卵数与雌蜂的体长和头宽呈正相关,相关系数分别为0.859和0.907。研究结果表明霍氏啮小蜂属卵育型(synovigeny)寄生蜂,本文还讨论了卵育型霍氏啮小蜂的卵成熟策略。     

Structural study of the ovary,oogenesis and brooding in Tonicia lebruni (Polyplacophora Chitonidae) from Patagonia

Tonicia lebruni, a common, lower intertidal and subtidal chiton inhabiting Patagonian rocky shores, is a gonochoristic iteroparous species producing large eggs (≈ 400 μm in diameter), which are fertilized and brooded within the pallial groves until released as juveniles. A free larval stage is absent, despite this, T. lebruni is widely distributed along the south‐western Atlantic. At Puerto Deseado, T. lebruni has a marked seasonality in the reproductive cycle, reproducing only once a year. The reproductive period is quite short and defined in time: spawning and brooding take place during the late austral winter and beginning of spring. Recovery of the female gonad starts very soon after spawning. Oogenesis takes about 10–11 months for completion. Brood size is correlated with length of maternal individual. The number of embryos per brood varied between 785 and 5945. Extensive resorption of abortive eggs is viewed as related to limitation of space available for brooding. The egg hull is formed by a large number of minute pentagonal or hexagonal plates each one bearing a short spine bent onto the egg surface. The morphology and the surface of the hull could contribute to the cohesiveness of the brooded egg mass within the pallial grooves.     

Ultrastructural study of oogenesis in Monocelis lineata (Turbellaria,Proseriata)

Summary

Oogenesis in the marine turbellarian proseriat Monocelis lineata was investigated at the ultrastructural level. Oocyte differentiation is not synchronous so that successive stages of germ cell maturation were simultaneously detected in each of the two ovaries. Each developing oocyte is enveloped by follicle cell projections which are presumably involved in a morphologically undetectable support of vitellogenesis. The main features evidenced during oocyte differentiation are: (1) The synthesis of cortical granules by the rough endoplasmic reticulum and Golgi complex, occurring in the earlier stages of oogenesis; (2) The synthesis of yolk globules by the rough endoplasmic reticulum (RER) and Golgi complex, occurring in the later stages of oogenesis, namely late meiotic prophase I. Neither morphologically visible endocytotic activity, nor the presence of intercellular bridges, nor even the development of microvilli were observed at the oolemma or cortical ooplasm, so that the sole mechanism of vitellogenesis appears to be autosynthetic. The significance of these findings is discussed in relation to the taxonomic position of M. lineata and more generally in relation to the phylogenetic history of the class Turbellaria.     

Molt cycle related changes and effect of short term starvation on the biochemical constituents of the blue swimmer crab Portunus pelagicus

Synthesis and hardening of a new exoskeleton are essential to the arthropod molting process. The present study emphasizes the variations in the levels of hemolymph total free sugars, hepatopancreas glycogen and cuticular proteins during the molting stages of Portunus pelagicus. It also reports the effect of short-term starvation conditions on the biochemical constituents of the hemolymph. Intermolt crabs were subjected to 6 days of starvation and hemolymph samples were taken. Standard biochemical procedures were followed toward the quantification of total proteins, total free sugars and total lipids. The total free sugar level in the hemolymph of P. pelagicus was observed to increase during early premolt D₀ (3.108 ± 0.032 g/ml) and a gradual decrease till late postmolt B stage (0.552 ± 0.124 g/ml), suggesting the need for total free sugars to provide energy for the apolysis process. Increase in the levels of hepatopancreas glycogen was observed from 1225 ± 0.04 μg/mg in early premolt D₀ to 1700 ± 0.3 μg/mg in late premolt D_2–3. This is in correlation with the decreased levels of free sugars during premolt stages, suggesting an increase in the storage of glycogen reserves in the hepatopancreas. Cuticular proteins increased during stage B (2.702 ± 0.093 g/ml) and stage C (3.065 ± 0.012 g/ml), indicating exoskeleton hardening and mineralization. Results of the starvation studies clearly showed a steady decline in the level of total free sugars till day 6 (0.099 ± 0.00 g/ml) when compared to the control (8.646 ± 0.08 g/ml). Gradual decrease of total lipids was also observed from the first day of the experiment (6.088 ± 2.44 g/ml) to the last day of the study (0.401 ± 0.20 g/ml) which was 85% lesser than the control (8.450 ± 0.49 g/ml)suggesting the efficient usage of total sugars to consolidate the loss of energy reserves during starvation. The knowledge of Molt-cycle events can be used as a tool for the evaluation of the developmental state providing a morphological reference system for physiological and biochemical studies related to crab aquaculture. Starvation studies enlightens that increasing carbohydrate levels in crab feed together with good protein content could alleviate the natural effects of starvation, improve farm productivity and reduce the deleterious impact of nitrogen pollution generated by rich-protein feeds used in crab farming.     

Old and new DNA unweave the phylogenetic position of the eastern Atlantic gall crab Detocarcinus balssi (Monod, 1956) (Decapoda: Cryptochiridae)

Phylogeny reconstructions based on mtDNA and nDNA have become the standard in studies on relationships between taxa. Difficulties in obtaining material, for example because of small endemic distributions, often lead to gaps in datasets. Collections in natural history museums can provide us with material to fill these gaps, but extracting DNA from historical specimens can be challenging. We used a PCR protocol for small amounts of sample material and high PCR yield on eggs of specimens of the coral‐dwelling gall crab family Cryptochiridae collected in 1984, including material from the eastern Atlantic species Detocarcinus balssi. We obtained DNA sequences from seven older museum specimens using newly developed primers, which we combined with COI sequences from recently collected material. Results show that D. balssi is closest to the Indo‐Pacific species Utinomiella dimorpha and not closely related to one of the other three Atlantic Cryptochiridae species. The remaining newly acquired DNA sequences from museum material cluster with the respective sequences from recently collected specimens.     

Female reproductive morphology of coral‐inhabiting gall crabs (Crustacea: Decapoda: Brachyura: Cryptochiridae)

Gall crabs are obligate associates of stony corals in which they induce skeletal modifications. In some cryptochirid species, females live in open depressions accessible to males; while in others, females are rather isolated in semiclosed galls, which necessitates elaborate sperm storage capabilities by the female. In this study we investigate the female gross morphology and reproductive systems of Fungicola syzygia lodged in semiclosed flattened pits, Opecarcinus cathyae with semiopen pits and Pseudocryptochirus viridis from shallow open depressions using line drawings and histological methods. The general morphology of the cryptochirids' reproductive systems is uniform and conforms to other thoracotreme brachyurans: paired muscular vaginae of the concave pattern lead from the sternal gonopores into paired seminal receptacles where sperm is stored. The seminal receptacle is internally lined by distinct types of epithelia: a cuticle underlined by a columnar epithelium ventrally, a monolayered secretory epithelium dorsally and a multilayered transfer tissue where the oviducts enter the seminal receptacle. In all studied specimens, the seminal receptacle contained free spermatozoa; however, in specimens of Pseudocryptochirus viridis it also contained spermatophores, indicating a recent insemination. In contrast to most other brachyurans ovaries of the investigated cryptochirids extend into the pleon. The specific degree of ovary extension differs between the studied species and is closely related to female body shape.     

孟氏隐唇瓢虫生殖系统结构和卵子发生的研究

【目的】孟氏隐唇瓢虫Cryptolaemus montrouzieri Mulsant生殖系统结构和卵细胞发生将为昆虫的系统进化关系及瓢虫分类提供依据,同时可作为瓢虫人工饲料研究开发的参考。【方法】利用组织石蜡切片技术和光学显微镜,观察孟氏隐唇瓢虫生殖系统结构,以及自成虫羽化后不同发育阶段卵巢发育状况和成熟卵巢管卵子发生过程。【结果】孟氏隐唇瓢虫雄性生殖系统包括2对附腺、1对精巢、1对输精管、1对贮精囊、射精管、弯管和阳基。雌性生殖系统包括2片生殖板、生殖腔、受精囊、中输卵管、1对侧输卵管和1对卵巢。单侧卵巢管数量在11~14根之间,卵巢管端部延伸出细长的端丝。卵巢管属于端滋式,分为原卵区和生长区。滋养细胞分散且细胞核几乎充满整个细胞,未见合胞体。卵细胞稀疏地集中在原卵区下端,并且可见营养索向卵巢管顶端延伸。根据卵细胞位置和形态,卵黄积累情况,滤泡细胞形态变化,将卵细胞发生分为前期,中期,中后期和后期。卵细胞发育后期,营养索消失,滤泡细胞排列疏松,细胞间隙增大。【结论】孟氏隐唇瓢虫卵巢管的滋养细胞是端滋式卵巢管滋养细胞中的原始类型,且推测瓢虫科昆虫卵巢管滋养细胞均属于此类。卵细胞早期发育过程中,卵细胞通过营养索从滋养细胞获取营养物质。     

Genetic differentiation in the mud crab Scylla serrata (Decapoda: Portunidae) within the Indian Ocean

Scylla serrata (Decapoda: Portunidae) is a swimming crab that is widespread in the Indo-Pacific region and commonly found in estuarine and mangrove waters. An extended planktonic larval phase suggests high dispersal potential and the possibility of extensive gene flow between conspecific populations at least on a geographic mesoscale (tens to hundreds of kilometres).Intraspecific variation of the mitochondrial DNA cytochrome oxidase subunit I (mtDNA COI) gene was investigated in 77 individuals from four representative mangrove swamps of the African tropics (Kenya and Zanzibar) by means of DNA sequencing. We examined 535 base pairs (bp) and identified 24 different haplotypes. Each population sample is characterised by a single most frequent haplotype, shared among all four populations, and a small number of rare ones, typically present in only one or two individuals and representative of a specific population.Analysis of molecular variance (AMOVA), F_ST statistics and χ² contingency analysis of spatial distribution of mtDNA haplotype frequencies revealed in toto a significant genetic differentiation among populations. These results could indicate that gene flow might be reduced, even between geographically close sites, despite the high potential for dispersal; anyway, at the recorded level of divergence and owing to the abundance of rare haplotypes and singletons in our data set, repeated sampling over time is necessary to establish whether the recorded pattern of genetic differentiation is stable and biologically significant.Finally, integration of our data with those reported by Gopurenko et al. [Mar. Biol. 134 (1999) 227] on S. serrata from South Africa, Red Sea and Mauritius Islands allowed to infer S. serrata population structure within a larger area of the Indian Ocean region.     

Effects of constant and varying temperatures on the development of blue swimmer crab (Portunus pelagicus) larvae: Laboratory observations and field predictions for temperate coastal waters

Temperature is widely held to be a critical factor for the development of marine invertebrate larvae. We investigated three specific aspects of this relationship for the blue swimmer crab, Portunus pelagicus, in a temperate gulf: (1) the effects of different but temporally constant temperatures on the survival and developmental period of larvae reared in the laboratory, (2) the effects of varying temperatures on the survival and developmental period of larvae reared in the laboratory, and (3) prediction of larval developmental periods under seasonal temperature changes found in the field. Temperature had a marked effect on larval survival. At constant temperatures of 22.5 and 25 °C larval survival was far greater than at lower temperatures down to 17 °C, and developmental period of the larval period was inversely related to (constant) temperature. However, larvae in temperate coastal waters will usually be exposed to seasonally varying, rather than constant, temperatures. To account for this, a larval developmental period model was created and then verified in the laboratory by rearing larvae under varying temperatures. Results of this work demonstrated that developmental periods were markedly different under constant versus varying temperature regimes. Using different temperature simulations for a temperate gulf (Gulf St Vincent, South Australia), the developmental period model predicted that in years of ‘average’ seasonal temperature changes, total larval developmental period could range from 26 to 45 days depending on the day of hatching. In such cases, peak postlarval settlement was predicted to occur between mid January and mid March. Results from this study also predict that larval survival (and thus postlarval settlement) will be maximised in years with abnormally warm summers. Whilst the developmental period model was used to make predictions of developmental period for P. pelagicus in a temperate gulf, it could readily be adapted to predict developmental periods in other coastal environments.     

Spermatozoa ultrastructure of the pink shrimp Farfantepenaeus paulensis (Decapoda: Dendrobranchiata)

Braga, A.L., Nakayama, C.L., Suita de Castro, L.A. and Wasielesky, W. 2011. Spermatozoa ultrastructure of the pink shrimp Farfantepenaeus paulensis (Decapoda: Dendrobranchiata). —Acta Zoologica (Stockholm) 00 : 1–6. The spermatozoa ultrastructure of the pink shrimp Farfantepenaeus paulensis was investigated in this morphological study. Spermatophores and spermatozoa were analyzed by electron microscopy. The pink shrimp spermatophore is divided into two regions: the appendage and the spermatophore main body, where spermatozoa are grouped in a spermatic mass. Pink shrimp spermatozoa are unistellate and are composed of main body and single spike. The spermatozoa body comprises a perinuclear cytoplasmic band, nucleus, acrosomal cap, and subacrosomal region. The spermatozoa cell mean total length was 10.71 μm, the mean body diameter was 5.56 μm, and the mean spike length and diameter were 5.15 μm and 0.85 μm, respectively.     

Yolk formation in an annelid (Ophryotrocha puerilis, polychaeta)

The polychaete Ophryotrocha does not show a distinct breeding season. Egg masses are produced throughout the year (continuous breeder sensu Olive and Clark, 1978). A female specimen may contain up to three different generations of oocytes with oocyte growth and maturation in each batch being well synchronized. Oogenesis takes about 18 days from proliferation of the oogonia to mature eggs. In each segment pairs of sister cells interconnected by cytoplasmic bridges are located in outpocketings of the ventral mesentery which form the gonad wall. Presumptive oocytes and nurse cells are not easily distinguished at that time. Vitellogenesis is initiated while both oocytes and nurse cells are still in the ovary. Mitochondria, multivesicular bodies (transformed mitochondria ?), dense bodies, preformed yolk bodies of smaller size and lipid droplets are probably passed through the cytoplasmic bridge from the nurse cell to the oocyte. Yolk formation includes different mechanisms and materials of different origin. Autosynthetic yolk formation predominates during the first intraovarial growth phase. After detachment of oocyte-nurse cell-complexes from the gonad pinocytotic activity of nurse cells and particularly oocytes, increases considerably. The existence of coated vesicles suggests that external sources of yolk precursors contribute to yolk formation. Prior to oocyte maturation the remnants of the nurse cell are incorporated by oocytes.     

Ovarian structure,folliculogenesis and oogenesis of the annual killifish Millerichthys robustus (Cyprinodontiformes: Cynolebiidae)

Cellular aspects of oocyte development of the Mexican rivulus Millerichthys robustus were morphologically described in order to analyze ovarian function and the cellular recruitment dynamics associating it with life history strategies of annual killifishes. Millerichthys is an iteroparous batch spawner with continuous oocyte recruitment and indeterminate fecundity with asynchronous development of the follicles. It has two ovaries of cystovarian type, with a central lumen, which communicates with the outside through the caudal region of the ovary, that is, the gonoduct. From the walls of the ovary, irregular lamellae composed of germinal epithelium and vascularized stroma project. Oogenesis starts with oogonial proliferation, found alone or in nests within the germinal epithelium. The oogonia come into meiosis becoming oocytes and advancing to the chromatin nucleolus stage and to early primary growth stage. Folliculogenesis is completed in the primary growth stage and cortical alveoli step. Follicles moves toward the stroma, but they continue to be attached to the germinal epithelium through the basement membrane until ovulation. The inclusion of fluid yolk in the follicles during the secondary growth stage was observed. During ovulation, the follicle collapsed, the oocyte was released into the lumen, and the constitutive elements of the post-ovulatory follicle complex remained in the stroma.