Brood Sac Research Articles

Infestation of sporocysts of parasite Leucochloridium in the snails Succinea daucina and Indosuccinea semiserica

Aim: The aim of the present study was to evaluate the differential beating rate of the parasite Leucochloridium within the tentacle of the infected snail Succinea daucina and Indosuccinea semiserica under different photoperiodic conditions. Methodology: In this study, the rhythmic beating pulse of sporocyst of the parasite invading the tentacle of the snail was recorded from multiple snails, periodically in light and dark hours. The numbers of beat per minute were subjected to univariate ANOVA using day and photoperiod as explanatory variables. A two-sample t-test was performed with the mean of pulse rate in light and dark phase to estimate the significant differences between the same. Results: The observations revealed that sporocyst brood sac of I. semiserica beat mean 99.23 ± 1.91 SE per minute ( 90 – 101) in day hours and mean 76.57 ± 4.64 SE per minute (48 – 88) in the night hours (n = 7 observations). In S. daucina, the sporocyst brood sac beat 65.85 ± 2.17 SE per minute (25 – 94) during day and 30.5 ± 3.55 SE per minute (0 – 80) in night (n = 9 separate observations). The beats of the sporocyst varied significantly diurnally and with the consecutive days revealed through t-test and ANOVA. Interpretation: Apparently, the colourful sporocyst broodsac of Leucochloridium faster beating rate under light may attract definite host, the insectivorous birds which prey on the Leucochloridium sporocycst disrupting the snail tentacle. Thus, the pulsating sporocyst inside the tentacle makes the infected host snail more conspicuous for the birds, linking the snail- Leucochloridium- bird cycle. Key words: Indosuccinea semiserica, Intermediate host, Land snail, Leucochloridium sp., Succinea daucina, Trematoda

Spectacular alterations in the female reproductive system during the ovarian cycle and adaptations for matrotrophy in chernetid pseudoscorpions (Pseudoscorpiones: Chernetidae)

Pseudoscorpions are small matrotrophic chelicerates. The embryos develop in a brood sac and feed on the nutritive fluid provided by the female. It was widely accepted that the nutritive fluid is synthesized in the ovary. Recent studies have shown that in Chelifer cancroides, a representative of Cheliferidae, considered one of the most derived pseudoscorpion families, the nutritive fluid is produced not only in the ovary but also in the oviducts. Since evolution of adaptations for matrotrophy in pseudoscorpions is poorly known, we aimed to verify our hypothesis that pseudoscorpions of the family Chernetidae, closely related to Cheliferidae, share the traits of adaptations to matrotrophy in the structure and function of the female reproductive system with C. cancroides. We analysed the structure of the ovary and oviducts in five representatives of chernetids with light, confocal, and transmission electron microscopy. The results confirmed our hypothesis and provided new data which broaden our knowledge of matrotrophic pseudoscorpions. We show that in chernetids, the ovary and oviducts undergo significant alterations including their size, multistep hypertrophy and polyploidization of the epithelial cells involved in secretion of the nutritive fluid, the complex secretory activity of the epithelial cells, massive degeneration of the epithelial cells that have completed secretion, and epithelium renewal.

Small body size of pseudoscorpions and a distinct architecture of the ovary: A step to miniaturization?

Chelicerata, the second largest subphylum of Arthropoda, includes invertebrates with a wide range of body size. Pseudoscorpions are among small or miniature chelicerates which exhibit several morphological, anatomical, and developmental features related to miniaturization, e.g., replacement of book lungs by tracheae, unpaired gonads, and matrotrophic development of the embryos outside the female body, in the brood sac. In this paper, we show the ovary structure of two pseudoscorpion species, Cheiridium museorum and Apocheiridium ferum (Cheiridiidae). Both cheiridiids are one of the smallest pseudoscorpions. The results of our observations conducted in light, transmission electron, and confocal microscopy demonstrate that the ovary of C.museorum and A.ferum, displays a significant structural difference that is unusual for chelicerates. The difference concerns the spatially restricted position of the germarium. We show that such ovary architecture results in a significantly reduced number of growing oocytes and in consequence a reduced number of deposited eggs. A centrally located germarium implies also a modified pattern of ovary development during oocyte growth due to long distance migration of the germline and the accompanying somatic cells. Herein, we postulate that such an ovary structure is related to the pseudoscorpion's small body size and it is a step towards miniaturization in the smaller pseudoscorpions species.

Reproductive biology, embryonic development and matrotrophy in the phylactolaemate bryozoan Plumatella casmiana

Bryozoa is a phylum of aquatic, colonial suspension-feeders within the Lophotrochozoa. In the Phylactolaemata embryonic development occurs in an internal brood sac on the body wall accompanied by extraembryonic nutrition. Owing to previous contradictive descriptions, many aspects of their sexual reproduction require restudy. Consequently, this study analyses embryogenesis of the freshwater bryozoan Plumatella casmiana by serial sections, 3D reconstruction and transmission electron microscopy. Early embryos cleave and soon develop into blastulae with a small central cavity. The mesoderm forms by delamination starting from the distal side towards the proximal end. In later embryos two polypides form on the posterior side that ultimately will be covered by a ciliated mantle in the larva. Embryos increase in size during development and form temporary cell contacts to the embryo sac. Mesodermal cells of the embryo sac show signs of transcellular transport indicating that embryos are nourished by transferring nutrients from the maternal coelom towards the brood cavity. This study clarifies several details such as mesoderm formation and the onset of bud development. Embryos are connected to their respective embryo sacs by a variety of temporary cytoplasmic processes formed by both tissues during embryogenesis, including a ‘placental’ ring zone. Although ultrastructural data of these cell contacts are not entirely conclusive about their function, we suggest that embryos absorb nutrients via the entire surface. The close opposition of embryos to the embryo sac implies placentation as matrotrophic mode in phylactolaemate bryozoans, with embryo sacs acting as placental analogues.

Adaptations for matrotrophy in the female reproductive system in the pseudoscorpion Chelifer cancroides (Chelicerata: Pseudoscorpiones, Cheliferidae).

Pseudoscorpiones (pseudoscorpions, false scorpions) is an order of small terrestrial chelicerates. While most chelicerates are lecithotrophic, that is, embryos develop due to nutrients (mostly yolk) deposited in the oocyte cytoplasm, pseudoscorpions are matrotrophic, that is, embryos are nourished by the female. Pseudoscorpion oocytes contain only a small amount of yolk. The embryos develop within a brood sac carried on the abdominal site of the female and absorb nutrients by a pumping organ. It is believed that in pseudoscorpions nutrients for developing embryos are produced in the ovary during a postovulatory (secretory) phase of the ovarian cycle. The goal of our study was to analyze the structure of the female reproductive system during the secretory phase in the pseudoscorpion Chelifer cancroides, a representative of the family Cheliferidae, considered to be one of the most advanced pseudoscorpion taxa. We use diverse microscopic techniques to document that the nutritive fluid is produced not only in the ovaries but also by the epithelial cells in the oviducts. The secretory active epithelial cells are hypertrophic and polyploid and release their content by fragmentation of apical parts. Our observations also indicate that fertilization occurs in the oviducts. Moreover, in contrast to previous findings, we show that secretion of the nutritive material starts when the fertilized oocytes reach the brood sac and thus precedes formation of the pumping organ. Summing up, we show that C. cancroides exhibits traits of advanced adaptations for matrotrophy due to coordinated secretion of the nutritive fluid by the ovarian and oviductal epithelial cells, which substantially increases the efficiency of nutritive fluid formation. Since the secretion of nutrients starts before formation of the pumping organ, we suggest that the embryos are able to absorb the nutritive fluid also in the early embryonic stages.

Molecular mechanisms underlying milk production and viviparity in the cockroach, Diploptera punctata

Viviparous reproduction is characterized by maternal retention of developing offspring within the reproductive tract during gestation, culminating in live birth. In some cases, a mother will provide nutrition beyond that present in the yolk; this is known as matrotrophic viviparity. While this phenomenon is best associated with mammals, it is observed in insects such as the viviparous cockroach, Diploptera punctata. Female D. punctata carry developing embryos in the brood sac, a reproductive organ that acts as both a uterus and a placenta by protecting and providing a nutritive secretion to the intrauterine developing progeny. While the basic physiology of D. punctata pregnancy has been characterized, little is known about the molecular mechanisms underlying this phenomenon. This study combined RNA-seq analysis, RNA interference, and other assays to characterize molecular and physiological changes associated with D. punctata reproduction. A comparison of four stages of the female reproductive cycle and males revealed unique gene expression profiles corresponding to each stage and between sexes. Differentially regulated transcripts of interest include the previously identified family of milk proteins and transcripts associated with juvenile hormone metabolism. RNA interference and methoprene application experiments established the potential impacts of bothbreakdown and synthesis reduction of juvenile hormone in maintaining pregnancy in D. punctata. These studies provide the comprehensive molecular mechanisms associated with cockroach viviparity, which will be a critical resource for comparative purposes among viviparity in insect systems.

Matrotrophic viviparity constrains microbiome acquisition during gestation in a live-bearing cockroach, Diploptera punctata.

The vertical transmission of microbes from mother to offspring is critical to the survival, development, and health of animals. Invertebrate systems offer unique opportunities to conduct studies on microbiome‐development‐reproduction dynamics since reproductive modes ranging from oviparity to multiple types of viviparity are found in these animals. One such invertebrate is the live‐bearing cockroach, Diploptera punctata. Females carry embryos in their brood sac, which acts as the functional equivalent of the uterus and placenta. In our study, 16S rRNA sequencing was used to characterize maternal and embryonic microbiomes as well as the development of the whole‐body microbiome across nymphal development. We identified 50 phyla and 121 classes overall and found that mothers and their developing embryos had significantly different microbial communities. Of particular interest is the notable lack of diversity in the embryonic microbiome, which is comprised exclusively of Blattabacteria, indicating microbial transmission of only this symbiont during gestation. Our analysis of postnatal development reveals that significant amounts of non‐Blattabacteria species are not able to colonize newborn D. punctata until melanization, after which the microbial community rapidly and dynamically diversifies. While the role of these microbes during development has not been characterized, Blattabacteria must serve a critical role providing specific micronutrients lacking in milk secretions to the embryos during gestation. This research provides insight into the microbiome development, specifically with relation to viviparity, provisioning of milk‐like secretions, and mother–offspring interactions during pregnancy.

Compositional differences among female-associated and embryo-associated microbiota of the viviparous Pacific Beetle cockroach, Diploptera punctata.

All cockroach species, except one, harbor the endosymbiont Blattabacterium, transmitted from females to embryos. Adult cockroaches acquire non-Blattabacterium bacteria as part of their gut microbiota over time, but our knowledge of the possible transmission of these non-Blattabacterium bacteria from females to embryos is rudimentary. We characterized the gut microbiota of gravid viviparous Diploptera punctata females and the non-Blattabacterium microbiota of associated developing embryos, as well as the gut microbiota of non-gravid females, and the microbiota of orphan embryos (females not included), following high-throughput sequencing of the 16S rRNA gene to assess bacterial transference. We determined significant differences in community composition between gravid females and associated embryos and overall greater similarity in community composition among embryos than adult females. Results suggest various routes of transference of bacteria from females or the environment to embryos. The bacterial families Halomonadaceae and Shewanellaceae were more abundant in embryos than in gravid females. The functional relevance of these families remains to be elucidated, but provisioning of amino acids deficient in the brood sac secretion is a possibility. Overall, our results highlight the need for further studies investigating the uptake and selective screening of microbes by D. punctata embryos, as well as their functions.

Origin of origami cockroach reveals long-lasting (11Ma) phenotype instability following viviparity.

Viviparity evolved in bacteria, plants, ˃141 vertebrate lineages (ichthyosaurs, lizards, fishes, mammals, and others), and in 11 of 44 insect orders. Live-birth cockroaches preserved with brood sac (3D recovered two times optically) included Diploptera vladimir, Diploptera savba, Diploptera gemini spp.n., D. sp.1-2, and Stegoblatta irmgardgroehni from Green River, Colorado; Quilchena, Republic; McAbee, Canada; and Baltic amber, Russia (49, 54, and 45Ma). They evolved from rare and newly evolved Blaberidae; they radiated circumtropically, later expanded into SE Asia, and have now spread to Hawaii and the SE USA. Association of autapomorphic characters that allow for passive and active protections from parasitic insects (unique wing origami pleating identical with its egg case-attacking wasp) suggest a response to high parasitic loads. Synchronized with global reorganization of the biota, morphotype destabilization in roaches lasted approximately 11-22Ma, including both the adaptation of novel characters and the reduction of others. Thus, while viviparity can be disadvantageous, in association with new Bauplans and/or behaviors, it can contribute to the evolution of taxa with viviparous representatives that are slightly selectively preferred.

The challenging role of life cycle monitoring: evidence from bisphenol A on the copepod Tigriopus japonicus

We used the marine model copepod taxon T. japonicus Mori, 1938 for a first short sublethal life cycle assay of the endocrine-disrupting chemical bisphenol A (BphA) where we studied life cycle and reproductive success. Individual copepodid-I-stages were reared in 96-well microplates to adulthood in as short as 6 days in only 200 μl of 0.1 mg bisphenol (BphA/L) or seawater solution as a control (CON). Males and females were then mated in one well and checked daily for the following endpoints: postmating days to female brood sac extrusion, days from brood sac extrusion to naupliar hatching, first clutch brood size, hatching success, and reproductive success. Mean naupliar hatching times ranged from 1.3 days for CON-reared mating pairs to 2.7 days for BphA-reared mating pairs; mean brood size was significantly lower for CON females mated with BphA males than for all other mating combinations; reproductive success of CON females mated with BphA males and BphA females mated with BphA males showed significant differences between females and males. In conclusion, T. japonicus complete life cycle exposures to BphA at an environmentally realistic concentration resulted generally in reproductive depression with effect values depending on the exposure history of the different gender.

Phoretic relationship between Lustrochernes grossus (Pseudoscorpionida: Chernetidae) and Odontotaenius striatopunctatus (Coleoptera: Passalidae)

Phoresy is considered a non-parasitic partnership between a small animal (phoront) and a large one (host) that result in phoront transportation by the host. Throughout two years of observation and nursing of 16 family groups of the passalid Odontotaenius striatopunctatus, we noticed the presence of the pseudoscorpion Lustrochernes grossus within 11 of their 16 nests, which consisted of rottenwood logs (30 cm long × 15-20 cm in diameter) cut into 2 cm segments and secured with rubber bands to facilitate observation. The presence of pseudoscorpions became evident from the moment the passalids started building their nests creating space and food for their young. Evidence was found that pseudoscorpions can go under the elytra of passalids. Their presence did not prevent the reproduction of the beetles, performed in 13 of the 16 set nests (Fischer’s exact test: P =0.71, N = 16): neither was the number of passalids offspring modified (Mann Whitney U = 11.50, P = 0.2553). On three occasions during spring, L. grossus females carrying brood sac was documented. In this facultative relationship, phoresy and phagophilia could be involved; some hypotheses on the factors that may interfere with it are posed.

Life history and distribution of the Arctic pseudoscorpion, <em>Wyochernes asiaticus</em> (Chernetidae)

The Pseudoscorpiones are a remarkable yet understudied order of arachnids. The northernmost species in North America, Wyochernes asiaticus (family Chernetidae), occurs under rocks beside rivers or creeks and can be found above the Arctic Circle in Canada. In North America, the species is limited to the northwest, although its global distribution includes parts of Asia. It is presumably a Beringian species with quite specialized habitat affinities. I report on some life history traits of this species, based on examination of nearly 600 specimens from 16 localities in the Yukon and Northwest Territories. All life stages were collected. Of the females, 17% were carrying brood sacs, with an average of 10.5 eggs per brood sac; larger females tended to have larger clutch sizes. Despite these data on the natural history and distribution of W. asiaticus, its phylogeographic history and how the species feeds, disperses, and recolonizes habitats after flooding remain largely unknown.

Reproduction of Pisidium casertanum (Poli, 1791) in Arctic lake.

Freshwater invertebrates are able to develop specific ecological adaptations that enable them to successfully inhabit an extreme environment. We investigated the brooding bivalve of Pisidium casertanum in Talatinskoe Lake, Vaigach Island, Arctic Russia. Here, quantitative surveys were conducted, with the collection and dissections of 765 molluscs, on the basis of which analyses on the brood sacs length (marsupia) and the number and size of embryos, were performed. In this study, the number of brooded embryos was positively correlated with the parent's shell length. The number of extramarsupial embryos was much lower than the number of intramarsupial embryos. Our research also showed that the brood sac length and embryos within one individual can vary significantly. Thus, we detected that P. casertanum has a specific brooding mechanism, accompanied by asynchronous development and embryos release by the parent. We suggest that such a mode could result in the coin-flipping effect that, presumably, increases the population breeding success in the harsh environment of the Arctic lake.

Diopatra nishii, a new brooding species of Onuphidae (Annelida) from Japan

A small tubeworm, Diopatra nishii, is newly described from the shallow waters of Boso Peninsula and Izu Peninsula, as the second Japanese species of this genus. It differs from D. sugokai, a common baitworm in Japan, by its much smaller size, single rather than double postchaetal lobes on the modified parapodia and direct development in the parental tube rather than broadcast spawning. Juveniles of two different stages of the new species were found in a brood sac in the parental tube; these and recently settled juveniles are described.

Female harlequin beetle riding pseudoscorpion (Cordylochernes scorpioides) with a brood of birthing nymphs. Pseudoscorpions are viviparous, with the embryos developing in an external brood sac overlying the mother_s genital aperture. After birth, nymphs remain with their mother for 2‐3 days until they are fully sclerotized. Photo reproduced by permission of Jeanne Zeh.

Female harlequin beetle riding pseudoscorpion (<i>Cordylochernes scorpioides</i>) with a brood of birthing nymphs. Pseudoscorpions are viviparous, with the embryos developing in an external brood sac overlying the mother_s genital aperture. After birth, nymphs remain with their mother for 2‐3 days until they are fully sclerotized. Photo reproduced by permission of Jeanne Zeh.

New Zealand mudsnails (Potamopyrgus antipodarum) in Boulder Creek, Colorado: environmental factors associated with fecundity of a parthenogenic invader

New Zealand mudsnails (Potamopyrgus antipodarum (Gray, 1853)) are non-native snails that are increasingly spreading in freshwater systems in North America. Most invasive populations are parthenogenic and threaten native freshwater diversity. We observed variability of P. antipodarum fecundity each month for 16 months at a recently invaded site in Boulder Creek, Colorado. We collected 100 snails each month and dissected them to count embryos in the brood sac. We used a general linear model analysis to examine water-quality variables as predictors of the monthly variability in P. antipodarum fecundity. After dissecting 1600 snails, we observed four male individuals (&lt;1%), brood sizes ranging from 0 to 70 embryos per snail, reproductively mature females at 3.2 mm in length or greater, and a significant relationship between snail length and embryo counts (r2= 0.38, p &lt; 0.001). The model with the highest level of support for predicting variability in snail fecundity included water temperature, snail shell length, water hardness (calcium carbonate), and nutrient levels (total phosphate) (adjusted r2= 0.53, p &lt; 0.01). These variables may be important for snail growth and promote increased rates of reproduction in this parthenogenic, invasive snail. These results can further inform efforts to model geographic areas at high risk of P. antipodarum establishment and rapid demographic growth.

Differentiation and function of the ovarian somatic cells in the pseudoscorpion, Chelifer cancroides (Linnaeus, 1761) (Chelicerata: Arachnida: Pseudoscorpionida)

Pseudoscorpion females carry fertilized eggs and embryos in specialized brood sacs, where embryos are fed with a nutritive fluid produced and secreted by somatic ovarian cells. We used various microscopic techniques to analyze the organization of the somatic cells in the ovary of a pseudoscorpion, Chelifer cancroides. In young specimens, the ovary is a cylindrical mass of internally located germline cells (oogonia and early previtellogenic oocytes) and two types of somatic cells: the epithelial cells of the ovarian wall and the internal interstitial cells. In subsequent stages of the ovary development, the oocytes grow and protrude from the ovary into the hemocoel (opisthosomal cavity). At the same time the interstitial cells differentiate into the follicular cells that directly cover the oocyte surface, whereas some epithelial cells of the ovarian wall form the oocyte stalks – tubular structures that connect the oocytes with the ovarian tube. The follicular cells do not seem to participate in oogenesis. In contrast, the cells of the stalk presumably have a dual function. During ovulation the stalk cells appear to contribute to the formation of the external egg envelope (chorion), while in the post-ovulatory phase of ovary function they cooperate with the other cells of the ovarian wall in the production of the nutritive fluid for the developing embryos.

Reexamination of the five species of Blaberidae deposited in the Entomological Collection of the Museu Nacional/UFRJ, Rio de Janeiro, Brazil

The family Blaberidae is diverse, including large wingless, brachypterous, or fully winged species. Wings of the fully winged species have many-branched veins and reach beyond the apex of the abdomen. Some species are smaller and green in color. These cockroaches are more than 40mm long; species have front femora with 1 to 3 spines on the ventroposterior margin and 1 at the tip; and lack arolia. The females of Blaberidae possess a brood sac, wherein the ootheca or loose eggs are retained until the eggs hatch (ovoviviparity). The phylogeny of roach families was explored by McKittrick (1964), Grandcolas (1994, 1996), Kambhampati (1996) and Klass (1997, 1998, 2001) (Grimaldi &amp; Engel, 2004). We update knowledge of five species of Blaberidae from Brazil, based on a literature review and examination of the Blattaria collection in the Department of Entomology at the Museu Nacional of Rio de Janeiro, Brazil (MNRJ).

First occurrence of the mysid Hemimysis anomala in an inland lake in North America, Oneida Lake, NY

Hemimysis anomala (Crustacea, Mysidae) is a recent invader to North America that until now was reported only from the Laurentian Great Lakes and their immediate embayments, along with the St. Lawrence River. In August 2009, we identified Hemimysis in diets of white perch and yellow perch in Oneida Lake, NY. Night time vertical plankton net tows detected Hemimysis at four sites across the lake. Hemimysis in fish diets (5.5–8.6 mm) were larger than in net tows (2.2–7.0 mm) and reproduction is occurring as some females had brood sacs. This is the first documented introduction of Hemimysis to an inland lake in North America, outside the Great Lakes. Oneida Lake is located 53 river km upstream from Lake Ontario, the nearest known source of Hemimysis. No genetic differences were found between Hemimysis in Oneida Lake and Lake Ontario, indicating this is likely the source of introduction. Several large rapids, locks, and dams separate the two lakes, and as a result the most likely vector of introduction to Oneida Lake is pleasure boat or light commercial traffic via the canal system or overland transport. The presence of Hemimysis in Oneida Lake 3 years after it was first found in Lake Ontario suggests this species may spread rapidly throughout the basin. Despite an intensive monitoring program on Oneida Lake directed at fish, zooplankton, and limnology, Hemimysis was only detected in fish diets and night time zooplankton tows, indicating it may go undetected in lakes for some time using traditional daytime net tows.

The Internal-Brooding Apparatus in the Bryozoan Genus Cauloramphus (Cheilostomata: Calloporidae) and Its Inferred Homology to Ovicells

We studied by SEM the external morphology of the ooecium in eight bryozoans of the genus Cauloramphus (Cheilostomata, Calloporidae): C. spinifer, C. variegatus, C. magnus, C. multiavicularia, C. tortilis, C. cryptoarmatus, C. niger, and C. multispinosus, and by sectioning and light microscopy the anatomy of the brooding apparatus of C. spinifer, C. cryptoarmatus, and C. niger. These species all have a brood sac, formed by invagination of the non-calcified distal body wall of the maternal zooid, located in the distal half of the maternal (egg-producing) autozooid, and a vestigial, maternally budded kenozooidal ooecium. The brood sac comprises a main chamber and a long passage (neck) opening externally independently of the introvert. The non-calcified portion of the maternal distal wall between the neck and tip of the zooidal operculum is involved in closing and opening the brood sac, and contains both musculature and a reduced sclerite that suggest homology with the ooecial vesicle of a hyperstomial ovicell. We interpret the brooding apparatus in Cauloramphus as a highly modified form of cheilostome hyperstomial ovicell, as both types share 1) a brood chamber bounded by 2) the ooecium and 3) a component of the distal wall of the maternal zooid. We discuss Cauloramphus as a hypothetical penultimate stage in ovicell reduction in calloporid bryozoans. We suggest that the internal-brooding genus Gontarella, of uncertain taxonomic affinities, is actually a calloporid and represents the ultimate stage in which no trace of the ooecium remains. Internal brooding apparently evolved several times independently within the Calloporidae.