Germ Balls Research Articles

An atlas of the germ ball-cercaria-schistosomulum transition in Schistosoma mansoni, using confocal microscopy and in situ hybridisation

Schistosomes are complex platyhelminth parasites with a genome comprising ∼12,000 protein-coding genes, three distinct generations, and at least seven distinct phenotypes. We chart here cellular and gene expression changes associated with development of the cercaria, in the intramolluscan daughter sporocyst, and its transformation into the skin stage schistosomulum upon infection of the mammalian host. We describe the morphology of the early daughter sporocyst and the increasing complexity of cellular organisation in germ balls as they rapidly develop into cercariae. We show how individual myocytes differentiate and combine to create the complex musculature of the head capsule and body wall. In situ hybridisation reveals that some transcripts encoding the secretory proteins, released during skin penetration, are expressed in gland-cell precursors very early in germ ball development. However, those for the projected anti-inflammatory protein Sm16-stathmin are widely expressed in germ ball tissues, suggesting the protein has intracellular functions. Transcripts for smkk7 are expressed in six cells of the larval body, while the KK7 protein is present throughout the peripheral nerve net, including sensory nerve bulbs, providing a marker for the nerve net in adult worms. We also note that the cercaria-schistosomulum transformation is accompanied by tissue remodelling without growth.

Parasitism by a Digenea in Lucina pectinata (Mollusca: Lucinidae).

Lucina pectinata is an important economic resource in the Brazilian coast. This study reports parasitism caused by a Digenea in this species. The specimens (n = 470) were collected in December 2012 in a mangrove swamp of the Cachoeira River estuary, Bahia, Brazil. They were measured along the anterior-posterior axis (length), and after macroscopic analysis for parasites and diseases cuts of 5 mm were fixated in Carnoy's solution and processed by routine histology technique wherein sessions of 7 μm were stained with Harris hematoxylin and eosin (H&E). The tissues were examined using an optical microscope. The mean length of L. pectinata was 4.0 ± 0.53 cm. Microscopic analysis showed sporocysts containing both germ balls as cercariae of an unidentified Digenea (Platyhelminthes), these in various stages of development. The prevalence was 1.48% (7/470). In a parasitized specimen was macroscopic evidence of tissue densification of gills. The sporocysts were observed in mantle, gills, digestive gland and gonads, with evident alteration/destruction of tissues, including parasitic castration. There were no other parasites found, which is probably related to inaccessibility and chemical conditions in which lives L. pectinata, i.e., between 10 and 20 cm in mangrove sediment.

Stage and tissue expression patterns of Schistosoma mansoni venom allergen-like proteins SmVAL 4, 13, 16 and 24

BackgroundSchistosoma mansoni venom allergen-like protein (SmVAL) is a gene family composed of 29 members divided into group 1 encoding proteins potentially secreted, and group 2 encoding intracellular components. Some members were found to be upregulated in the transition of germ ball - cercariae - day 3 schistosomula, suggesting that group 1 SmVAL proteins are associated with the invasion of the human host, although their functions are not completely established. Recently, we have described the localization of SmVAL7 (group 1) and SmVAL6 (group 2) transcripts in the oesophageal gland and in the oral and ventral suckers of adult parasites, respectively. The expression patterns of the two genes suggest that SmVAL7 protein plays a role in the blood-feeding process while SmVAL6 is associated with the parasite attachment and movement in the vasculature. In this way, searching for additional secreted SmVAL proteins that could be involved in key processes from skin penetration to the beginning of blood-feeding, we investigated the tissue localization of SmVAL4, 13, 16 and 24 by whole-mount in situ hybridization (WISH).ResultsWe report here the localization of group 1 SmVAL4 and 24 transcripts in the pre-acetabular glands of developing germ balls. Time course experiments of in vitro cultured schistosomula after cercariae transformation demonstrated that SmVAL4 protein is secreted during the first 3 h of in vitro culture, correlating with the emptying of acetabular glands as documented by confocal microscopy. In addition, the localization of SmVAL13 transcripts in adult male anterior oesophageal gland suggests that the respective protein may be involved in the first steps of the blood-feeding process. SmVAL16 was localized close to the neural ganglia and requires further investigation.ConclusionsOur findings demonstrate that SmVAL proteins have localizations that place them in strategic positions to be considered as potential vaccine candidates as some members are exposed to interaction with the immune system and may participate in key processes of mammalian invasion and parasitism establishment.

Germs within Worms: Localization of Neorickettsia sp. within Life Cycle Stages of the Digenean Plagiorchis elegans.

Neorickettsia spp. are bacterial endosymbionts of parasitic flukes (Digenea) that also have the potential to infect and cause disease (e.g., Sennetsu fever) in the vertebrate hosts of the fluke. One of the largest gaps in our knowledge of Neorickettsia biology is the very limited information available regarding the localization of the bacterial endosymbiont within its digenean host. In this study, we used indirect immunofluorescence microscopy to visualize Neorickettsia sp. within several life cycle stages of the digenean Plagiorchis elegans Individual sporocysts, cercariae, metacercariae, and adults of P. elegans naturally infected with Neorickettsia sp. were obtained from our laboratory-maintained life cycle, embedded, sectioned, and prepared for indirect immunofluorescence microscopy using anti-Neorickettsia risticiihorse serum as the primary antibody. Neorickettsiasp. was found within the tegument of sporocysts, throughout cercarial embryos (germ balls) and fully formed cercariae (within the sporocysts), throughout metacercariae, and within the tegument, parenchyma, vitellaria, uteri, testes, cirrus sacs, and eggs of adults. Interestingly, Neorickettsia sp. was not found within the ovarian tissue. This suggests that vertical transmission of Neorickettsia within adult digeneans occurs via the incorporation of infected vitelline cells into the egg rather than direct infection of the ooplasm of the oocyte, as has been described for other bacterial endosymbionts of invertebrates (e.g.,Rickettsia and Wolbachia).

Additional study of the morphology of eggs and miracidia of Eurytrema coelomaticum (Trematoda)

Abstract The morphology of the eggs and miracidia of Eurytrema coelomaticum was analyzed. The eggshell presented different composition according to the maturation stage of the egg, as seen by distinct birefringence after polarized light microscopic analysis. Detailed morphology of the brownish eggs, measuring 44.97±3.83 (38.67 - 51.40) μm in length and 30.71±2.54 (25.79 -34.47) μm in width, and the different electrondensities of the inner, middle and outer layers of the eggs were determined using light (bright field and DIC) and electronic microscopy (scanning and transmission). The morphometry of the eggs is in accordance to those measures presented in previous studies. The miracidia were observed using light microscopy, inside and outside the egg. It was possible to observe the terebratorium at the anterior end of the body, the ciliated epidermal plates, placed according the formula E=2E1 + 2E2, and the interepidermal ridge, but eyespots were not observed. The excretory vesicle was laterally placed in the posterior middle of the body and the germ balls were observed. The miracidium inside the egg was observed by routine techniques, but the miracidium was not well preserved. Thus, the eggs were processed using high pressure-freeze substitution technique and reasonable preservation was achieved. The cilia covering the larval body, with a typical ciliary organization, the terebratorium with folds and the junction of the epidermal ciliated cell with the larval body were seen. For the first time detailed information on the eggs and miracidia of E. coelomaticum is given, which can be the basis for new studies.

Reproductive dysfunction in the edible oyster, Crassostrea madrasensis due to larval bucephalid infection – a case study

The present case study provides an insight into the tissue level pathological alterations inflicted by Bucephalus sp. infections in the edible oyster, Crassostrea madrasensis. Oysters collected during a routine study (from the oyster beds along the Southeast coast of India during July 2012) was found to harbor larval bucephalid infection. Though the prevalence of infection was low (1.37%), the intensity of infection was high, rated as two on the semi-quantitative scale. Infected oyster did not exhibit any external manifestation, but the developing parasitic stages, after replacing the host gonadal tissues have further spread to the gills and digestive glands. Regions of gonadal tissues were found packed with slender, branching sporocysts containing germ balls and crecariae, destroying the acinar architecture of the gonads. Immune responses like encapsulation against the infective stages were totally absent. The altered acinar architecture and reduced volume of the gonadal acini appeared to be insufficient to support any gametic release, resulting in gonadal dysfunction leading to parasitic castration of the host. An increase in the prevalence of Bucephalus sp. infection in the

Gene expression patterns in larval Schistosoma mansoni associated with infection of the mammalian host.

BackgroundThe infective schistosome cercaria develops within the intramolluscan daughter sporocyst from an undifferentiated germ ball, during which synthesis of proteins essential for infection occurs. When the aquatic cercaria locates the mammalian host it rapidly penetrates into the epidermis using glandular secretions. It then undergoes metamorphosis into the schistosomulum, including replacement of its tegument surface membranes, a process taking several days before it exits the skin. Patterns of gene expression underlying this transition have been characterised.Methods and Principal FindingsAll gene models from the S. mansoni genome (www.GeneDB.org) were incorporated into a high-density oligonucleotide array. Double-stranded cDNA from germ balls, cercariae, and day 3 schistosomula was hybridised to the array without amplification. Statistical analysis was performed using Bioconductor to reveal differentially transcribed loci. Genes were categorised on the basis of biological process, tissue association or molecular function to aid understanding of the complex processes occurring. Genes necessary for DNA replication were enriched only in the germ ball, while those involved in translation were up-regulated in the germ ball and/or day 3 schistosomulum. Different sets of developmental genes were up-regulated at each stage. A large number of genes encoding elastases and invadolysins, and some venom allergen-like proteins were up-regulated in the germ ball, those encoding cysteine and aspartic proteases in the cercaria and schistosomulum. Micro exon genes encoding variant secreted proteins were highly up-regulated in the schistosomulum along with tegument and gut-associated genes, coincident with remodelling of the parasite body. Genes encoding membrane proteins were prominently up-regulated in the cercaria and/or day 3 schistosomulum.Conclusions/SignificanceOur study highlights an expanded number of transcripts encoding proteins potentially involved in skin invasion. It illuminates the process of metamorphosis into the schistosomulum and highlights the very early activation of gut-associated genes whilst revealing little change in the parasite's energy metabolism or stress responses.

Morphology, development and probable systematic position of Cercariaeum crassum Wesenberg-Lund, 1934 (Digenea), a parasite of Pisidium amnicum in eastern Finland

Cercariaeum crassum Wesenberg-Lund, 1934 is redescribed at the cercariaeum stage and the daughter-rediae and cercaria are also described on the basis of new material from Pisidium amnicum collected in the Liikasepuro River (eastern Finland). The species is allocated to the family Allocreadiidae, although its generic affiliation remains unknown. The probable life-cycle (based on the developmental stages observed in daughter-redia) appears to eliminate the cercarial stage and, instead, a cercariaeum (a type of cercaria without a tail) may develop directly from germ balls or, rarely, through the stages of an ophthalmoxiphidiocercaria that transforms into a young caudate cercariaeum. Their morphology and development are shown to be consistent with the family Allocreadiidae. The probable lack of a second intermediate host in the life-cycle is discussed.

Saci-1, -2, and -3 and Perere, four novel retrotransposons with high transcriptional activities from the human parasite Schistosoma mansoni.

Using the data set of 180,000 expressed sequence tags (ESTs) of the blood fluke Schistosoma mansoni generated recently by our group, we identified three novel long-terminal-repeat (LTR)- and one novel non-LTR-expressed retrotransposon, named Saci-1, -2, and -3 and Perere, respectively. Full-length sequences were reconstructed from ESTs and have deduced open reading frames (ORFs) with several uncorrupted features, characterizing them as possible active retrotransposons of different known transposon families. Alignment of reconstructed sequences to available preliminary genome sequence data confirmed the overall structure of the transposons. The frequency of sequenced transposon transcripts in cercariae was 14% of all transcripts from that stage, twofold higher than that in schistosomula and three- to fourfold higher than that in adults, eggs, miracidia, and germ balls. We show by Southern blot analysis, by EST annotation and tallying, and by counting transposon tags from a Serial Analysis of Gene Expression library, that the four novel retrotransposons exhibit a 10- to 30-fold lower copy number in the genome and a 4- to 200-fold-higher transcriptional rate per copy than the four previously described S. mansoni retrotransposons [corrected]. Such differences lead us to hypothesize that there are two different populations of retrotransposons in S. mansoni genome, occupying different niches in its ecology. Examples of retrotransposon fragment inserts were found into the 5' and 3' untranslated regions of four different S. mansoni target gene transcripts. The data presented here suggest a role for these elements in the dynamics of this complex human parasite genome.

Schistosoma mansoni miracidia transformed by particle bombardment infect Biomphalaria glabrata snails and develop into transgenic sporocysts

Miracidia (and adults) of Schistosoma mansoni which had been subjected to particle bombardment with a plasmid DNA encoding enhanced green fluorescent protein (EGFP) under control of the S. mansoni heat shock protein 70 (HSP70) promoter and termination elements were shown to express the reporter gene. Bombarded miracidia were able to penetrate and establish in Biomphalaria glabrata the intermediate host snail. Gold particles could be detected in the germ balls of parasites in paraffin-sections of snail tissue. The bombarded miracidia were able to develop normally and to transform into mother sporocysts. Reporter gene activity could be determined at 10 days post-infection by RT-PCR in snail tissues, but not by microscopy or Western blot which probably reflected sub-optimal expression levels of constructs. Our findings indicated that it is feasible to return transgenic miracidia to the life cycle, a crucial step for the establishment of a transgenesis system for schistosomes. Index Descriptors and Abbreviations: schistosome, miracidium, sporocyst, particle bombardment, gold particles, transgene, Biomphalaria glabrata; germ ball; EGFP, enhanced green fluorescent protein; RT-PCR, reverse transcription-polymerase chain reaction; gDNA, genomic DNA; PBS, phosphate buffered saline; TBS, Tris buffered saline; FCS, fetal calf serum; DEAE, diethylaminoethyldextran

Some specific and non-specific phosphatases of the sporocyst of Fasciola hepatica. II. Enzymes associated with the membrane transport.

Using histochemical and cytophotometric methods, enzymes responsible for the membrane transport (alkaline phosphatase, adenosine triphosphatase, and 5-nucleotidase) in the developing sporocyst of Fasciola hepatica (L., 1758) were studied. The most active metabolism occurred in the germ balls of sporocysts on the 8th and 15th days of development, which is associated with intensive proliferation and subsequently differentiation of embryos within the germ balls.

Radix auricularia coreana: natural snail host of Clinostomum complanatum in Korea.

An epidemiological survey was carried out to determine the first intermediate host of Clinostomum complanatum among freshwater snails in Korea. Two species of snails belonging to the family Lymnaeidae were collected in Kaum-ji (pond), Uisong-gun, Kyongsangbuk-do. Twelve (0.9%) out of 1,273 Radix auricularia coreana examined were found to liberate cercariae of C. complanatum, which were identified by morphological characteristics and experimental infections in freshwater fish. Pseudorasbora parva. The cercariae were brevifurcate and clinostomatoid. They had a transparent dorsal fin, a well developed penetrating organ and a pair of eye spots. The body measured 119-147 x 33-36 microns, tail stem, 275-370 x 19-26 microns, and furcae, 72-104 microns. Rediae were demonstrated in the infected snail after crushing. Redia, 527-1,630 microns long and 121-368 microns wide, contained 10-45 germ balls and cercariae in various developmental stages. The metacercariae recovered from fish experimentally infected with C. complanatum cercariae were morphologically identical to those from naturally infected fish.

Paramphistomum daubneyi: the development of redial generations in the snail Lymnaea truncatula.

Rediae of Paramphistomum daubneyi were counted and measured in Lymnaea truncatula to elucidate the variability in the numbers of free rediae and cercariae occurring between naturally infected snails and experimental single-miracidium infections. Experiments were performed using one miracidium per snail and snail raising was carried out at 20 degrees C. Two redial generations succeeded each other in the snail until day 49. A mean of 8-10 rediae differentiated in the sporocyst at days 7 and 14; the remaining germ balls and redial embryos decreased in number after day 21. First-generation rediae became free in the snail's body starting at day 14 and their number increased to a mean of 7.5 at day 49, with a maximum of 10 rediae being detected in 1 snail. They produced second-generation rediae, which exited from the body starting at day 28, and then cercariae, which exited from day 42 onward. The count of second-generation rediae was 6-6.2/snail at day 49, with a maximum of 12 being detected in 1 snail; they produced only cercariae.

Echinostoma macrorchis: Life History, Population Dynamics of Intramolluscan Stages, and the First and Second Intermediate Hosts

Eggs of Echinostoma macrorchis were laid at the 1-cell stage and took 9 days (26-27 C) or 6 days (30 C) to hatch. The following accounts are based on studies undertaken at 24-26 C. Maximum survival of miracidia was 7 hr. Miracidia reached the ventricle of Gyraulus chinensis 15-20 hr after penetration and transformed into sporocysts. The earliest degeneration of sporocysts occurred 20 days postinfection (DPI), but some survived for 120 days. Mature mother rediac were first seen inside the sporocyst at 7 DPI, then were released to the ventricular cavity and migrated to other anatomical locations of the host. Among the germ balls present in a sporocyst, 1-3 developed concurrently to the mother redia stage. Young daughter rediae first appeared at 15 DPI and mature ones at 19 DPI. Snails of larger size produced more daughter rediae and cercariae than smaller ones. Initial shedding of cercariae from infected snails occurred between 25 and 48 DPI, with a mean of 34 DPI. The cercariae were phototactic and each snail shed a daily average of 34 cercariae. Gyraulus chinensis was proven to be the only snail capable of serving as the first intermediate host. Tadpoles of Rana latouchi and Bufo bufo, 9 species of gastropods, the bivalve Corbicula fluminea, and 1 unidentified planarian species served as experimental second intermediate hosts.(ABSTRACT TRUNCATED AT 250 WORDS)

Lecithochirium furcolabiatum (Jones, 1933), Dawes 1947: the miracidium and mother sporocyst

Experimental infections of the marine topshell Gibbula umbilicalis with Lecithochirium furcolabiatum (Digenea: Hemiuroidea) have allowed the development of a model system which will enable further studies of the molluscan host response. The long-lived intertidal prosobranch host is easily maintained in the laboratory, and experimental infection rates of 98% were consistently achieved. The miracidium and mother sporocyst have been studied at both light and ultrastructural levels, providing the first account of the morphology of these stages in Hemiuridae. The ingested egg hatches within the host intestine, treatment with L-cysteine and alkaline pH stimulating miracidial emergence in vitro. The general body surface of the miracidium is devoid of spines or cilia, the latter being restricted to four plates near the anterior extremity. The miracidium swims actively prior to penetration of the gut wall, the sporocyst being released from the miracidial epidermal coat within the haemocoel. Within 5 weeks of infection, the filamentous mother sporocyst contains 1 to 3 oval germ balls, daughter sporocysts being recorded free within the digestive gland haemocoel 7 weeks later. Twenty three weeks after ingestion of eggs, the daughter sporocyst extends into the host gill filaments, containing cystophorous cercariae ready for emergence.

The Larvae That Never Develop

In most animals other than vertebrates, an embryo develops into a motile larva that feeds and then develops (either gradually or suddenly) into an adult. There are some, however, in which the larva itself reproduces before it develops. This reproduction is generally asexual, by fragmentation or budding, and it does not interfere with the life cycle. A strikingly different form of larval reproduction occurs in many trematode flatworms. In these, at each of two or three successive larval stages, more individuals (larvae) appear. In general, a miracidium larva develops into a sporocyst, which gives rise to many rediae, each of which gives rise to many cercariae. The cercariae develop into adults, but the sporocysts and rediae all eventually die, after the escape of their offspring. There is much diversity in the details, and even in the number of successive larval stages (which may include a second generation of sporocysts and at least two extra stages of daughter rediae). It was once unclear how these larvae are produced, but it is now seen that there is no reproduction by any of these larvae, merely cell multiplication of an unusual sort. In the miracidium there is a mass of germ cells, often called a germ ball, derived from a single cell in the early embryo. The germ ball divides into daughter balls, which remain when the miracidium transforms into a sporocyst. Then each daughter germ ball forms a redia around itself, which thus contains a remnant of the original germ ball. At each successive larval stage (redia or cercaria), the new individuals are produced by one of the fragments of the germ ball, and each contains its own part of that earlier germ ball. Thus, this sequence of larval forms is not produced by one metamorphosing into another, nor by one producing the next generation by asexual reproduction. Instead, every larva after the sporocyst (the second larval stage) is produced by the original germ ball and its fragments. Each larva acts as a carrier for a germ ball that will itself produce the next stage by a sort of embryonic development. These later larvae are actually dead ends, and the continuity of the life cycle is solely through the germ balls that they carry temporarily. The first germ ball is really an early embryo of undifferentiated cells. Its division into several germ balls is similar to a well-known process that produces identical twins. This is polyembryony, production of many embryos by the break-up of the one early embryo that resulted from sexual reproduction. In these trematodes the germ ball in the embryo and early larva undergoes this polyembryony repeatedly, in the sense that each fragment in the sporocyst divides several times to form the fragments that become the rediae. And these fragment again to form the cercariae. This succession of fragmentations gives rise to the term successional polyembryony. Polyembryony occurs in many of the major phyla, particularly in insects, where as many as several thousand fragments may develop into new individuals. It also occurs in mammals: the nine-banded armadillo, for example, always produces four embryos, all genetically identical. Even identical twins in humans are formed by this process, which is one of the many forms of asexual reproduction. However, nothing as highly complicated as the successive fragmentations of the trematode germ ball into successively different larval forms is known in other animals. Polyembryony in its various forms is one of the unique and fascinating occurrences in the development of animals.

Bucephaloides gracilescens: A quantitative study of the lysosomal cellular stress response induced by the sporocysts and developing cercariae within the white furrow shell,Abra alba

Reproductive stress is apparent inAbra alba as a result of infection with the sporocysts ofBucephaloides gracilescens, culminating in castration in heavily infected specimens. The bivalve is also subject to mechanical stress from actively growing sporocyst tubules and nutritional stress due to the nutrient requirement of large numbers of germ balls within the sporocysts. Using the digestive cell lysosomal system ofAbra as a monitor, it was possible to demonstrate quantitatively a parasite-induced cellular stress response by applying a sensitive cytochemical test for lysosomal stability. Lysosomal stability was determined as the labilisation period for latent Nacetyl-β-hexosaminidase (NAH), measured by microdensitometry. In uninfectedAbra, digestive cell lysosomal NAH expressed structure-linked latency. Hence a significantly longer labilisation period was required compared with infectedAbra, where the parasitic burden with its associated stress effects resulted in a destabilisation of the lysosomal membrane. This reduced the latency of the enzyme, so that a much shorter labilisation period was required for the stressed tissue to express maximum lysosomal enzyme activity. It is suggested that the lysosomal system of the digestive cells inAbra can be used as a sensitive monitor of the stress induced by the sporocysts and developing cercariae ofBucephaloides.

The origin and development of the epidermis and associated structures in the cercaria of Cryptocotyle lingua (Creplin) (Digenea: Heterophyidae) from Littorina littorea (L.).

Cercariae of Cryptocotyle lingua develop from intraredial germinal cells which divide to form ‘naked’ cell aggregates and later germ balls covered, first, by a syncytial primitive epithelium and later a syncytial epidermis formed, in sequence, from superficial cells of the embryo. The primitive epithelium is soon lost. The original nuclei of the syncytial epidermis degenerate when the first series of epidermal cell bodies, formed immediately below and having the characters of protein synthesizing cells, become connected with it. The first cell bodies are replaced by a series of five types of epidermal (secretory) cell bodies developing in the parenchyma and giving off cytoplasmic processes which become connected, in sequence, with the outer cytoplasmic layer. Secretion bodies from four of the five types are discharged into the outer cytoplasmic layer, before the cercaria leaves the molluscan host, and remain there in the free swimming cercaria. The secretions of the fifth type are retained in their epidermal (secretory) cell bodies. The arrangement of the secretion bodies in the outer cytoplasmic layer and their histochemical reactions suggest possible functions concerned, later, with entry into and encystment within the second intermediate fish host.

Autoradiographic studies on the uprake of (14C)-glucose by Bithynia tentaculata (mollusca : Gastropoda) and its larval digeneans.

AbstractAutoradiographs revealed that sporocysts of Cercaria helvetica XII and rediae of Sphaeridiotrema globulus, incubated in a medium containing [14C]-glucose, exhibited slight radioactivity in their body wall after only 30 seconds incubation. Activity in sporocysts and rcdiae gradually increased as the incubation period was extended until after 4 minutes, a heavy reaction was observed in the parasite body wall as well as in the germ balls and developing cercariae within the parasite brood chambers. The redial caecum appeared to incorporate the labelled compound much more intensely than did the redial body wall.When uninfected Bithynia tentaculata, as well as B. tentaculata infected with either S. globulus or C. helvetica XII, were maintained in pond water containing [14C]-glucose, activity could be detected in the gills of both infected and uninfected snails after 5 hours incubation. After 12 hours a much heavier labelling was apparent in the gills, together with moderate activity in the haemocoelic spaces, foot musculature, stomach wall, and gut contents. After 20 hours a more intense labelling was apparent in these regions of the snail body and, in addition, some activity could be detected in the digestive gjand tubules of the host and, in infected snails, in the bodies of developing parasites. After 30 hours incubation the labelled compound had become incorporated into most of the host tissues but was most intense in the digestive gland tubules. Heavy activity was also evident in the parasites at this stage although certain cystogenous gland cells of the cercariae of both parasite species did not incorporate the labelled compound.

Himasthla quissetensis: Uptake and utilization of glucose by rediae as determined by autoradiography and respirometry

Daughter rediae of Himasthla quissetensis removed from the digestive gland of Nassarius obsoletus were placed in sterilized seawater fortified with antibiotics. When [ 3H]-glucose was added to this medium and autoradiographs were made after 3, 9, and 24 hr of exposure, labeling was observed associated with the redial walls and developing germ balls and cercariae within the brood chambers. Respirometric determinations on starved rediae suspended in the seawater medium with and without glucose revealed the rate of oxygen utilization by rediae exposed to exogenous glucose is significantly elevated. These results are interpreted to mean that the daughter rediae of H. quissetensis can take up and utilize glucose.