Gastropod Biomphalaria Glabrata Research Articles

Analysis of some factors that may modify the bioavailability of cadmium and lead byBiomphalaria glabrata

Laboratory acute bioassays (t = 96 h) were used to determine the uptake, distribution, and elimination of cadmium and lead by the freshwater gastropod Biomphalaria glabrata. Experiments were conducted on nonpigmented and pigmented organisms separately. The influence of food and different concentrations of humic acids were also investigated. The results showed that, after the treatments, the digestive gland presented the highest level of metal uptake for both elements. Food did not prove to induce significant changes in the pattern of accumulation and distribution within the different tissues. Instead, different concentrations of humic materials modified the pattern of accumulation and distribution of cadmium and lead but in a different way according to the metal. Elimination of cadmium from the soft tissues was considerably slow, presenting a redistribution over the time of depuration. Elimination of lead was faster in the digestive gland and gonads. After all the treatments, no significant differences were observed between nonpigmented and pigmented snails.

Production of reactive oxygen species by hemocytes of Biomphalaria glabrata: carbohydrate-specific stimulation

Recognition of specific carbohydrate structures, which occur commonly on the surfaces of invading pathogens, is thought to elicit internal defense mechanisms in invertebrates. To investigate the nature of carbohydrates that evoke a defensive response in hemocytes of the gastropod Biomphalaria glabrata, we tested eight different carbohydrates, conjugated to bovine serum albumin (BSA), for generation of reactive oxygen species (ROS). Six of the carbohydrate moieties tested are thought to be present on the S. mansoni sporocyst surface (mannose, galactose, fucose, N-acetyl-glucosamine, N-acetyl-galactosamine, and lactose); the other two carbohydrates tested were glucose and melibiose. ROS generation was measured using the fluorescent probe — 2′,7′-dichlorofluorescein-diacetate (DCFH-DA). Hemocytes were derived from two different strains of B. glabrata: one of the strains used (MO) is susceptible to infection by the trematode Schistosoma mansoni (PR-1 strain), while the other snail strain (13–16-R1) is resistant to infection with PR-1. Three of the BSA-carbohydrate conjugates (BSA-galactose, BSA-mannose, and BSA-fucose), stimulated generation of reactive oxygen species in the molluscan hemocytes. The responses of the hemocytes were similar whether they were derived from susceptible or resistant snails. If the carbohydrate structures we found, to stimulate ROS generation are involved in parasite recognition, our results suggest that parasite killing may involve either qualitative differences in production of reactive oxygen species, or additional factors.

Molecular studies of Biomphalaria glabrata, an intermediate host of Schistosoma mansoni

The freshwater gastropod Biomphalaria glabrata is one of the most important invertebrate hosts of the helminth parasite Schistosoma mansoni. Investigators are using different strategies to determine the molecular basis of this snail–parasite relationship. Of particular interest are the identification of parasite resistance genes in the snail, and the application of molecular probes to better understand the epidemiology of schistosomiasis. This review will focus on recent advances that have been made on genome analysis of B. glabrata. Much of this work has centred on the use of random amplification of polymorphic DNA–PCR-based technology, with restriction fragment length polymorphism analysis and the generation of expressed sequence tags from the snail. A brief discussion of how parasite products may complicate this analysis is also given, along with an indication of the scope of the problems that lie ahead.

Effect of temperature on survival and infectivity of Echinostoma trivolvis cercariae: a test of the energy limitation hypothesis

SUMMARYTrematode cercariae typically become unable to successfully infect a host many hours before they die. We examined the hypothesis that both time to 50% mortality and time to loss of infective capacity are controlled to the same degree by rates of energy expenditure, by determining the relative effects of temperature on both parameters. Infective capacity was assessed by exposing Echinostoma trivolvis cercariae of different ages to a suitable second intermediate host (the gastropod Biomphalaria glabrata) and counting 1–2 days later the number of metacercarial cysts formed. Temperature had a remarkably similar effect on time to 50% mortality and loss of infective capacity, supporting the hypothesis that both absolute and functional cercarial life-spans are limited by the rates at which energy stores are utilized.

Characterization of immunoreactive TNFα molecules in the gastropod Biomphalaria glabrata

In recent years, several studies have demonstrated the existence of cytokine-like molecules in invertebrates, therefore suggesting that cytokines may have been conserved throughout evolution. In this study, we investigated the presence of immunoreactive TNFα (ir TNFα) in the gastropod mollusc Biomphalaria glabrata, the specific intermediate host for the trematode Schistosoma mansoni. Immunocytochemical study indicated the presence of ir TNFα in mollusc hemocytes corresponding to a 53-kDa molecule detected by western blot analysis. Using ELISA tests, we demonstrated the presence of substantial amounts of ir TNFα in hemolymph, that were significantly decreased during the S. mansoni infection. The possible role of ir TNFα in the regulation of mollusc immune functions and in the host-parasite relationship is discussed.

Carbohydrate-binding plasma proteins from the gastropod Biomphalaria glabrata: Strain specificity and the effects of trematode infection

As lectins are believed to mediate non-self-recognition in molluscs, carbohydrate-binding proteins (CBP) from the circulating plasma of the gastropod Biomphalaria glabrata were harvested by affinity chromatography using six different monosaccharides as ligands. Pools of plasma were derived from B. glabrata of either the M line strain, which is susceptible to infection with the PR1 strain of the digenetic trematode Schistosoma mansoni, or from the 13-16-R1 strain, which is resistant to infection. For each strain, plasma was obtained from control snails and from snails exposed to infection 1 or 8 days previously with S. mansoni or the related digenean, Echinostoma paraensei, which is able to develop in either host strain. For control snails, only minor interstrain differences were noted. In M line snails exposed 8 days previously to either parasite, marked changes in CBP populations were observed. Only E. paraensei infections produced comparable alterations in 13-16-R1 snails. The most conspicuous changes noted were the increased production of 80–120 kDa CBP in both strains, 150–210 kDa in 13-16-R1 snails, and 190–210 kDa in M line snails. The results demonstrate 1) interstrain differences in CBP, particularly following exposure to trematodes; 2) that infection provokes increased production and diversity of CBP that bind with greater affinity to the columns; and 3) that snails of the same strain respond differently to the two parasites used.

The site of ultrafiltration in the kidney sac of the pulmonate gastropod Biomphalaria glabrata

Ultrastructural observation of the renopericardial region in B. glabrata revealed the presence of a site for ultrafiltration located in the proximal region of the kidney sac, in the vicinity of the pericardium. This region comprises podocyte-like cells covering large hemal spaces supplied by a renal artery. Extracellular material observed in spaces between podocyte foot processes resembled small septate junctions rather than diaphragms such as observed in various kidneys. The effective ultrafiltration barrier may be the thick basal lamina of the podocytes. The hemal spaces have a connective tissue frame rich in fixed phagocytes and migrating hemocytes which may play a role in the internal defence system of the snail.

Biological control studies: Bacteria associated with moribund Biomphalaria glabrata (Mollusca) in the laboratory

The bacteria associated with moribund specimens of the gastropod Biomphalaria glabrata were identified over a 4-year period. A total of 12 species and one or more representatives of Enteric Group 17 were found. Among these, Citrobacter freundii, a common aquatic species, was associated with 99%, and Aeromonas hydrophila, a sometimes pathogenic species, was associated with 66% of the moribund snails.

Biomphalaria glabrata amoebocytes: Assay of factors influencing in vitro phagocytosis

A simple and short-term in vitro assay system has been adapted for quantitative evaluation of the rate of phagocytosis of amoebocytes obtained from hemolymph of the pulmonate gastropod Biomphalaria glabrata. The variation in the rate of phagocytosis exhibited by amoebocyte monolayers prepared from individual snails is reduced by utilizing amoebocytes from pooled hemolymph from snails of similar sizes. The study demonstrates that the rate of phagocytosis depends on incubation time, temperature, and pH. Substantial inhibition of phagocytosis is exerted by 2-deoxy- d-glucose and 2-iodoacetamide but not by potassium cyanide, indicating that energy consumed during phagocytosis is generated by the glycolysis pathway.

Edema-horn, an abnormal mutant of Biomphalaria glabrata

An abnormal condition is reported in the pulmonate gastropod Biomphalaria glabrata involving body edema and flaring of the shell aperture. Selection and selfing of isolated snails suggested this character is inherited. Crossing experiments indicated the character is inherited as a simple recessive.

Leucocytosis in Biomphalaria glabrata sensitized and resensitized to Echinostoma lindoense

Leucocytosis was shown to occur in the pulmonate gastropod Biomphalaria glabrata exposed to the trematode Echinostoma lindoense. In these sensitized snails, the leukocyte count in the hemolymph was elevated 3 to 5 days postexposure to miracidia, and prior to complete encapsulation of sporocysts. This increase continued 1 to 5 days after destruction of sensitizing, irradiated E. lindoense sporocysts. Counts returned to normal levels after this period. A significant and more rapid increase in numbers of circulating leukocytes occurred 1 to 6 hr after reexposure of snails to a sensitizing dose of nonirradiated E. lindoense sporocysts. The leukocyte counts usually returned to normal levels after this period, except in snails in which some resensitizing sporocysts remained alive.

Experimentally induced elevations in acid phosphatase activity in hemolymph of Biomphalaria glabrata (Mollusca)

The levels of acid phosphatase activity in the hemocytes and serum of the following four groups of the pulmonate gastropod Biomphalaria glabrata were ascertained: (1) those injected with heat-killed Bacillus megaterium, (2) those challenged with sterile distilled water, (3) those shaminjected, and (4) those left untampered. It was determined that challenge with heat-killed bacteria sesulted in significant elevations in acid phosphatase activity in both cells and serum at 1, 2, and 4 hr postinjection, with the level being highest at 2 hr. Injection with water resulted in a significant elevation in cellular enzyme level at 1 hr postinjection but not at 2 and 4 hr; however, there were significant elevations at 2 and 4 hr in serum. This is interpreted to indicate that the elevated intracellular enzyme was subsequently released into serum. Sham injection resulted in significant elevations in acid phosphatase levels in hemocytes at 2 and 4 hr postinjection but no increase of enzyme activity in serum during the course of the experiment. This is interpreted to mean that this hydrolase was not released as a result of sham injection. The source of acid phosphatase was apparently the cytoplasmic granules of granulocytes, which are true lysosomes.

Aminopeptidase and lysozyme activity levels and serum protein concentrations in Biomphalaria glabrata (Mollusca) challenged with bacteria

The total serum protein concentrations and levels of aminopeptidase and lysozyme activities in the sera of the gastropod Biomphalaria glabrata have been determined. The groups of snails from which hemolymph samples were taken for study included (1) untampered controls, (2) sham-injected snails, (3) heat-killed Bacillus megaterium-injected, and (4) live B. megaterium-injected ones. Our results indicate that there are significant elevations in the levels of aminopeptidase activity in 2 hr in the sera of snails that had been sham-, dead bacteria-, and liver bacteria-injected. The levels of lysozyme activity were not altered in sham-, dead bacteria-, and live bacteria-injected snails. This is contrary to an earlier finding ( T. C. Cheng, M. J. Chorney, and T. P. Yoshino, 1977. J. Invertebr. Pathol., 29, 170–174), and the difference is believed to be due to the age of the snails employed. Comparisons of total serum proteins have revealed that the concentration in snails injected with live B. megaterium is significantly higher than in sham-injected ones. This may be due to increase of some yet undetermined serum protein fraction.

Hemocytes of the pulmonate gastropod Biomphalaria glabrata

Scientific researches have clearly indicated that ocean acidification and warming poses serious threats to coral reef ecosystems. In coral reef ecosystems, herbivorous gastropods have an important function in maintaining the stability of the ecosystem due to controlling the abundance and growth of macroalgal, which compete for nutrients and space with coral. However, limited knowledge is available on the physiological responses of the specific keystone species to the increased ocean acidity and thermal stress. In this study, we evaluated the effects of ocean acidification (OA) and warming (OW) on an herbivorous gastropod Trochus niloticus commonly found on intertidal and shallow subtidal coral reefs in the South China Sea, on the aspect of immune responses (total hemocyte counts, reactive oxygen species level and apoptosis rate), oxidative stress (lipid peroxidation level, antioxidant enzyme activities), neurotoxicity (acetylcholinesterase activity), and energy metabolism (respiration rate and cellular energy allocation), after a 28-day exposure experiment to acidic (pH 7.6) and/or thermal (30 °C) seawater. Our results demonstrated that both OA and OW could lead to physiological disturbances of the herbivorous top-shells, including impaired immune functions and oxidative balance, neurotoxicity, and disorder of energy metabolism. Furthermore, results of integrated biomarker response (IBR) confirmed that the overall fitness of T. niloticus were deleteriously impacted by OA and OW, and were more stressed under the co-exposure condition. These results indicated that increased acidity and temperature in the future ocean might impair the viability of T. niloticus in the long-run, which will indulge the proliferation of macroalgae and lead to degradation of the coral reef ecosystem.

Aminopeptidase activity in the hemolymph and body tissues of the pulmonate gastropod Biomphalaria glabrata

The levels of aminopeptidase activity in the whole hemolymph, serum, hemocytes, headfoot, and visceral mass of Biomphalaria glabrata were determined. The highest enzyme level occurs in the serum and the lowest in the hemocytes. Both the headfoot and visceral mass include subequal levels of aminopeptidase activity. From our data, it now appears possible that the serum aminopeptidase in B. glabrata, which has an open circulatory system, could have originated in hemocytes as well as in other tissues. The biologic function of serum aminopeptides is uncertain; however, because of the known chemical function of this enzyme, it could serve to degrade foreign proteins in serum prior to phagocytosis.

Alterations in the osmoregulation of the pulmonate gastropod Biomphalaria glabrata due to copper

Specimens of Biomphalaria glabrata were exposed to 0.06 ppm of copper in the form of CuSO 4, and the resulting changes in the wet and dry weights of the soft tissues and in the osmolality of the hemolymph were measured. The wet weights of snails exposed to copper increased as a function of time, while those of the controls decreased. The dry weights of both the experimental and control snails decreased equally. Finally, the ratio of wet weight to dry weight of the experimental snails was significantly higher than that of the controls after 24 and 48 hr of exposure to copper. In addition, the osmolality of the hemolymph of snails exposed to copper was significantly lower than that of the controls after 12, 24, and 36 hr of exposure. These data have led to the conclusion that exposure of B. glabrata to copper results in an osmotic influx of water into its tissues and thereby causes death.

The encapsulation process in Biomphalaria glabrata experimentally infected with the metastrongylid Angiostrongylus cantonensis: Enzyme histochemistry

Enzyme histochemistry has revealed that encapsulation reactions surrounding larvae of the nematode Angiostrongylus cantonensis 4 wk after infection of the gastropod Biomphalaria glabrata are the sites of highly localized acid phosphatase and nonspecific esterase activities. Lesser amounts of alkaline phosphatase and β-glucuronidase activities also occur within such capsules, but aminopeptidase activity cannot be demonstrated. In addition, it has been ascertained that acid phosphatase activity gradually increases as the encapsulation reaction progresses during the first to the fourth week postinfection.

5-Hydroxytryptamine and Dopamine in Biomphalaria glabrata

5-hydroxytryptamine (5-HT) and dopamine (DA) have been isolated and identified in the tissues of the snail, Biomphalaria glabrata. The brain contained approximately 15 tAg, the cephalopedal region 2 ,ug, and the mantle region 1 jug of 5-HT per gram of fresh weight. The concentrations of DA in the brain and the cephalopedal region were of the same order of magnitude as those of 5-HT, and 5 times lower in the mantle region. Neither 5-HT nor DA were detectable in the hepatopancreas-ovotestis region. The distribution of these 2 biogenic amines was also studied by the histochemical fluorescence technique of Falck and Owman (1965). Only green (catecholamine) fluorescent neurons were observed in the buccal ganglia; they are tentatively considered as DA neurons. Neuropile areas probably contain a mixture of 5-HT and DA. The nerve fibers emanating from the ganglia predominantly contain DA. With the exception of the buccal ganglia, 5-HT neurons were more numerous and comparatively large in the other ganglia. Green catecholamine fluorescence was detected in areas nearest to the epithelium of the foot. The presence of two biogenic amines, 5hydroxytryptamine (5-HT) and dopamine (DA), has been demonstrated in a variety of gastropods (Carpenter et al., 1971; Cottrell and Maser, 1967; Dahl et al., 1966; Gerschenfeld, 1971; Sweeney, 1963; Twarog, 1968; Walker et al., 1968; Kerkut and Cottrell, 1963; Kerkut et al., 1966). These amines are localized predominantly in the nervous system of this class of animals, and evidence for their function as neural transmitters has been reported (Carpenter et al., 1971; Dahl et al., 1966; Gerschenfeld, 1971; Gerschenfeld and Stefani, 1968; Sweeney, 1963; Twarog, 1968). The gastropod Biomphalaria glabrata is an intermediate host of the parasitic trematode, Schistosoma mansoni. 5-HT and norepinephrine have been identified as constituents of the adult form of this parasite (Bennett et al., 1969; Bennett and Bueding, 1971). Furthermore, recent evidence indicates the presence of dopamine receptors in the longitudinal muscle of adult S. mansoni, a function of 5-HT as an excitatory neuromuscular transmitter in the parasite. (T. Tomosky et al., 1974), and a high affinity uptake Received for publication 2 November 1973. * This investigation was supported by NIH training grant AI-149, and research grants AI-03515 and AI-08022. t Present address: National Institute of Child Health and Human Development, Gerontology Research Center, Baltimore City Hospitals, Baltimore, Maryland 21224. mechanism for 5-HT providing a neurotransmitter which the adult worm cannot synthesize de novo. The question arises whether biogenic amines play a role in the development of the larval stages of schistosomes in their intermediate host; this, in turn, requires information about the distribution of these amines in B. glabrata. The present study is concerned with this problem. MATERIALS AND METHODS Biomphalaria glabrata with shell diameters of 9 to 14 mm were used. Determination of biogenic amines The tissues were dissected, quickly frozen in liquid nitrogen, and homogenized in a solution containing perchloric acid (0.4 M), ascorbic acid (10 mM), and EDTA (5.4 mM) (Anden and Magnusson, 1967) in a motor-driven all-glass homogenizer at 4 C. One ml of solution was used per 100 mg tissue. The mixture was transferred into a centrifuge tube and, after washing of the container with a measured volume of the same solution, was centrifuged at 10,000 g for 15 min at 4 C. The pH of the supernatant solution was adjusted to 5 to 6 by addition of 0.5 ml of 2 M potassium phosphate buffer (pH 6.5) and a measured volume of 2 N KOH. After centrifugation (10,000 g, 15 min, 4 C) the supernatant solution was used for the determinations of 5-HT and of DA.

Cytochemical Observations on the Midpiece of the Spermatozoa of the Gastropod Biomphalaria glabrata

Cytochemical Observations on the Midpiece of the Spermatozoa of the Gastropod Biomphalaria glabrata