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Praziquantel treatment of Biomphalaria glabrata infected with Schistosoma mansoni--influence on snail fecundity.

Praziquantel, administered over a 72 h period in the food of mature Biomphalaria glabrata harbouring 7-week-old Schistosoma mansoni infections, dramatically reduced the numbers of cercariae shed. Doses of 20-30 micrograms/g body weight (including shell weight) reduced shedding by 85-95% over 5 weeks before recovery was evident. Suppression of cercarial shedding in these infections was accompanied by the temporary recovery of the snail reproductive regression due to S. mansoni infection. Snail fecundity was subsequently re-suppressed 2 weeks prior to recovery of the parasite as evidenced by a resumption of cercarial shedding. Praziquantel destroyed mature and developing cercariae within the daughter sporocysts but had no apparent effect on daughter sporocysts; this may account for the eventual resumption of cercarial production. Reproductive failure of the snail is apparently related to the latter stages of cercarial development specifically. Reproductive recovery did not occur when snails were infected as juveniles or when mature-infected snails harboured 12.5 week i.e. older infections: drug treatment temporarily inhibited cercarial production but no snails produced eggs.

Animals↗

Interactions between the plasma proteins of Biomphalaria glabrata (Gastropoda) and the sporocyst tegument of Schistosoma mansoni (Trematoda).

The tegumental surface of Schistosoma mansoni sporocysts is the site of both nutritive and immunological interactions with haemolymph cells and plasma of Biomphalaria glabrata, the schistosome intermediate host. Within minutes of being placed in host plasma, sporocysts acquire plasma antigens, and within 3 h host plasma antigens are present on the surface at near steady state. Though a wide variety of peptides is acquired, there is selection. Furthermore, some differences occur in the peptides acquired from the plasma of susceptible and resistant strains of snail. Acquired antigens are rapidly processed, and are predominantly undetectable in tegumental extracts after a few hours. In contrast, rabbit antibody on sporocysts remains in situ for at least 48 h, so under some conditions there is stable expression of certain tegumental antigenic determinants. These data, obtained using antibodies to snail plasma antigens and to sporocyst tegumental antigens, are discussed in the light of current ideas on the cellular and molecular basis of susceptibility and resistance in this host-parasite system.

Adhesiveness↗

Elevated agglutination titres in plasma of Biomphalaria glabrata exposed to Echinostoma paraensei: characterization and functional relevance of a trematode-induced response.

Production of elevated haemolymph agglutination titres by Biomphalaria glabrata following exposure to Echinostoma paraensei miracidia was investigated, to characterize this parasite-induced response and to understand its functional relevance. Both the dose of infection (1, 10 or 100 miracidia per snail) or the number of separate exposures to infection (between one and three, over a 4 or 8 day interval) were varied, and assuming a threshold dosage (10 miracidia per snail or higher) was exceeded, titres of juvenile snails peaked at 8-16 times the values for unexposed control snails, regardless of the exposure regimen. Adult snails, which are relatively refractory to infection, have slightly higher resting titres than juveniles, but exhibit only a 2- to 4-fold increase in titre following exposure. Juveniles exposed to infection but lacking demonstrable infection had lower titres than snails with confirmed infections. Exposure to infection increased heterogeneity of plasma agglutinins and provoked production of unique specificities not found in unexposed snails. However, the overall pattern of agglutination responses for snails with successfully developed parasites did not differ from those in which parasite development was unsuccessful. Agglutinating activity was inhibitable by several different monosaccharides, although plasma from infected snails was relatively unaffected by N-acetyl-glucosamine or N-acetyl-galactosamine. Wounding of snails provoked no change in plasma agglutination activity. As the highest agglutination titres were produced in snails with successfully developing parasites and agglutinin composition did not differ between snails with successful or unsuccessful parasites, the functional relevance of the response remains enigmatic. The production of unique agglutinins following exposure deserves additional study.

Agglutination↗

Biochemical and morphological pathology of the foot of the schistosome vector Biomphalaria glabrata infected with Schistosoma mansoni.

Infection by Schistosoma mansoni resulted in morphological and biochemical changes to the foot of its intermediate host, Biomphalaria glabrata. Migration through, and emergence of cercariae from, the foot was observed and evidenced by lesions on the ciliated foot surface. This was accompanied by a significant decrease in the velocity of movement by infected individuals. In vivo 31P NMR spectral analyses demonstrated that the foot of infected snails had a lower phosphoarginine (PA)/adenosine 5' triphosphate (ATP) ratio than that of uninfected controls. Moreover, kinetic experiments, employing saturation transfer, demonstrated the pseudo-first-order rate constant for the arginine kinase-catalysed exchange reaction in the forward direction, that is, PA-->ATP was decreased by infection. The reverse reaction was not observed by the NMR methods used. PA was depleted upon exposure to hypoxic conditions suggesting its traditional role in preserving ATP level. Partly oxidized metabolic end-products were not observed in snails maintained under aerobic conditions, but succinate, propionate, acetate and lactate rapidly accumulated under hypoxic conditions.

Adenosine Triphosphate↗

Increased oviposition and growth in immature Biomphalaria glabrata after exposure to Schistosoma mansoni.

Biomphalaria glabrata snails are known to be castrated by infection with the trematode parasite Schistosoma mansoni 4-6 weeks post-infection. The pattern of oviposition in the first 35 days post-exposure (p.e.) was investigated, in snails aged 14 weeks and measuring 7-10 mm diameter which had not commenced egg-laying, by counting the numbers of eggs laid in 7-day intervals. A group of exposed snails was compared with a control non-exposed group. The exposed group included both parasitized and non-parasitized snails, and showed a significant increase in the median number of eggs laid during the periods 14-21 and 22-28 days p.e. Throughout the entire 35-day period exposed non-parasitized snails laid significantly more eggs than control snails, while parasitized snails laid significantly more eggs than controls during days 22-28 p.e. and significantly fewer during days 29-35 p.e. Parasitized snails also laid significantly more eggs/egg mass in the period 16-28 days p.e. than did control snails. Growth of the snails was measured. By day 28 p.e. the mean diameter of the exposed group was significantly greater than that of the control group. The increase in oviposition by snails soon after exposure is discussed in terms of a compensatory response for expected future suppression of egg-laying. The fact that parasitized and non-parasitized snails both show increased oviposition indicates that normal development of the parasite is not necessary to trigger the response.

Animals↗

Parasite (Schistosoma mansoni) and host (Biomphalaria glabrata) genetic diversity: population structure in a fragmented landscape.

Random amplified polymorphic DNA (RAPD) markers were used to quantify genetic diversity within and between 5 populations of Schistosoma mansoni within its definitive host (Rattus rattus) and the 5 corresponding populations of the snail intermediate host (Biomphalaria glabrata) from a limited endemic area of murine schistosomiasis on the island of Guadeloupe. Analysis of molecular variance (AMOVA) and canonical correspondence analysis (CCA) were used to test the significance of genetic differentiation between populations. Both methods gave similar results. Of total gene diversity, 15.1% (AMOVA) and 18.8% (CCA) was partitioned between localities for S. mansoni with an absence of association between genetic and geographical distances. Geographical localities accounted for 20.5% (CCA) of the total diversity for B. glabrata populations. The genetic distances between pairs of parasite populations were not correlated with the genetic distances between the corresponding pairs of snail host populations. Such strong patterns of local differentiation of both parasite and snail populations are consistent with predictions based on metapopulation dynamics and may have implications on host-parasite susceptibility relationship through local adaptation processes.

Animals↗

Refractoriness of host haemocytes to parasite immunosuppressive factors as a putative resistance mechanism in the Biomphalaria glabrata-Echinostoma caproni system.

In contrast to the growing knowledge accumulated on plant resistance to pathogens, mechanisms of parasite resistance largely remain to be elucidated in animal species. In the present study we investigated mechanisms underlying resistance/susceptibility in the snail-trematode system Biomphalaria glabrata-Echinostoma caproni. In particular, we compared the effect of the parasite excretory-secretory (E-S) products on the defence functions of haemocytes from 2 susceptible and 2 resistant snail strains. In vitro experiments showed that E. caproni E-S products inhibit adhesion and phagocytosis of haemocytes from susceptible snails. A partial biochemical characterization also suggested that the interfering factor(s) is (are) heat-labile glycosylated polypeptides of molecular mass between 10 and 30 kDa. Interestingly, haemocytes from resistant snails remained unaffected by the parasite E-S products, suggesting that a constitutive difference results in their refractoriness to the parasite's immunosuppressive factor(s).

Animals↗

The relationship between Schistosoma mansoni and Biomphalaria glabrata: genetic and molecular approaches.

Biomphalaria glabrata is a major intermediate host for the helminth parasite Schistosoma mansoni. Beginning in the mid-20th century, studies were carried out with this snail species to identify the immunological and genetic components that might be involved in controlling schistosome development. A number of genetically well-defined snail stocks were derived as a direct result of these studies and have since played major roles in helping investigators to identify important cellular and humoral components in the snail/schistosome relationship. This review will explore the historical development of these stocks and describe some of the major advances in several areas of medical malacology that hawe been made possible be their use.

Animals↗

Molecular approaches in the study of Biomphalaria glabrata--Schistosoma mansoni interactions: linkage analysis and gene expression profiling.

Gene mapping and the generation of linkage groups are fundamental to an understanding of the organization and relationships of genes and marker sequences, providing a framework with which to investigate their association with traits of interest. The abundance of techniques available for generating polymorphic molecular markers, and recent advances in high throughput screening, have allowed the extension of map analysis to the tropical freshwater snail Biomphalaria glabrata, an important intermediate host for Schistosoma mansoni. Direct comparison of gene expression by differential display screening, without prior identification of candidate genes, can be combined with mapping to quantify the involvement of specific sequences in the schistosome resistance response, and other important host-parasite interactions. Here we discuss the application of current and emergent technologies to gene characterization and linkage analysis in snail-schistosome interactions. Preliminary results from the analysis of comparative gene expression in resistant and susceptible snails are also presented.

Animals↗

Schistosoma mansoni and Echinostoma caproni excretory-secretory products differentially affect gene expression in Biomphalaria glabrata embryonic cells.

Biomphalaria glabrata embryonic (Bge) cells have been shown to be a valuable in vitro cellular model for the study of snail host-parasite interactions. They both promote the growth and differentiation of various trematode species including Schistosoma mansoni, and Echinostoma caproni and share some morphological and functional features with circulating haemocytes. As an approach to investigate snail genes potentially regulated following exposure to trematode excretory-secretory (ES) products, we compared gene expression profiles of Bge cells exposed to saline solution, or saline solution containing ES products from S. mansoni or E. caproni, two trematode species parasitizing B. glabrata. Following differential display RT-PCR analysis we characterized 23 differentially displayed cDNAs and we focussed on the 5 cDNAs showing sequence similarity to known genes for expression validation. Using RT-PCR, we confirmed that ES products from S. mansoni and E. caproni differentially affect the expression levels of 4 out of the 5 transcripts. These partial transcripts corresponded to novel B. glabrata sequences, and showed significant sequence similarity to genes coding for (i) cytochrome C, (ii) methyl-binding proteins, (iii) glutamine synthetases, and (iv) protease inhibitors from the Kunitz family. The possible significance of these gene expression changes in host-parasite molecular interactions is discussed.

Amino Acid Sequence↗

Compatibility in the Biomphalaria glabrata/Echinostoma caproni model: new candidate genes evidenced by a suppressive subtractive hybridization approach.

In order to elucidate mechanisms underlying snail/echinostome compatibility, numerous molecular studies comparing transcripts and proteins of Biomphalaria glabrata susceptible or resistant to Echinostoma caproni were undertaken. These studies focused on plasma and haemocytes of the two strains and revealed that some transcripts and/or proteins were differentially expressed between strains. The aim of the present study was to develop a complementary transcriptomic approach by constructing subtractive libraries. This work revealed some candidate transcripts already identified in previous studies (calcium-binding proteins and glycolytic enzymes) as well as novel candidate transcripts that were differentially represented between strains of B. glabrata. Among these newly identified genes, we revealed several genes potentially involved in immune processes encoding proteases, protease inhibitors, a lectin, an aplysianin-like molecule, and cell adhesion molecules.

Animals↗

Failure of Schistosoma mansoni to reinfect Biomphalaria glabrata snails: acquired humoral resistance or intra-specific larval antagonism?

Failure of snail reinfection by Schistosoma mansoni has been demonstrated in susceptible Biomphalaria glabrata infected with 1 miracidium and subsequently re-exposed to 1 or 5 homologous parasite larvae. The acquisition of 'resistance' to secondary parasite infection was time dependent since complete inhibition was observed at 2 weeks and longer following monomiracidial exposure. This phenomenon was still observed in snails challenged 8 weeks after primary infection. Histological observations revealed that sporocysts from the challenge infection were free of encapsulation, their development was stopped and they degenerated slowly in the absence of haemocytic reaction of the host. Under the hypothesis of an acquired homologous resistance mechanism, this strongly suggests that 1 or several unidentified humoral factors are responsible for the non-development of the sporocysts from the challenge infection. However, considering the time-dependent nature of the phenomenon, an intraspecific larval antagonism process between sporocysts resulting from the primary infection and those from the challenge infection may be involved.

Animals↗

Compatibility of Schistosoma mansoni and Biomphalaria pfeifferi in northern Senegal.

The construction of the Diama dam on the Senegal River and the ensuing ecological changes have led to a massive outbreak of Schistosoma mansoni infection in Northern Senegal, associated with very high intensity of infections, due to extremely intense transmission. The vectorial capacity of Biomphalaria pfeifferi from Ndombo, near Richard-Toll was investigated in order to assess the role of the snail-parasite relationship in this particular epidemiological situation. The results revealed an unusually high compatibility between the Senegalese S. mansoni strain and its local snail intermediate host, B. pfeifferi. The snail infection rate after exposure to a single miracidium per snail was 87%. The cercarial production of infected snails was very high, with a mean total production of 50,456 cercariae per snail. No significant difference was found in the total cercarial output between snails exposed to 1 miracidium and those exposed to 5 miracidia. The increase in the rate of cercarial output was significantly greater in snails exposed to 5 miracidia, but there was a higher mortality in this group. The chronobiological cercarial production pattern showed a peak around mid-day. The implications of these findings on the epidemiology of schistosomiasis in Northern Senegal are discussed.

Animals↗

Glycosidase activities in plasma of naive and schistosome-infected Biomphalaria glabrata (Gastropoda).

Activity of the following glycosidases was detected in the plasma of the freshwater snail Biomphalaria glabrata: beta-D-fucosidase, beta-D-glucosidase, beta-D-galactosidase, beta-D-mannosidase, beta-D-glucuronidase, N-acetyl-beta-D-galactosaminidase, N-acetyl-beta-D-glucosaminidase, and lysozyme. At the physiological pH (7.2-7.4) of snail haemolymph, enzymatic activity was about 10-50% of the maximum activity at each enzyme's respective acid pH-optimum. Schistosome-susceptible B. glabrata showed lower plasma protein concentration and significantly lower enzymatic activities (U/mg protein) than schistosome-resistant snails. Changes in glycosidase activity levels correlate with the progress of infection. After successful schistosome invasion, activities of plasma glycosidases but not the concentration of total plasma proteins increased significantly during the first 2 days in both snail strains. Thus, most tegumental glycoproteins of schistosome larvae can be altered by humoral host glycosidases. The detection of only very low activities of hexosaminidases leads to the hypothesis that GalNAc/GlcNAc may be involved in the process of non-self recognition. At 4 days post-infection, glycosidase activities were identical or slightly below the levels found in naive snails. At this time of infection the parasite is encapsulated and destroyed by haemocytes of resistant snails. In susceptible snails, however, the schistosomes have transformed into sporocysts and will complete their life-cycle without eliciting effective defence reactions. After > 30 days post-infection, when cercariae are fully developed in susceptible snails, plasma protein concentration decreased significantly, whereas glycosidase activities were elevated.

Animals↗

Genetic differentiation, dispersal and mating system in the schistosome-transmitting freshwater snail Biomphalaria glabrata.

Biomphalaria glabrata is the main intermediate host of Schistosoma mansoni in America and one of the most intensely studied species of freshwater snail, yet very little is known about its population biology. Here, we used seven highly polymorphic microsatellite loci to analyse genetic diversity in populations from three regions (Lesser Antilles, Venezuela and southern Brazil). Considerable genetic variation was detected, with an average (s.d.) H(0) = 0.32 (0.24). More diversity per population was found in the Valencia lake basin in Central Venezuela, which suggests an influence of dispersal (via inter-population connectivity) on the restoring of genetic diversity after the demographic bottlenecks recurrently experienced by populations. A marked population structure was detected and there seems to be a relationship between mean differentiation and genetic diversity within regions. There is also a significant isolation-by-distance pattern. The Lesser Antilles populations appear clearly differentiated from the rest, which suggests a single colonisation event followed by local radiation within these islands or multiple colonisation events from the same source area. Our results indicate that B. glabrata essentially cross-fertilises, with little variation in selfing rates among populations. However, significant deficits in heterozygotes and linkage disequilibria were detected in two Venezuelan populations suggesting a mixture of at least two different genetic entities, probably with differences in their respective mating systems.

Animals↗

Evolutionary history and phylogeography of the schistosome-vector freshwater snail Biomphalaria glabrata based on nuclear and mitochondrial DNA sequences.

The phylogeography of the freshwater snail Biomphalaria glabrata remains poorly known, although this species is the major vector of schistosomiasis in the New World. It was here investigated in South America and the Lesser Antilles, based on partial mitochondrial large ribosomal subunit (16S rDNA) and nuclear internal transcribed spacer-2 (ITS-2) gene sequences. Sampling included 17 populations from a large part of the current geographic range of the species (Brazil, Venezuela and Lesser Antilles). Substantial variability was detected, as well as a high amount of phylogenetically informative signal. The molecular phylogeny inferred splits B. glabrata into Northern and Southern clades separated by the Amazon river, and may even suggest a supra-specific status for B. glabrata. Brazilian populations were the most diverse and appeared basal to the other populations. Venezuelan haplotypes formed a single clade, albeit not strongly supported. Two Venezuelan haplotypes appear rather similar to Brazilian haplotypes. Similarly, Lesser Antilles haplotypes clustered in the same monophyletic clade, which suggests that the recent colonisation of the Antilles has a northern South American origin. However, the estimated divergence time between Antilles and Venezuelan sequences is extremely large (conservatively higher than 10(5) years). These results are discussed in the light of (i) phylogeographic patterns at South American scale, and (ii) recurrent introduction of molluscs, especially in the Antilles, as a consequence of human activities.

Animals↗

Purification and characterization of a tetrameric alpha-macroglobulin proteinase inhibitor from the gastropod mollusc Biomphalaria glabrata.

The alpha-macroglobulin proteinase inhibitors (alpha Ms) are a family of proteins with the unique ability to inhibit a broad spectrum of proteinases. Whereas monomeric, dimeric and tetrameric alpha Ms have been identified in vertebrates, all invertebrate alpha Ms characterized so far have been dimeric. This paper reports the isolation and characterization of a tetrameric alpha M from the tropical planorbid snail Biomphalaria glabrata. The sequence of 18 amino acids at the N-terminus indicates homology with other alpha Ms. The subunit mass of approx. 200 kDa was determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and SDS/PAGE. The quaternary structure was determined by sedimentation equilibrium centrifugation and native pore-limit electrophoresis. Evidence for a thioester is provided by the fact that methylamine treatment prevents the autolytic cleavage of the snail alpha M subunit and results in the release of 4 mol of thiols per mol of snail alpha M. The snail alpha M inhibited the serine proteinase trypsin, the cysteine proteinase bromelain and the metalloproteinase thermolysin. The spectrum of proteinases inhibited, together with the demonstration of steric protection of the proteinase active site and a "slow to fast' conformational change after reacting with trypsin, all suggest that the inhibitory mechanism of the snail alpha M is similar to the "trap mechanism' of human alpha 2-macroglobulin.

Amino Acid Sequence↗

Fine-scale population structure and dispersal in Biomphalaria glabrata, the intermediate snail host of Schistosoma mansoni, in Venezuela.

Biomphalaria glabrata is the main intermediate host of Schistosoma mansoni in America and one of the most intensely studied species of freshwater snails, yet very little is known about its population biology. Here, we used seven highly polymorphic microsatellite loci to analyse genetic diversity in the Valencia lake basin, which represents the core of the endemic area for schistosomiasis in Venezuela. Populations were sampled at short spatial scale (a few kilometres), both inside the lake and in ponds or rivers near the lake. Our results indicate that B. glabrata essentially cross-fertilizes, with little variation in selfing rates among populations. Our markers detected considerable genetic variation, with an average heterozygosity of 0.60. More diversity per population was found within than outside the lake, suggesting an influence of connectivity among populations on the levels of genetic diversity. A marked population structure was detected and lake populations were less structured than other populations. Most individuals were assigned to their population of origin using an assignment test. No strong demographic signal (e.g. bottleneck) was detected, though lake populations are likely to experience bottlenecks more frequently than the other populations analysed. Differences in gene flow therefore seem to play an important role in population differentiation and in the restoring of genetic diversity in demographically unstable populations.

Animals↗