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PubMed1994

cDNA cloning and deduced amino acid sequence of two ferritins: soma ferritin and yolk ferritin, from the snail Lymnaea stagnalis L.

Pulmonate freshwater snails contain two different ferritin types, soma ferritin and yolk ferritin. A cDNA library was constructed from midgut gland poly(A)-rich RNA of the snail Lymnaea stagnalis L. and recombinant clones encoding both ferritin types were obtained by immunoscreening. The longest cDNA inserts had a length of 859 bp (soma ferritin) and 1548 bp (yolk ferritin) and the specificity of these inserts was confirmed by immunoprecipitation of both ferritin types translated in vitro from hybrid-selected mRNAs. The 5' untranslated region (UTR) of the soma ferritin mRNA contains a 28-bp element which shows 64% sequence identity with the iron-responsive element (IRE) of vertebrate ferritin mRNAs. The soma ferritin mRNA is strongly translated in the wheat germ system but poorly translated in rabbit reticulocyte lysate. The yolk ferritin mRNA, which contains no IRE, is equally well translated in both in vitro translation systems. The deduced amino acid sequence of the soma ferritin subunit (174 amino acid residues, M(r) 20140) shows 50-70% sequence identity with subunits of vertebrate ferritins. After removal of an 18-amino-acid-residue signal sequence the deduced protein sequence of yolk ferritin contains 221 amino acids (M(r) 25438). Sequence identity of this chain with other eukaryotic ferritin chains is only 31-42%. Both snail ferritin sequences are more similar to the H-subunit type of vertebrate ferritins than to the L-type and both have the H-specific amino acid residues of the ferroxidase centre. The yolk ferritin sequence has a 42-amino-acid-residue insertion predicted to reside in the L loop of the subunit.

Amino Acid Sequence↗

PubMed1995

In the biosynthesis of N-glycans in connective tissue of the snail Lymnaea stagnalis of incorporation GlcNAc by beta 2GlcNAc-transferase I is an essential prerequisite for the action of beta 2GlcNAc-transferase II and beta 2Xyl-transferase.

Using a series of relevant substrates, connective tissue of the snail Lymnaea stagnalis was shown to contain beta 1-2 xylosyltransferase (beta 2Xyl-T), beta 1-2 N-acetylglucosaminyltransferase I (beta 2GlcNAc-T I), and beta 1-2 N-acetylglucosaminyltransferase II (beta 2GlcNAc-T II) activities. These enzymes are probably involved in the biosynthesis of the N-linked carbohydrate chains, like those present in hemocyanin. The products formed by incubation of GlcNAc beta 1-2Man alpha 1-6(GlcNAc beta 1-2Man alpha 1-3)Man beta 1-R [where R = -4GlcNAc beta 1-4GlcNAc or O-(CH2)7CH3] with UDP-Xyl and connective tissue microsomes have been purified and characterized by 1H-NMR spectroscopy in conjunction with methylation analysis to be GlcNAc beta 1-2Man alpha 1-6(GlcNAc beta 1-2Man alpha 1-3)(Xyl beta 1-2)Man beta 1-R. Substrate specificity studies focused on connective tissue beta 2Xyl-T show that the minimal structure requirements are fulfilled in GlcNAc beta 1-2Man alpha 1-3Man beta 1-O-(CH2)7CH3. The enzyme activity can therefore be characterized as UDP-Xyl:Glc-NAc beta 1-2Man alpha 1-3Man beta-R (Xyl to Man beta) beta 1-2 xylosyltransferase. In substrate-specificity studies directed to connective tissue beta 2GlcNAc-T I, it could be demonstrated that the enzyme is active towards acceptors having at the minimum a Man alpha 1-3Man beta-R sequence, and that introduction of a beta Xyl residue at C2 of beta Man totally abolishes the enzyme activity. Xylose-containing oligosaccharides are not acceptors for beta 2GlcNAc-T I. In combination with the substrate specificity of beta Xyl-T, this shows that in snail connective tissue beta 2GlcNAc-T I must act before beta 2Xyl-T. The connective tissue beta 2GlcNAc-T II activity follows the earlier established biosynthetic routes. Based on the substrate specificities of the various connective tissue glycosyltransferases known so far, and the structures isolated from L. stagnalis hemocyanin, a partial biosynthetic scheme for N-glycosylation in snail connective tissue is proposed.

Animals↗

PubMed1983

Chemosterilization and its reversal in the snail Lymnaea acuminata.

Treatment of the snail Lymnaea acuminata, the vector for the liver flukes Fasciola hepatica and Fasciola gigantica, with two alkylating agents (cyclophosphamide and busulfan) and one thiocarbamide (thiourea) caused reduction in the number of eggs and production of non-viable embryos. All the three drugs caused reduction in the levels of DNA, RNA and protein and the activity of alkaline phosphatase in the gonadal tissues of this snail. There was enhancement in free amino acid levels and activity of acid phosphatase by these drugs. Discontinuation of treatment could not reverse the cytotoxic effects of these drugs. Treatment of sterilized snails with prostaglandin E1 (PGE1), prostaglandin F2 alpha (PGF2 alpha) or hydrogen peroxide not only restored the number of eggs but also caused full embryonic development. Also, there was near total recovery of RNA, DNA and protein levels and small changes in free amino acid levels as well as the activity of alkaline and acid phosphatase. It has been suggested that the prostaglandins play a modulatory role in protein synthesis. The possible therapeutic use of PGE1, PGF2 alpha and H2O2 has been pointed out.

Alprostadil↗

PubMed1978

Schistosoma mansoni: identification of chemicals that attract or trap its snail vector, Biomphalaria glabrata.

A new bioassay for chemical attractants of aquatic snails demonstrated that Biomphalaria glabrata could be attracted to or trapped in the vicinity of homogenates of lettuce. Fractionation of homogenates revealed the amino acids glutamate and proline and the primary attractants. Attraction was specific for the L form of glutamate. Proline appeared to stimulate reproductive activity. Glutathione, gamma-aminobutyric acid, and a number of other compounds had no effect. Extracts of lyophilized snail tissue also attracted other snails and may thus contain pheromones. These results permit formulation and testing of controlled-release attractants designed to overcome the repellant effects of slow-release molluscicides, as well as the design of stimulants to be used with no-release poisons.

Amino Acids↗

PubMed1990

Identification in Xenopus of a structural homologue of the Drosophila gene snail.

We have cloned a Xenopus cDNA that is related to snail, a gene that is required for mesoderm formation in Drosophila. The cDNA encodes a protein that contains five zinc-fingers that closely resemble those of snail. In the non-canonical parts of the DNA-binding loop, there is almost 90% homology between snail and xsna. The corresponding mRNA (xsna) is expressed strongly at the start of zygotic transcription simultaneously with the transcription factor EF1 alpha. In early gastrulae, xsna is equally distributed between the dorsal and ventral halves of the equatorial zone. The possibility that the capacity to synthesise xsna is more localised before the start of zygotic transcription has been investigated by culturing fragments of stage 8 embryos until xsna is synthesised. The capacity to synthesise xsna at stage 8 is located principally in the dorsal half of the equatorial zone. A small amount of maternal xsna is localised in the vegetal hemisphere before zygotic transcription starts. xsna is not present in isolated animal caps but can be induced by the mesoderm-inducing factors XTC-MIF and bFGF. Synthesis of xsna does not occur autonomously in dispersed cells but is restored when cells reaggregate in the presence of calcium and magnesium.

Animals↗

PubMed1993

The expression of a zebrafish gene homologous to Drosophila snail suggests a conserved function in invertebrate and vertebrate gastrulation.

Snail, a zinc finger protein, is required for the formation of the ventral furrow and the mesoderm during gastrulation of the Drosophila embryo. snail homologues have been cloned from Xenopus and mouse. We have isolated a zebrafish homologue of snail, designated sna-1. Like its Drosophila counterpart, Sna-1 protein is nuclear. Maternal and zygotic sna-1 transcripts are ubiquitously distributed in zebrafish embryos of cleavage and blastula stages. In gastrulating embryos, sna-1 is expressed in involuting cells of the germ ring, but not in those at the dorsal midline, the presumptive notochordal region. After involution, the expression is maintained in the paraxial mesoderm and becomes prominent in the muscle pioneer precursors, followed by expression at the posterior somite boundaries. Later, sna-1 is expressed in neural crest and mesodermal derivatives of the head region. Sna-1 expression is induced in animal cap cells by activin A. The early sna-1 expression pattern in gastrulating zebrafish no tail (ntl) mutant embryos is normal except a reduction in the level of sna-1 transcription, suggesting that Ntl protein is not the key activator of sna-1 transcription in vivo, but might be involved in the enhancement or maintenance of sna-1 transcription. Data obtained in studies with ectopic ntl expression support this model.

Amino Acid Sequence↗

PubMed1999

Effects of anoxia on the extra- and intracellular acid-base status in the land snail helix lucorum (L.): lack of evidence for a relationship between pyruvate kinase down-regulation and acid-base status

The aims of the present study were to describe a possible correlation between the regulation of the key glycolytic enzyme pyruvate kinase and the acid-base status in the haemolymph and in several other tissues of land snails during anoxia. To illustrate whether such a relationship exists, we determined (i) the acid-base variables in the haemolymph and tissues of the land snail Helix lucorum, (ii) the kinetic properties of pyruvate kinase from several tissues and (iii) the levels of the anaerobic end-products d-lactate and succinate in the haemolymph and tissues of aerobic and anoxic Helix lucorum. The results showed that the pH of haemolymph (pHe) decreased significantly over the first 20 h of anoxia and then recovered slowly towards control values. A similar pattern was observed for intracellular pH (pHi), which decreased significantly over the first 16 h of anoxia and slowly returned towards control levels. The reduction and recovery of pHi and pHe seem to reflect the rate of anaerobic metabolism. The main anaerobic end-products, d-lactate and succinate, accumulated rapidly during the initial stages of anoxia and more slowly as anoxia progressed. The decrease in the rate of accumulation of anaerobic end-products during prolonged anoxia was due to the conversion of tissue pyruvate kinase to a less active form. The results demonstrate a correlation between pyruvate kinase down-regulation and the recovery of acid-base status in the haemolymph and the tissues of land snails during anoxia.

Journal Article↗

PubMedDate not supplied

Biological and morphological characteristics of Schistosoma mansoni from Ribeira Valley, state of São Paulo, Brazil I--Susceptibility of Biomphalaria tenagophila snail to sympatric S. mansoni strain.

In the São Paulo State, Brazil, where the Biomphalaria tenagophila is the intermediate host, the Ribeira Valley is an important endemic schistosomiasis mansoni area. During last eleven years there has been intense control measures focusing on schistosomiasis. The efforts have been concentrated in the municipalities of Pedro de Toledo and Itariri. We determined the susceptibility of B. tenagophila to sympatric strain of S. mansoni, both recently isolated from Itariri field. In 1988, this strain was isolated and maintained in the experimental model: Swiss mice--sympatric B. tenagophila. The second generation of the worm was evaluated. The snail were divided in the three groups of 60 snails each. One group was exposed to 1 miracidium and other to 10. The third group was the control. The mortality and the shedding of cercariae were checked during 78 days. After that, the positive snails were observed until they ceased to shed cercariae. The exposed molluscs showed mortality rates of 23% and 31% and infection indexes were of 8% and 60% to 1 and 10 miracidia respectively. The mortality was of 22% in the control group. The periods of shedding cercariae in the two groups were 82 and 104 days. We can conclude that B. tenagophila is an effective intermediate host to the sympatric strain of S. mansoni sympatric strain.

Animals↗

PubMed1992

Ecological studies on the intermediate host snails and the relevance to schistosomiasis control.

A detailed knowledge of distribution patterns of schistosome intermediate hosts and their population dynamics and factors affecting these patterns will provide useful information about the possibilities and desirability of conducting snail control measures in various transmission situations. On the basis of various case studies the association between the occurrence of human water contacts and the presence of schistosome intermediate hosts or infections in the intermediate hosts is illustrated. Other parameters affecting snail distribution patterns and density fluctuations are discussed. It is concluded that ecological studies on the intermediate hosts are extremely relevant, either to optimally apply existing control measures or to develop alternative measures of snail control, such as ecological or biological control.

Animals↗

PubMedDate not supplied

Effects of Bayluscide WP 70 on the survival and water-leaving behaviour of Biomphalaria straminea, snail host of schistosomiasis in northeast Brazil.

The toxic and behavioural effects of niclosamide (Bayluscide WP 70) on Biomphalaria straminea from a highly endemic area of schistosomiasis in northeastern Brazil were investigated through laboratory bioassays. The LD50 and LD90 were 0.114 mg/l and 0.212 mg/l, respectively. Water-leaving behaviour occurred among 14% to 30% of the snails in the presence of sublethal doses of niclosamide and among 16% of the controls. It was concluded that both the relatively low susceptibility to niclosamide and water-leaving behaviour of local B. straminea may be responsible for the recolonization of transmission foci after mollusciciding. It was suggested that recently improved measures of snail control, such as controlled-release formulations of niclosamide and plant molluscicides should be considered in areas where snail control is recommended.

Animals↗

PubMed2002

Identification of snails within the Bulinus africanus group from East Africa by multiplex SNaPshot trade mark analysis of single nucleotide polymorphisms within the cytochrome oxidase subunit I.

Identification of populations of Bulinus nasutus and B. globosus from East Africa is unreliable using characters of the shell. In this paper, a molecular method of identification is presented for each species based on DNA sequence variation within the mitochondrial cytochrome oxidase subunit I (COI) as detected by a novel multiplexed SNaPshotTM assay. In total, snails from 7 localities from coastal Kenya were typed using this assay and variation within shell morphology was compared to reference material from Zanzibar. Four locations were found to contain B. nasutus and 2 locations were found to contain B. globosus. A mixed population containing both B. nasutus and B. globosus was found at Kinango. Morphometric variation between samples was considerable and UPGMA cluster analysis failed to differentiate species. The multiplex SNaPshotTM assay is an important development for more precise methods of identification of B. africanus group snails. The assay could be further broadened for identification of other snail intermediate host species.

Animals↗

PubMed2005

Isolation and characterization of the full-length cDNA encoding a member of a novel cytochrome p450 family (CYP320A1) from the tropical freshwater snail, Biomphalaria glabrata, intermediate host for Schistosoma mansoni.

Cytochrome p450s (cyp450s) are a family of structurally related proteins, with diverse functions, including steroid synthesis and breakdown of toxins. This paper reports the full-length sequence of a novel cyp450 gene, the first to be isolated from the tropical freshwater snail Biomphalaria glabrata, an important intermediate host of Schistosoma mansoni. The nucleotide sequence is 2291 bp with a predicted amino acid sequence of 584aa. The sequence demonstrates conserved cyp450 structural motifs, but is sufficiently different from previously reported cyp450 sequences to be given a new classification, CYP320A1. Initially identified as down-regulated in partially resistant snails in response to S. mansoni infection, amplification of this gene using RT-PCR in both totally resistant or susceptible snail lines when exposed to infection, and all tissues examined, suggests ubiquitous expression. Characterization of the first cyp450 from B. glabrata is significant in understanding the evolution of these metabolically important proteins.

Amino Acid Sequence↗

PubMed2001

Effect of miracidial dose on adoptively transferred resistance to Schistosoma mansoni in the snail intermediate host, Biomphalaria glabrata.

Adoptively transferred resistance to Schistosoma mansoni in the snail intermediate host Biomphalaria glabrata was measured as a function of miracidial challenge dose. Schistosome-susceptible snails implanted with the amebocyte-producing organ (APO) from resistant donors showed 29 and 39% prevalences of infection after challenge with 5 and 10 miracidia, respectively, but 68-83% prevalences when exposed to 25-200 miracidia. Prevalences in control (untampered) susceptible snails ranged from 97 to 100% at the different miracidial doses. Higher infection prevalences at elevated doses suggest that a range of transferred resistance occurs and possibly that low levels of APO-derived plasma factors or hemocytes in some recipients can be overwhelmed by larger numbers of parasites.

Adoptive Transfer↗

PubMed2004

Mitotic responses to extracts of miracidia and cercariae of Schistosoma mansoni in the amebocyte-producing organ of the snail intermediate host Biomphalaria glabrata.

Suspensions of miracidia and cercariae of Schistosoma mansoni were subjected to repeated freeze-thaw cycles and then injected into resistant Salvador strain Biomphalaria glabrata snails. A pronounced increase in the number of mitotic figures, relative to uninjected, sham-injected, or diluent (water)-injected controls, was observed in the amebocyte-producing organ (APO) at 3 days postinjection (PI). After centrifugation of miracidia freeze-thaw extract (FTE), the resulting supernatant (FTS) and pellet possessed equal stimulatory activity that was approximately half that seen with FTE. Ultracentrifugation of miracidia FTS resulted in a supernatant that retained full activity, indicating a soluble molecule. Heat treatment of miracidia FTE reduced but did not eliminate activity, suggesting a nonprotein active component. Concentration or dilution of FTS by a factor of 10 gave a nonlinear dose-response relationship. Susceptible NIH albino snails injected with miracidia FTE had increased mitotic activity in the APO, which was much less than that seen in Salvador snails, whereas injection of miracidia FTE into Helisoma duryi had no discernable effect. Measurement of mitotic activity as a function of time PI showed no increase in numbers of mitotic figures in the APO at 18 hr but a large increase at 24 hr PI. Mitotic activity returned to preinjection levels by 96 hr PI, although a subsequent increase occurred at 120 hr PI.

Analysis of Variance↗

PubMed1989

Central and peripheral expression of genes coding for egg-laying inducing and insulin-related peptides in a snail.

Egg laying in the hermaphrodite freshwater snail Lymnaea stagnalis is a highly complex activity, including a series of internal activities (ovulation, egg and egg mass formation) which are closely correlated to a pattern of behaviours (alteration of locomotion and feeding, specific postures, oviposition). In this snail egg laying is induced by the neuroendocrine caudodorsal cells (CDCs), consisting of two homogeneous clusters at a total of 100 neurons. At egg laying these neurons release their products during a 60 min period of firing. The genes coding for these products have been cloned and characterized. There are two genes, CDCH-I and -II. Each gene codes for 12 peptides; one of these is the ovulation hormone (CDCH). The genes display over 90% homology. The most striking difference is a 17 bp deletion near the carboxy-terminal region. With immunocytochemistry and in situ hybridization both CDCH genes appeared to be expressed in the CDC and in paired groups of ectopic CDC-like neurons in the pleural ganglia, while a group of small neurons in the cerebral ganglia expresses the CDCH-I gene only. In addition, a widespread expression of the CDCH genes has been demonstrated in peripheral tissues. In the female part of the reproductive tract neurons were found to express the CDCH-I gene. In the male part of the tract exocrine secretory cells express the CDCH-I or -II gene. The gene products are secreted into the male tract and transferred to the female partner during copulation. Finally, sensory neurons in the head skin and mantle edge were found to express the CDCH-I gene. The presence of insulin-related peptides has been demonstrated in the brain as well as the digestive system of Lymnaea. The brain insulin-related peptides are produced in 4 groups of 50 giant neurons each (Light green cells, LGC). These neurons are involved in various physiological activities, related to body growth and glycogen metabolism. The major gene products expressed in the LGC have been cloned and characterized. It appeared that the predicted proteins represent three types of insulin-related molecules (MIP, molluscan insulin-related peptide). In these MIPs, those elements important in the determination of the tertiary structure, have been conserved. The MIP of the digestive system has been characterized up to now only at the peptide level. The snail gut MIP is more hydrophobic compared to bovine insulin. Cells containing MIP have been identified immunocytochemically in the gut epithelium.

Animals↗

PubMed1976

The chemical ecology of Biomphalaria glabrata: the effects of ammonia on the growth rate of juvenile snails.

When juvenile specimens of Biomphalaria glabrata were subjected to concentrations of ammonia ranging from 1-100 mug/ml in various media the following effects were observed: the addition of ammonia to borate buffered media caused mortality. Both borate and tris-buffered media caused a decrease in the growth rate of snails when compared with controls in SSW. The growth rates of the snails could be enhanced by increasing the concentration of ammonia to critical thresholds, but further increases beyond these thresholds resulted in growth inhibition. The toxicity of ammonia in ambient water was augmented by an an increase in pH. The possible causation and ecological significance of these effects are discussed. There are indications that the snails are physiologically well-adapted to utilize ammonia when required and also to control its excretion and uptake from the medium.

Ammonia↗

PubMed1988

Metacercariae of Echinostoma cinetorchis encysted in the fresh water snail, Hippeutis(Helicorbis) cantori, and their development in rats and mice.

The fresh water snail, Hippeutis(Helicorbis) cantori, caught from the rice paddies in Namyangju-gun, Gyeonggi-do, was found to be a new second intermediate host of E. cinetorchis in Korea. The metacercariae were found from 89(76.7 %) out of 116 snails examined and the average metacercarial burden was 4.7 per infected snails. Some developmental characteristics of E. cinetorchis were observed after experimental infection of the metacercariae to rats and mice.

Journal Article↗

PubMed1998

Transfection and heat-inducible expression of molluscan promoter-luciferase reporter gene constructs in the Biomphalaria glabrata embryonic snail cell line.

Studies were initiated to begin developing a genetic transformation system for cells derived from the freshwater gastropod, Biomphalaria glabrata, an intermediate host of the human blood fluke Schistosoma mansoni. Using a 70-kD heat-shock protein (HSP70) cDNA probe obtained from the B. glabrata embryonic (Bge) cell line, we cloned from Bge cells a complete HSP70 gene including a 1-kb genomic DNA fragment in its 5'-flanking region containing sequences indicative of a HSP promoter. Identified in the 5'-half (416 nucleotides) of this genomic fragment were TATA and CAAT boxes, two putative transcription initiation sites, and a series of palindromic DNA repeats with shared homology to the heat-shock element consensus sequence (Bge HSP70(0.5k) promoter). The 3'-half of this upstream flanking region was comprised of a 508-base intron located immediately 5' of the ATG start codon. To determine the functionality of the putative snail promoter sequence, Bge HSP promoter/luciferase (Luc) reporter gene constructs were introduced into Bge cells by N-(1-(2,3-dioleoyloxy) propyl)-N,N,N-trimethylammonium methylsulfate (DOTAP)-mediated transfection methods, and assayed for Luc activity 48 hr following a 1.5-hr heat-shock treatment (40 degrees C). Compared with control vectors or the Bge HSP70(0.5k/1.0k) promoter constructs at 26 degrees C, a 10- to 300-fold increase in Luc expression was obtained only in the Bge HSP70 promoter/Luc-transfected cells following heat-shock. Results of transfection experiments demonstrate that the Bge HSP70(0.5k) DNA segment contains appropriate promoter sequences for driving temperature-inducible gene expression in the Bge snail cell line. This report represents the first isolation and functional characterization of an inducible promoter from a freshwater gastropod mollusc. Successful transient expression of a foreign reporter gene in Bge cells using a homologous, inducible promoter sequence now paves the way for development of methods for stable integration and expression of snail genes of interest into the Bge cell line.

Animals↗