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The molluscicidal activity of the latex of Euphorbia splendens var. hislopii on Melanoides tuberculata (Thiaridae), a snail associated with habitats of Biomphalaria glabrata (Planorbidae).

The use of the latex of Euphorbia splendens var. hislopii was considered as an effective control method for Biomphalaria glabrata in Sumidouro, Rio de Janeiro. However, the appearance and expansion of the snail Melanoides tuberculata since August 1997, with the concomitant reduction of the population of B. glabrata suggest that competitive exclusion might be taking place. Depending on the susceptibility of the thiarid to the E. splendens toxin, the natural control that is occurring could be interrupted by the employment of the latex if the planorbid were less susceptible to the toxin. The aim of this study is to investigate the molluscicidal activity of the latex on M. tuberculata. We used 420 M. tuberculata, from Sumidouro. Fourteen different latex concentrations were tested using World Health Organization general methodology. Probit analysis was used for LD90 and LD50 determination. The LD50 was 3.57 mg/l and LD90 was 6.22 mg/l. At the highest concentration (10 mg/l) there was no survival. No significant differences among replicas (chi2 = 8.31; gl = 13; p > 0.05) were found. The LD90 dose for M. tuberculata was 13.8 times greater than that for B. glabrata, so that the molluscicide in the presence of the thiarid may have a synergic effect on reduction of Biomphalaria populations.

Animals↗

Geographical distribution of Biomphalaria snails in the State of Minas Gerais, Brazil.

Published and unpublished observations on geographical distribution of Biomphalaria snails in the State of Minas Gerais, Brazil, were compiled. This work is aimed at knowing the present occurrence of Biomphalaria species in this region, and at contributing to the elaboration of the planorbid chart of Minas Gerais. In malacological surveys, performed by several researchers, the presence of seven species of this genus was recorded. Those planorbids were found in 12 mesoregions, in 283 (33.1%) municipalities out of 853 with the following distribution: B. glabrata (185 municipalities), B. straminea (125), B. tenagophila (58), B. peregrina (57), B. schrammi (26), B. intermedia (20) and B. occidentalis (2). B. glabrata and B. tenagophila are found naturally infected by Schistosoma mansoni in Minas Gerais. In 24 municipalities the three snail hosts of S. mansoni in Brazil, B. glabrata, B. tenagophila and B. straminea, are present.

Animals↗

Anesthesia of Biomphalaria spp. (Mollusca, Gastropoda): sodium pentobarbital is the drug of choice.

The anesthetic effect of some water-soluble anesthesic or narcotic drugs currently used in mice was tested in molluscs of the Biomphalaria genus. Sodium thiopental was very toxic to the snails resulting in high rates of mortality in all the treatment schedules tested. Cetamine base, at concentration of 0.25 mg/ml of water, resulted in partial snail anesthesia (40% of snails were anesthetized) only after 20 h of exposition. The association of Cetamine base with Tiazine chloridrate did not improve the anesthesic effect, and higher concentrations of these drugs were toxic to the snails. Sodium pentobarbital at 0.4 mg/ml in water for 8 h was the best treatment schedule to anesthetize Biomphalaria snails. In this schedule, the snails were anesthetized without any toxic effect. The procedure provides a powerful tool for in vivo studies that demande a complete state of snail anesthesia.

Adjuvants, Anesthesia↗

Genetic markers between Biomphalaria glabrata snails susceptible and resistant to Schistosoma mansoni infection.

The analysis of the genetic variability related to susceptibility to Schistosoma mansoni infection in the vector of the genus Biomphalaria is important in terms of a better understanding of the epidemiology of schistosomiasis itself, the possible pathological implications of this interaction in vertebrate hosts, and the formulation of new strategies and approaches for disease control. In the present study, the genetic variability of B. glabrata strains found to be resistant or susceptible to S. mansoni infection was investigated using DNA amplification by random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR). The amplification products were analyzed on 8% polyacrylamide gel and stained with silver. We selected 10 primers, since they have previously been useful to detect polymorphism among B. glabrata and/or B. tenagophila. The results showed polymorphisms with 5 primers. Polymorphic bands observed only in the susceptible strain. The RAPD-PCR methodology represents an adequate approach for the analysis of genetic polymorphisms. The understanding of the genetic polymorphisms associated to resistance may contribute to the future identification of genomic sequences related to the resistance/susceptibility of Biomphalaria to the larval forms of S. mansoni and to the development of new strategies for the control of schistosomiasis.

Animals↗

Antigenic community between Schistosoma mansoni and Biomphalaria glabrata: on the search of candidate antigens for vaccines.

We have previously confirmed the presence of common antigens between Schistosoma mansoni and its vector, Biomphalaria glabrata. Cross-reactive antigens may be important as possible candidates for vaccine and diagnosis of schistosomiasis. Sera from outbred mice immunized with a soluble Biomphalaria glabrata antigen (SBgA) of non-infected B. glabrata snails recognized molecules of SBgA itself and S. mansoni AWA by Western blot. Recognition of several molecules of the SBgA were inhibited by pre-incubation with AWA (16, 30, 36, 60 and 155 kDa). The only specific molecule of AWA, inhibited by SBgA, was a 120 kDa protein. In order to determine which epitopes of SBgA were glycoproteins, the antigen was treated with sodium metaperiodate and compared with non-treated antigen. Molecules of 140, 60 and 24 kDa in the SBgA appear to be glycoproteins. Possible protective effects of the SBgA were evaluated immunizing outbred mice in two different experiments using Freund's Adjuvant. In the first one (12 mice/group), we obtained a significant level of protection (46%) in the total worm load, with a high variability in worm recovery. In the second experiment (22 mice/group), no significant protection was observed, neither in worm load nor in egg production per female. Our results suggest that SBgA constitutes a rich source of candidate antigens for diagnosis and prophylactic studies.

Animals↗

Analysis of the first and second internal transcribed spacer sequences of the ribosomal DNA in Biomphalaria tenagophila complex (Mollusca: Planorbidae).

The first and second internal transcribed spacer regions (ITS1 and ITS2) of the ribosomal DNA of Biomphalaria tenagophila complex (B. tenagophila, B. occidentalis, and B. t. guaibensis) were sequenced and compared. The alignment lengths of these regions were about 655 bp and 481 bp, respectively. Phylogenetic relationships among the Biomphalaria species were inferred by Maximum Parsimony and Neighbor-joining methods. The phylogenetic trees produced, in most of the cases, were in accordance with morphological systematics and other molecular data previously obtained by polymerase chain reaction and restriction fragment length polymorphism analysis. The present results provide support for the proposal that B. tenagophila represents a complex comprising B. tenagophila, B. occidentalis and B. t. guaibensis.

Animals↗

Primary culture of the region of the amebocyte-producing organ of the snail Biomphalaria glabrata, the intermediate host of Schistosoma mansoni.

Biomphalaria glabrata snails are major hosts for the digenetic trematoda Schistosoma mansoni, the causative agent of human schistosomiasis. The success or failure of the infection will be dependent on the mobilization of the molluskan internal defense system, where a major role will be played by circulating hemocytes produced by the APO (amebocyte-producing organ) of the snail. In this report, the primary culture of the APO region of B. glabrata was obtained for the first time, as well as a control culture of the ovotestis. Three different cell populations migrated easily from the explants in culture, with no need of any dispersion agent. The cells grew in suspension at an incubation temperature of 15 degrees C and the cultures were maintained viable for up to two weeks. Two of these cell populations obtained resembled cell types known to be present in the hemolymph of Biomphalaria. The availability of APO cells in culture may contribute to a better understanding of the internal defense in mollusks, in general, as well as the specific response of B. glabrata to S. mansoni infection.

Animals↗

Rescue of sporocysts of Schistosoma mansoni in nonsusceptible Biomphalaria by head-foot transplantation into susceptible snails.

To measure the longevity of sporocysts of Schistosoma mansoni in nonsusceptible snails (13-16-R1 and Salvador strains of Biomphalaria glabrata, and Biomphalaria obstructa), the head-foot (HF) of miracidia-exposed snails was transplanted into the hemocoel of a susceptible NIH albino recipient at 1-36 days postexposure (DPE). Recipient snails which were not exposed to miracidia then were monitored for infection transferred by the implant, and infection prevalences in recipients of HF transplants from nonsusceptible donors were compared to those in snails implanted with an HF from NIH albino donors. Transplants from NIH albino snails between 1 to 15 DPE infected 98% of recipients. Similarly, at 1 DPE, 69-85% of transplants from nonsusceptible snails contained viable sporocysts, as shown by resulting patent infections in the recipients. Recipient infection prevalence, and presumably numbers of transplants containing viable sporocysts, declined as a function of DPE, and by 5-9 DPE this decrease was significant for all 3 types of nonsusceptible donors. However, viable sporocysts still occurred in B. obstructa and 13-16-R1 B. glabrata as late as 19 and 20 DPE, respectively, and in Salvador B. glabrata as late as 33 DPE. Thus, sporocysts persist in nonsusceptible snails considerably longer than suggested by results of previous histological studies.

Animals↗

The mitochondrial genome of Biomphalaria glabrata (Gastropoda: Basommatophora), intermediate host of Schistosoma mansoni.

The complete mitochondrial (Mt) genome of the gastropod Biomphalaria glabrata, a major intermediate host for the human parasite Schistosoma mansoni, was sequenced. The circular genome, the first determined from a basommatophoran snail, is AT rich (74.6%) and the smallest Mt genome (13,670 nucleotides [nt]) characterized from mollusks to date. Sequences from 2 B. glabrata strains, M-line and 1742, differed by only 18 nt. Phylogenetic analysis of 16S and ND1 sequences confirmed the Brazilian ancestry of both B. glabrata strains. Gene predictions indicated 22 transfer RNA, 12S and 16S ribosomal RNA (rRNA), and 13 protein-encoding genes, as is typical for metazoans. Of the mollusk Mt genomes currently known, the gene order was most similar to that of stylommatophoran gastropods, concordant with the monophyly of pulmonate gastropods. Screening of GenBank (expressed sequence tags database [dbEST]) with the Mt sequence identified 108 entries from B. glabrata as Mt-derived sequences, including 12S and 16S rRNA sequences. Moreover, 11 sequences originating from the Mt genome of B. glabrata were identified among EST entries ascribed to intramolluskan stages of S. mansoni. The availability of this Mt sequence will facilitate further molecular investigations into the biology of Biomphalaria sp. and interactions between this intermediate host and intramolluskan stages of S. mansoni.

Animals↗

Effects of copper sulfate toxicity on cercariae and metacercariae of Echinostoma caproni and Echinostoma trivolvis and on the survival of Biomphalaria glabrata snails.

Copper in the form of copper sulfate (CuSO4) decreases the survival of Biomphalaria glabrata snails, but the effects of this molluscicide on Echinostoma caproni and Echinostoma trivolvis, 2 species of digeneans that use B. glabrata as intermediate hosts, are not known. Studies were done on the effects of various concentrations of CuSO4 in artificial spring water (ASW) on the survival and infectivity of E. caproni and E. trivolvis cercariae. Solutions containing 1.0, 0.1, and 0.01% CuSO4 were 100% lethal within 2 hr of exposure for both species. Time to 50% mortality in 0.001% CuSO4 was 8 hr for E. caproni and 16 hr for E. trivolvis; at 24 hr, the controls showed 50 and 65% mortality, respectively. Treatment of cercariae of both species for 0.5 hr in 0.001% CuSO4 had no effect on the ability of cercariae to form normal cysts in juvenile B. glabrata snails. However, treatment with 0.01% CuSO4 for 0.5 hr caused a significant reduction in the ability of cercariae of both species to encyst in snails. Treatment of encysted metacercariae of both species in 0.001% CuSO4 for I hr had no effect on subsequent excystation of these echinostomes in a trypsin-bile salts medium, whereas concentrations of 1.0, 0.1, and 0.01% CuSO4 and 1.0 and 0.1% CuSO4 decreased chemical excystation of E. caproni and E. trivolvis cysts, respectively. Survival studies on the effects of CuSO4 in Locke's solution on chemically excysted metacercariae of both species were also done. Excysted metacercariae of both species were killed by 2 hr in either 0.1 or 0.01% CuSO4 in Locke's solution. However, time to 50% mortality for both species of excysted metacercariae in 0.001% CuSO4 was approximately 5 hr. Time to 50% mortality for the controls was about 12 hr. Survival of juvenile B. glabrata snails was also examined. All B. glabrata snails were dead by 6 hr in 1 and 0.1% CuSO4 in ASW. Biomphalaria glabrata snails showed 50% mortality by about 6 hr in 0.01% CuSO4 and about 80% were still alive at 24 hr in 0.001% CuSO4. All controls were alive at 24 hr, at which time the experiment was terminated. Concentrations greater than 0.001% CuSO4 increased snail mortality, as well as that of the cercariae and excysted metacercariae of E. caproni and E. trivolvis. Our findings suggest that concentrations of copper sufficient to eliminate juvenile B. glabratta snails are also sufficient to kill the cercariae and excysted metacercariae of these digeneans but not the encysted metacercariae, which may be protected by their cyst walls.

Analysis of Variance↗

Multiplex PCR for both identification of Brazilian Biomphalaria species (Gastropoda: Planorbidae) and diagnosis of infection by Schistosoma mansoni (Trematoda: Schistosomatidae).

A simple and single-step technique based on multiplex PCR (multiplex polymerase chain reaction) has been developed for simultaneous identification of Brazilian Biomphalaria species, the intermediate hosts of Schistosoma mansoni, and their diagnosis of infection by the trematode. We used species-specific primers directed both to the internal transcribed spacer 2 (ITS2) of ribosomal DNA from 3 of the S. mansoni host species and to the mitochondrial DNA (mtDNA) from the trematode. Those primers were used simultaneously in a single multiplex-PCR reaction, and template DNA was obtained from S. mansoni-infected and noninfected snails. The results were visualized in silver stained polyacrylamide gels, revealing the presence of specific bands. The methodology has shown to be efficient, fast, and reproducible for Biomphalaria species identification and diagnosis of snails infected by S. mansoni during prepatent periods.

Animals↗

Glucose-6-phosphate dehydrogenase of Biomphalaria glabrata (Say , 1818) (Mollusca: Pulmonata). Characterization and inheritance pattern.

Glucose-6-phosphate dehydrogenase of Biomphalaria glabrata has been characterized by electrophoresis, kinetic properties and inheritance pattern. It exists as a single electrophoretic band with no polymorphism in the natural population studied during the present investigations. There are no organ-specific differences in electrophoretic pattern. The enzyme from the homogenates of various organs e.g., albumen gland, digestive gland, ovotestis and columellar muscle show similar physico-chemical parameters, such as optima for pH, NADP and glucose-6-phosphate. Incorporation of NADP is essential for maintaining the stability of the enzyme, failing which, there appear artifactual variations in electrophoretic mobility of the enzyme. The apparent lack of polymorphism in natural population of Biomphalaria glabrata has been discussed.

Animals↗

[Monthly variations in the level of infestation and the potential for transmission of Biomphalaria pfeifferi in 2 aquatic systems in Lwiro, Democratic Republic of Congo].

This study of variations in the rate of infestation by Biomphalaria pfeifferi and risk of schistosomiasis transmission was carried out to determine optimal timing for control measures. The study was conducted in two aquatic ecosystems, i.e. creek and fish pond, in the Lwiro region located in the eastern part (sud-Kivu) of the Democratic Republic of Congo. Biomphalaria pfeifferi infestation rates were high in the creek from November to January and consistently low in the fish pond. The risk of schistosomiasis transmission to man was highest in the creek especially during the months of January, February, May, and September. The risk of transmission was always less than 16% in the fish pond. Based on these findings, the optimal timing for control measures is the dry season for creeks and the rainy season for fish ponds.

Animals↗

Susceptibility of Biomphalaria alexandrina to infection with Schistosoma mansoni: correlation with the activity of certain glycolytic enzymes.

The importance of the glycolytic flux for the success of Biomphalaria-Schistosome sporocyst interaction was acertained in this study. Hexokinase (HK), pyruvate kinase(PK), glucose phosphate isomerase(GPI) and lactate dehydrogenase (LD) as four important glycolytic enzymes were markedly stimulated in trematode infected Biomphalaria alexandrina when measured two weeks post exposure to infection with Schistosoma mansoni miracidia. On the other hand phosphoenolpyruvate carboxy kinase (PEPCK), glucose-6-phosphatase (G6Pase) and fructose 1,6 diphosphatase(FDPase) as three gluconeogenic enzymes were slightly affected which confirm the importance of the glycolytic pathway for schistosome-exposed snails. Effect of LC25 of Solanum nigrum leaves dry powder as plant molluscicide on HK, PK and GPI were tested. Treatment with this plant resulted in a significant inhibition of these three investigated enzymes. LC10 concentrations of S. nigrum reduced considerably the infection rate of B. alexandrina with S. mansoni to be 34% compared to an infection rate of 80% in control, non-treated snails. Longer prepatent period and remarkable decrease in cercarial production was also recorded in snails treated with the sublethal concentrations of the molluscicide. As conclusion, susceptibility of B. alexandrina to infection with the digenetic trematode S. mansoni is correlated to the activity levels of the glycolytic enzymes. Moreover, sublethal and less pollutant concentration of S. nigrum could be recommended to control schistosomiasis by disturbing the intramolluscan environment of the parasite.

Animals↗

Sublethal concentration of Ambrosia maritima(Damsissa) affecting compatibility of Biomphalaria alexandrina snails to infection with Schistosoma mansoni through disturbing the glycolytic pathway.

High glycolytic flux as an emergency pathway for generating ATP was recorded as the most important metabolic pathway required for the success of Biomphalaria-Schistosome sporocyst interaction. Effect of LC25 of dry powdered Ambrosia maritima (Damsissa) as plant molluscicide on hexokinase (HK), pyruvate kinase(PK), glucose phosphate isomerase(GPI) was tested. It resulted in a significant inhibition of the three investigated enzymes. Treatment of snails with LC10 concentrations of A. maritima reduced considerably the infection rate of Biomphalaria alexandrina with Schistosoma mansoni to be 34% compared to an infection rate of 80% in control non-treated snails. Longer prepatent period and remarkable decrease in cercarial production was also recorded in snails treated with the sublethal concentrations of this molluscicide.

Animals↗

SOME FACTORS AFFECTING THE FECUNDITY OF BIOMPHALARIA PFEIFFERI (KRAUSS) IN GLASS AQUARIA.

Little being known of the effect of artificial conditions on Biomphalaria snails maintained in laboratory aquaria, experiments were conducted to determine some of the basic requirements of these snails, expressed in terms of the influence of various factors on their growth, fecundity and mortality. Among the factors studied were diet, artificial aeration, the chemical and physical properties of the water, and the presence of human urine in the water.The results obtained suggest that a diet of dehydrated lettuce and lucerne, no artificial aeration, a CaCO(3) concentration of approximately 18 p.p.m., a sodium/calcium ratio of 1, and mild "pollution" give optimum fecundity and growth and low mortality with Biomphalaria pfeifferi (Krauss).

Africa↗

Important aspects of Biomphalaria snail-schistosome interactions as targets for antischistosome drug.

The Biomphalaria species are freshwater snails which have a wide distribution and are significant both medically and economically as intermediate hosts for the schistosome parasite, a digenetic trematode causing schistosomiasis, a disease that infects 200 million people, and domestic animals throughout the tropics. The host-parasite relationship is, in principle, a powerful determinant of the biology of infection and disease. Research on snail-schistosome interactions has the potential for making an important contribution to the study of co-evolution or reciprocal adaptation. The association between Biomphalaria and Schistosoma mansoni could well be an excellent model for studies aimed at elucidating some aspects of the compatibility or resistance of this species to schistosomes. Snail hosts and schistosomes appear to have effects on each other's phenotype and genotype. The objective of this review is to clarify the nature of the relationship between schistosome parasites and their freshwater snail hosts. Aspects of snail-schistosome interactions will be traced in relation to behavioral (growth, reproduction, locomotion), immunological, and biochemical changes induced in the host's tissues by the developing intramolluscan stages of the parasite. This may help to identify biochemical or genetic targets for drug design. Manipulation of the intermediate host through these targets could break the cycle of human and snail infection by schistosomes.

Animals↗