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The influence of mating system, demography, parasites and colonization on the population structure of Biomphalaria pfeifferi in Madagascar.

Current evolutionary forces and historical processes interact to shape the distribution of neutral genetic variability within and among populations. Focusing on the genetics of recently introduced organisms offers a good opportunity to understand the relative importance of these factors. This study concerns variation at 8 polymorphic microsatellite loci in 30 populations of Biomphalaria pfeifferi. The sampling area spans most of the species' range in Madagascar where it was probably introduced recently. Extremely low variation was found within all populations studied, which may partly result from high selfing rates. However, this cannot account for the variance of variation across populations, which is better explained by habitat openness (that reflects environmental stochasticity), the prevalence of the parasitic trematode Schistosoma mansoni and historical demography (colonization and subsequent bottlenecks). Large global differentiation was also observed, suggesting that current gene flow among populations is limited to small distances, within watersheds and to few individuals. Our data set also allowed us to test several hypotheses regarding colonization, based on bottleneck and admixture tests. The observed pattern requires at least two independent introductions from slightly differentiated genetic sources in the western part of Madagascar. Another introduction, from a very different genetic origin, should also be postulated to explain the genetic composition of eastern populations. That this introduction occurred recently suggests that the colonization of Madagascar by B. pfeifferi is an ongoing process.

Animals↗

Anthraquinones in Rheum palmatum and Rumex dentatus (Polygonaceae), and phorbol esters in Jatropha curcas (Euphorbiaceae) with molluscicidal activity against the schistosome vector snails Oncomelania, Biomphalaria and Bulinus.

Hot water extracts of Rheum plamatum and Rheum dentatus (from China) showed molluscicidal activity against the snails Oncomelania hupensis, Biomphalaria glabrata and Bulinus globosus, which are vectors of Schistosoma japonicum, S. mansoni and S. haematobium, respectively. Activity was correlated with antraquinones which were identified by HPLC: rhein and chrysophanol-anthron were most active (> 50% dead snails after 2 days in a 0.03% solution). Molluscicidal activity was intermediate with Rheum-emodin and physcion and was not detectable with cinnamic acid or Aloe-emodin. The snail O. hupensis tended to be more sensitive for several compounds than B. glabrata. Extracts of Jatropha curcas seeds (from Mali) showed molluscicidal activity against both B. glabrata and O. hupensis, the latter being the more sensitive snail. The activity was associated with phorbol esters extracted from Jatropha oil. Of the pure phorbol esters tested, 4 beta-phorbol-13-decanoate killed both snail species at a concentration of 0.001% (10 p.p.m). As Jatropha is locally grown in Mali for other purposes, it might potentially be exploited for schistosomiasis control.

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[Influence of some medium factors on the fecundity of Biomphalaria glabrata (Say, 1818) Mollusca, Pulmonata) in laboratory conditions].

In different samples of fresh waters we studied the fecundity of Biomphalaria glabrata. We tested, too, the influence of different concentrations of Na, Ca, Mg, and K-salts. The snail, B. glabrata, is not affected by very high rates of those salts but eggs laying is getting down when the amount of NaCl is between 2000 and 4000 mg/1, of MgCl2 between 410 and 763 mg/1 and KCl between 330 and 760 mg/1. The rate in Ca seems inactive in the field of fecundity but, in an other hand, high concentrations of Ca suppressed the inhibiting effect of the high rates in Mg. B. glabrata is not affected by high rates in Mg, when Mg/Ca is Low. The inhibiting effect of Mg begins when Mg/Ca is around 30. The importance of different kinds of substratum for eggs lying on fecundity was also studied.

Animals↗

[Experimental study of the life cycle of Echinostoma togoensis n. sp., a larval parasite of Biomphalaria pfeifferi in Togo (author's transl)].

The adult and larval stages of Echinostoma togoensis are described. E. togoensis develops into rediae in the ovotestis of Biomphalaria pfeifferi which is totally sterilized. The metacercariae were recovered either in the pericardial cavity of several aquatic Pulmonata, or in the kidney of amphibian tadpoles. The adult was experimentally obtained in the laboratory mice exclusively.

Animals↗

[Effects of hyperinfestations by Echinostoma togoensis Jourdane and Kulo, 1981 on growth and survival of Biomphalaria pfeifferi snails].

A study of larval development of Echinostoma togoensis in Biomphalaria pfeifferi under hyperinfestation conditions (number of miradia equal to or greater than 10) showed the following characteristics: --Exposure to E. togoensis resulted in considerably increased growth in relation to the miracidial exposure dose; --Exposure to E. togoensis resulted in considerably increased mortality of B. pfeifferi. A population of Planorbs infested with 20 miracidia is completely eliminated 6 weeks after exposure. This last character, discovered for the first time in an Echinostome species, which is also a sterilizing species for the Planorb and a dominant competitor over Schistosoma mansoni, emphasizes the choice of E. togoensis as a biological control agent of intestinal schistosomiasis in Africa.

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[In vitro development of hemolymph granulocytes of Biomphalaria glabrata (Gastropoda, Planorbidae). Study by scanning electron microscopy].

The morphological evolution in vitro of Biomphalaria glabrata granulocytes on a cover-glass is described by scanning electron microscopy. 4 stages have been characterized: stage 1: globular cells showing low adhesion to cover-glass; stage 2: flattened cells in contact with substrate over a large surface, which present two well differentiated cytoplasmic zones: an endoplasm in which the nucleus is embedded and an ectoplasm which extends into very thin filopodia with a spherical terminal bulb; stage 3: cells which are very flattened and which adhere to each other; stage 4: cells which are completely spread on the cover-glass, in which the ectoplasm includes the filopodia. These 4 stages probably represent the granulocyte ontogenetic stages in their role in cellular defense against foreign substances.

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Observed disparity on schistosome infection rates in field Biomphalaria pfeifferi (Krauss) between two areas of the Jos Metropolis, Nigeria.

Two regions of the Jos Metropolis in Plateau State, Nigeria, with contrasting topographic features and harbouring many snails infested water bodies were surveyed for the presence of cercariae shedding Biomphalaria pfeifferi (Krauss) for a period of 12 months. A significantly marked (P < 0.01) fluctuation in infection rates in field B. pfeifferi was observed between the two areas. The factors contributing to the disparity in shedding capacities are linked to human behavioural pattern, and the drying up of water bodies. Their importance with respect to the control of intestinal schistosomiasis in the region are discussed.

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Biomphalaria tenagophila: genetic variability within intermediate snail hosts susceptible and resistant to Schistosoma mansoni infection.

DNA analysis by molecular techniques has significantly expanded the perspectives of the study and understanding of genetic variability in molluscs that are vectors of schistosomiasis. In the present study, the genetic variability of susceptible and resistant B. lenagophila strains to S. mansoni infection was investigated using amplification of their genomic DNA by RAPD-PCR. The products were analyzed by PAGE and stained with silver. The results showed polymorphism between tested strains with four different primers. We found two bands of 1,900 and 3,420 bp that were characteristic of the susceptible strains with primer 2. The primers 9 and 10 identified a single polymorphic band that was also characteristic of (3,136 and 5,041 bp, respectively) susceptible snails. Two polymorphic bands were detected by primer 15: one with 1,800 bp was characteristic of the resistant strain and the other with approximately equal to 1,700 bp in the susceptible one. These results provide additional evidence showing that the RAPD-PCR technique is adequate for the study of polymorphisms in intermediate hosts snails of S. mansoni. The obtained results are expected to expand the knowledge about the genetic variability of the snails and to permit the future identification of genomic sequences specifically related to the resistance/susceptibility of Biomphalaria to the larval forms of S. mansoni.

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The identification of markers segregating with resistance to Schistosoma mansoni infection in the snail Biomphalaria glabrata.

Both snail and parasite genes determine the susceptibility of the snail Biomphalaria glabrata to infection with the trematode Schistosoma mansoni. To identify molecular markers associated with resistance to the parasite in the snail host, we performed genetic crosses between parasite-resistant and -susceptible isogenic snails. Because resistance to infection in adult snails is controlled by a single locus, DNA samples from individual F2 and F1 backcross progeny, segregating for either the resistant or susceptible phenotypes, were pooled (bulked segregant). Genotypes for both parents were determined with 205 arbitrary decamer primers by random amplified polymorphic DNA-PCR. Of the 205 primers, 144 were informative, and the relative allele frequencies between the pools for these primers were determined. Two primers, OPM-04 and OPZ-11, produced fragments in the resistant parent of one cross that were inherited in a dominant fashion in the resistant F2 and backcross-bulked segregant progeny. Subsequent typing of DNA samples of individual progeny snails showed that the 1.2-kb marker amplified by primer OPM-04 and the 1.0-kb marker produced by primer OPZ-11 segregated in the same dominant fashion with the resistant phenotype. Sequence analysis of the 1.2-kb marker showed that it corresponds to a repetitive sequence in the snail genome with no homology to existing DNA sequences in the public databases. Analysis of the 1. 0-kb marker showed that it also corresponds to a repetitive sequence in the B. glabrata genome that contains an imperfect ORF, with homology to retrovirus-related group-specific antigens (gag) polyprotein.

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Emergence of cercariae of Echinostoma caproni and Schistosoma mansoni from Biomphalaria glabrata under different laboratory conditions.

Release of Echinostoma caproni cercariae and Schistosoma mansoni from experimentally infected Biomphalaria glabrata snails maintained under different laboratory conditions was studied. Infected snails were isolated individually for 1 h in Stender dishes containing 5 ml of artificial spring water and the number of cercariae released during this time was recorded. Of numerous conditions tested, the addition of lettuce, the use of water conditioned by B. glabrata snails and a temperature of 35 degrees C significantly increased the release of E. caproni cercariae. A significant increase in cercarial release of S. mansoni was seen only in cultures fed lettuce. A temperature of 12 degrees C caused a significant decrease in cercarial release of both E. caproni and S. mansoni. Increased snail activity associated with feeding behaviour was probably responsible for the enhanced cercarial sheds observed in this study.

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Effects of snail size on encystment of Echinostoma caproni in juvenile Biomphalaria glabrata (NMRI strain) and observations on the survival of infected snails.

The effects of snail size on encystment of Echinostoma caproni cercariae in neonatal and juvenile Biomphalaria glabrata (NMRI strain) snails were studied. Encystment in neonatal (0.7-1.1 mm shell diameter) and juvenile (2-3 mm shell diameter) snails was compared 24 h post-infection (PI) following individual exposure of snails of each size to 1, 5, 10, 25 and 50 cercariae. Significantly more cysts were recovered from juveniles exposed to 1, 5, 10 and 50 cercariae than from neonatals with comparable exposure. Size of B. glabrata was a major factor in determining cyst burden in this planorbid. Survival of infected versus uninfected neonatals and juveniles was also examined for 7 days. Neonatals exposed to 10 cercariae showed a significant decrease in survival at 3, 6 and 7 days PI when compared to the uninfected controls. There was no significant decrease in the survival of juveniles exposed to 10 cercariae compared to uninfected controls at any time point. Snail size was a factor in mortality associated with echinostome cercarial penetration and encystment.

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Effects of Schistosoma mansoni infection on inorganic elements in the snail Biomphalaria glabrata.

Inductively coupled plasma atomic emission spectrometry (ICP-AES) was used to study element ions in whole bodies of uninfected Biomphalaria glabrata snails and those experimentally infected with larval Schistosoma mansoni trematodes. Infected snails were analysed 8 weeks post-infection. Cohort snails that were left uninfected were analysed at the same time as the infected snails. Sixteen elements (aluminum, boron, barium, calcium, cadmium, copper, iron, potassium, magnesium, manganese, sodium, nickel, lead, selenium, tin and zinc) were found to be present in infected and uninfected whole bodies at concentrations above the detection limit of the ICP-AES analysis. Of these, calcium, cadmium, manganese and sodium were present in significantly higher amounts (Student's t-test, P<0.05) in whole infected versus whole uninfected snails. Variations in the present results compared with other studies reflect intrinsic differences in the larval trematode-snail systems used.

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Are Biomphalaria snails resistant to Schistosoma mansoni?

Among Biomphalaria glabrata/Schistosoma mansoni snail-trematode combinations, it appears that some parasites succeed whilst others fail to infect snails. Snails that become infected are termed susceptible hosts. Those which are not infected are traditionally determined as 'resistant'. Here the concept of B. glabrata resistance to S. mansoni is re-examined in the light of additional observations. It is suggested that, in B. glabrata/S. mansoni, compatibility is tested independently for each individual miracidium and host, and that the success or failure of an infection does not depend on the snail susceptibility/resistance status, but on the 'matched' or 'mismatched' status of the host and parasite phenotypes.

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Population genetics of the schistosome intermediate host Biomphalaria pfeifferi in the Zimbabwean highveld: implications for co-evolutionary theory.

Co-evolutionary theory proposes that polymorphisms in co-evolved traits may be maintained through differential selection in spatially heterogeneous environments. Spatial heterogeneity of the schistosome intermediate host, Biomphalaria pfeifferi, was investigated here, using RAPD markers. Overall, 256 individuals, collected at 32 sites on 13 rivers in the Chiweshe region of Zimbabwe, were analysed. Significant genetic differentiation was demonstrated, both between populations from the different rivers and between populations collected at different sites on each of several of the rivers investigated. However, the presence of spatial differentiation between populations from individual rivers varied with river type. It was not apparent in permanent, deep, fast-flowing rivers, where high levels of migration, through passive dispersal along the rivers, are likely. The snails collected from shallow, semi-permanent rivers not only showed relatively high levels of population subdivision but also high levels of within-site genetic diversity, consistent with the existence of 'co-evolutionary hot-spots' where schistosome infection may be prevalent. These results are discussed with reference to the population biology of B. pfeifferi and to host-parasite co-evolution.

Analysis of Variance↗

Spatio-temporal genetic variability in the schistosome intermediate host Biomphalaria pfeifferi.

Genetic variation among temporally and spatially defined populations of the schistosome intermediate host, Biomphalaria pfeifferi, was analysed using RAPD markers. Snails were collected from each of eight sites that were 1-20 km apart, on four occasions, over a 2-year period. High levels of genetic variability were found within populations, between populations and between collections. Spatial differentiation and intrapopulation similarity were highest during periods of drought and were lowest following heavy rainfall. Temporal variability, consistent with founder effects, genetic drift and/or differential parasite pressure, was of similar or greater magnitude to that of the spatial variability. The results not only demonstrate that spatio-temporal sampling is necessary to understand the population genetics of short-lived species in variable habitats, but also provide empirical data to support current theoretical models of host-parasite co-evolution.

Analysis of Variance↗

The influence of stream geology on the distribution of the bilharzia host snails, Biomphalaria pfeifferi and Bulinus (physopsis) sp.

The Gladdespruit and Komati River surveys have shown that similar environmental conditions have developed where the watercourses flowed over particular rock types. In these watercourses a potentially useful association was found between the occurrence of permanent, lentic habitats produced by the weathering of bedrock with a hardness above 5 in Mohs' Scale of Hardness and the longitudinal distribution of persistent populations of the bilharzia intermediate host snails Biomphalaria pfeifferi and Bulinus (Physopsis) sp. Bilharzia transmission follows this pattern in these waterways. Weathering of bedrock with hardness below 5 in Mohs' scale, and/or flow over beds of alluvial sand, produced perpetually lotic (flowing) environments without persistent host snail populations because of the snail's inability to tolerate high current speeds. However a few temporary populations survived in temporarily lentic habitats during the dry winter season.

Africa↗

Study of growth of Biomphalaria glabrata Say and other Planorbidae in Guadeloupe West Indies.

The growth of Biomphalaria glabrata, B. schrammi, Drepanotrema lucidum and D. kermatoides was studied in three habitatis on Guadeloupe. Theortical growth curves were calculated for each species by applying the Walford method and the von Bertalanffy equation. On Guadeloupe, the growth of B. glabrata was faster in the ponds of Grand Terre than in the small high altitude streams of Basse Terre. There is good correlation between the observations made in Guadeloupe and previous data obtained on Pureto Rico and St Lucia.

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Preliminary trials against Biomphalaria glabrata of a new molluscicide formulation: geletin granules containing Bayluscide wettable powder.

Tests were conducted with gelatin granules containing Bayluscide (niclosamide) w.p., 23-3% w/w a.i. Granules rendered a simulated field habitat (banana drain) lethal to Biomphalaria glabrata for 40 days when applied at a rate calculated to give 5 mg/1 in the water. At this dose, Bayluscide e.c. was effective for 8-12 days. Granules exposed to various combinations of natural or simulated tropical climatic factors for up to 56 days gave variable kills of snails, apparently unrelated to the factors tested. The rate at which the molluscicide was released from the granules was affected by climatic factors: high temperatures and low humidities slowed the rate, probably by hardening the granules; rainfall and high humidities accelerated the rate, probably by softening or disintegrating the granules. There was no apparent loss of molluscicidal activity in granules exposed for 56 days to various combinations of climatic factors. The results suggest that this formulation may be applied during droughts to temporary habitats when these are dry and accessible. The molluscicide would then be released at the unpredictable start of the next wet season and kill any snails which survive the drought by aestivation.

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