Calcium as a limiting factor in the biology of Biomphalaria pfeifferi (Krauss), (Gastropoda: Planorbidae)

The minimum calcium requirements, relative importance of buffering and optimum ratio of calcium to magnesium, calcium to sodium, and calcium to potassium ions were determined for laboratory populations ofBiomphalaria pfeifferi and related to suggested limiting factors for the natural distribution of this species. Snails were reared in a range of concentrations of both calcium bicarbonate and unbuffered calcium sulphate from 0.5 to 20 mg/l as Ca⁺⁺ and also in a series of media with a constant concentration of 2 mg/l as Ca⁺⁺ but with a range of Ca/Mg, Ca/Na and Ca/K ratios of 4.0 to 0.1. Shell growth, survivorship, fecundity, egg fertility, and the net reproductive rate were compared. In calcium bicarbonate cultures a concentration of 2mg/l Ca⁺⁺ appeared to be the lower limit for the survival of laboratory populations but a concentration of 4 mg/l Ca⁺⁺ was needed for a population to thrive. The calcium sulphate salt gave much poorer results, emphasizing the importance of the bicarbonate buffer. In the cationic ratio experiments the low Ca/Mg ratios proved to have the most damaging effects on snail populations but the effects of very low Ca/Na and Ca/K ratios could also be measured. A parallel experiment on the hatching rate of snail eggs, using similar experimental solutions, gave comparable results. The significance of these findings to snail ecology is discussed.     

Effects of intermittent starvation on mature Biomphalaria glabrata (Gastropoda: Planorbidae)

Mature Biomphalaria glabrata, submitted to four weeks of varied starvation times (0, 1, 3, 5, 6 & 7 d.week^–1, were thereafter refed during four weeks. The different intermittent starvation times had no significant effect on snails survival. As weekly starvation increased, the rate of change in body weight and fecundity decreased. In snails fed one or two d.week^–1, the rate of change in body weight was negative, while fecundity remained at a low level. Continuous hunger stopped oviposition.Starvation had no further effect on body growth after the first week of refeeding; however, its effect on fecundity remained significant over the two first weeks.     

Water relations and osmotic pressure in Biomphalaria pfeifferi (Krauss), Biomphalaria glabrata (Say) and Helisoma trivolvis (Say) (Gastropoda : Planorbidae) in response to cationic alterations of the medium

The percentage of water in the total living body weight is restricted within a narrow range for each species, even in snails cultured under differing controlled ionic concentrations in the medium. The water level in Helisoma trivolvis is consistently higher than in Biomphalaria pfeifferi and B. glabrata under the same culture conditions.The whole cationic fraction of the total osmoconcentration of the haemolymph is maintained at a constant hypertonic level, even though individual cation levels vary with the cationic composition of the surrounding medium.     

Neotropical schistosomiasis: African affinities of the host snail Biomphalaria glabrata (Gastropoda: Planorbidae)

Schistosoma mansoni , the blood fluke responsible for human intestinal schistosomiasis in the Neotropics, was imported repeatedly with African slaves during the period 1500–1800. This trematode, and its intermediate host snails of the genus Biomphalaria , are widely distributed across Africa and the disease is thought to have quickly become established in South America and the West Indies because of the presence of an endemic susceptible congener, B. glabrata. We compared B. glabrata with four other Neotropical and three African species of Biomphalaria using 20 allozyme loci and found that it is phenetically and phylogenetically more like the African species; both parasite and American host snail are apparently of historically or geologically recent African origin. Furthermore, genetic distances, cladistic analyses and fossil data suggest the African Biomphalaria species may themselves have evolved from Neotropical founders following an initial trans-Atlantic dispersal in the reverse direction 2.3–4.5 Mya. Interpretation of existing patterns remains problematic as few African snails have been characterized genetically and both B. glabrata and African B. pfeifferi appear to comprise several cryptic species.     

Cationic responses of organs and haemolymph of Biomphalaria pfeifferi (Krauss), Biomphalaria glabrata (Say) and Helisoma trivolvis (Say) (Gastropoda: Planorbirdae) to cationic alterations of the medium

The cationic responses of haemolymph, mantle collar, headfoot, gut + digestive diverticulum and ovotestis + albumin gland of three planorbids to cationic changes in the media are explored. Body organs exhibited cationic homeostasis, although attained with difficulty under very low Ca : Mg and Ca : Na ratios. Haemolymph imbalances were obtained at very low calcium concentrations and at very low Ca : Mg and Ca : Na ratios in the medium; this may be linked to competition for Ca²⁺ uptake sites in the epithelium by Mg²⁺ and possibly Na⁺ ions.Fecundity and shell growth in relation to calcium concentrations and to Ca : Mg and Ca : Na ratios are examined.Normal internal cationic levels, under optimal conditions, were obtained for each species. Significant interspecific differences for haemolymph sodium were found; magnesium levels were slightly higher in Biomphalaria spp. than in Helisoma trivolvis; potassium levels were all similar. Amounts of tissue calcium decreased as follows: Mantle collar head-foot ovotestis + albumin gland-gut + diverticulum. Tissue magnesium levels in the gut were low. H. trivolvis had highest tissue calcium and sodium; B. glabrata had highest tissue potassium.     

Effects of starvation on growth and reproductive apparatus of two immature freshwater snails Biomphalaria pfeifferi and Biomphalaria glabrata (Gastropoda: Planorbidae)

Starvation of immature snails of Biomphalaria pfeifferi and B. glabrata results in an arrest of growth and animals remain immature. Spermatogenesis is limited to spermatogonia (B.g.) or spermatocytes 1 (B.p.). The number and the size of oocytes remain inferior to that of controls. Animals show reduced genital tracts.Once feeding is restored, growth is resumed but wet weight and shell diameter do not reach the same level as in controls. Fecundity and gametogenesis do not differ from that in controls. Genital tracts weight is proportional to body weight.     

Effects of environmental calcium on fecundity and cercarial production of Biomphalaria glabrata (Say) infected with Schistosoma mansoni Sambon

Biomphalaria glabrata were reared in stock culture and subjected to either 7-day or 60-day acclimation periods in complex CaCO3 media with calcium values ranging from 1.5 mg/L to 75 mg/L. Following 60-day acclimation, snails from series I were each exposed to 8 miracidia of Schistosoma mansoni. Snails of series II were each exposed to a single miracidium. Snails of both experimental regimens were observed for mortality, growth, rate of infection, and number of cercariae shed. Series I snails were also monitored for fecundity during acclimation and following miracidial exposure. Calcium levels of 1.5 and 75 mg/L resulted in significant snail mortality. Shell growth and rates of infection were positively correlated with calcium maintenance level. Snails with high fecundity prior to miracidial exposure subsequently shed more cercariae. In contrast, post-exposure (PE) fecundity of snails reared in media with up to 30 mg/L Ca++ were negatively correlated with calcium level, infection rate, and number of cercariae shed. Maximal cercarial emergence occurred at 30 mg/L Ca++. These results suggest that environmental calcium affects both the distribution patterns of snail hosts of human schistosomes and the productivity of intramolluscan schistosome infection.     

Characterization of the bacterial microbiota of Biomphalaria glabrata (Say, 1818) (Mollusca: Gastropoda) from Brazil

Roughly 200 000 000 people in 74 countries infected with schistosomes all share the fact that they came in contact freshwater harbouring infected snails. The aim of the study is to characterize the microbiota of wild and laboratory‐reared snails of Biomphalaria glabrata from Pernambuco, Brazil. The microbiota of these molluscs was identified biochemically by the VITEK 2 automated microbiological system. Antimicrobial susceptibility testing was carried out by the disc diffusion method with ß‐lactam antibiotics, aminoglycosides, quinolones, folate pathway inhibitors, fenicols and tetracyclines. The results showed that all bacteria identified were gram‐negative, including 11 bacterial genera: Aeromonas, Citrobacter, Enterobacter, Cupriavidus, Rhizobium, Stenotrophomonas, Pseudomonas, Klebsiella, Acinetobacter, Vibrio and Sphingomonas. Regarding the antimicrobial susceptibility, all the isolates exhibited resistance to amoxicillin and sensitivity to meropenem (beta‐lactam antimicrobials). The microbiota of the wild snails consisted predominantly of Enterobacter cloacae, while the laboratory‐reared snails predominantly showed Citrobacter freundii and Aeromonas sobria.

Significance and Impact of the Study

Biomphalaria glabrata is a Brazilian freshwater Planorbidae of great medical relevance as an intermediate host of Schistosoma mansoni. About a month after being infected by one or more miracidia larvae of a compatible schistosome, B. glabrata sheds thousands of cercariae into the water where they seek human skin and, if successful, penetrate to establish infection, eventually taking up residence and maturing in blood vessels of the small intestine. Results obtained from this study aim at targeting novel biological control strategies for schistosomiasis such as paratransgenesis. This is the first study on the microbiota of B. glabrata from Brazil.     

The effect of calcium concentration on the crushing resistance, weight and size of Biomphalaria sudanica (Gastropoda: Planorbidae)

Biomphalaria sudanica (intermediate host of Schistosoma mansoni) were raised in the laboratory in media with seven different calcium concentrations ranging from 0.02 to 2.0 mmol/l. After 10 weeks, snails were killed and shell diameter, inorganic dry weight, ash free dry weight and crushing resistance were measured. Snails raised at lower calcium concentrations were found to be significantly smaller than snails raised at higher concentrations. Furthermore, relatively thinner shells were developed at low concentrations and crushing resistance relative to snail size was found to be lower for these snails. The ratio between crushing resistance and ash free dry weight (as a measure for the energetic cost/benefit-ratio) was also found to be correlated with calcium concentration. The relevance of the results for predation risk of B. sudanica is discussed.     

Impact of geographical origin on mating behaviour in two species of Biomphalaria (Planorbidae: Gastropoda)

Studies of inbreeding and outcrossing have traditionally concentrated on matings within populations. The influence of geographical origin on mate choice in animals from different populations has received less attention. We investigated whether planorbid snails mated preferentially within their own population or with snails from other populations. Snails from three Biomphalaria pfeifferi strains and three B. glabrata strains were allowed to mate with conspecifics in the laboratory. We recorded their matings at night using time-lapse video. When they could choose between sympatric and allopatric snails, Biomphalaria snails significantly preferred the former: snails of each population mated more often with sympatric than with allopatric snails. This tendency to avoid outcrossing may indicate that, in some species, local adaptations can be more valuable than genetic novelties. Copyright 1999 The Association for the Study of Animal Behaviour.     

Microgeographic differentiation among closely related species of Biomphalaria (Gastropoda: Planorbidae) from the Andean Altiplano

Direct development and water dependence entail limited vagility in freshwater fauna. In these organisms, the population structure is probably linked to restrictions imposed by the habitat. In this study we investigate the relative contribution of processes stimulating the divergence of populations of Biomphalaria costata (Biese, 1951) and Biomphalaria crequii (Courty, 1907), two freshwater snails occurring in two contiguous and fragmented closed basins from the Andean Altiplano using mitochondrial DNA (cytochrome c oxidase subunit I) sequences, shell morphometric and radular morphology. In order to clarify the species boundaries, a third allopatric species was included: Biomphalaria aymara Valdovinos & Stuardo, 1991. Molecular analyses recovered two distinct clades: one composed of B. aymara from the Isluga swamps and B. costata from Spring 1 in Salar de Carcote, the single spring occupied by this species, and another integrated by snails from 12 springs spread across the Salar de Carcote and the Salar de Ascotán assigned to B. crequii, originally described from the Salar de Ascotán. Unlike shell morphometrics, radular morphology was informative for distinguishing these species. The division of the lineages occurred in the Late Pleistocene. A subclade that includes snails from the southernmost springs in Salar de Ascotán suggests fragmentation of the distribution of B. crequii associated with landscape discontinuities. In addition to microvicariance signals, the private haplotypes scattered around both salt spans show that close‐range dispersal is a common biogeographic process in this species. As evolutionary units, the single isolated and restricted population of B. costata has a high priority for conservation. © 2013 The Linnean Society of London     

Influence of Echinostoma paraensei (Lie and Basch, 1967) infection on the calcium content in Biomphalaria glabrata (Say, 1818)

The calcium content in the hemolymph and shell of Biomphalaria glabrata (Say, 1818) was determined after exposure to different parasite burdens (5 and 50 miracidia) of Echinostoma paraensei (Lie and Basch, 1967). The snails were dissected 1, 2, 3, and 4 weeks after infection to collect the hemolymph and shell. An increase in calcemia was observed in snails infected with both miracidial doses. A significant decrease in the calcium ions in the shell was observed, coinciding with the calcemia peak in the hemolymph. This indicates greater mobilization of calcium between the shell and hemolymph to regulate the calcium content in the body when the snail is exposed to stress conditions, as has also been observed in some other infected snail species. The results obtained indicate that in this model, the calcium metabolism depends on the miracidial dose used.     

Relationship between light conditions and behavior of the freshwater snail Biomphalaria glabrata (Say)

The influence of light upon behavior of Biomphalaria glabrata was investigated in snails submitted for 48 h to one of the following regimes: normal light cycle, continuous darkness, continuous light. Time-lapse cinematography was used to provide data about snail locomotor activity in response to (a) continuous light or darkness; (b) light or dark phases; (c) light transitions. Snails were significantly less active under continuous light than under continuous or intermittent darkness. Under the normal light cycle, the activity rate was significantly higher in the dark than in the light. Changes from light to dark corresponded to increases in the activity rate which persisted long afterwards. No significant variation in activity occurred upon changes from dark to light.     

Phylogenetic relationships and taxonomy of Altiplano populations of Biomphalaria (Gastropoda: Planorbidae): inference from a multilocus approach

Isolated closed basins provide a natural laboratory to study the differentiation among wild populations. Here we examined the phylogenetic relationships of the Southern Altiplano populations of Biomphalaria, a genus with medical importance, using nuclear (ITS1, ITS2) and mitochondrial (16S) ribosomal gene markers and a species of Helisoma as outgroup. Phylogenetic trees based on separate and combined analyses show that these populations form a particular lineage within Biomphalaria along with Biomphalaria peregrina (d'Orbigny, 1835) and Biomphalaria oligoza Paraense, 1974. The origin of this clade was estimated to have occurred in the middle Pleistocene. Molecular analyses showed that the nominal species Biomphalaria crequii ( Courty, 1907 ) from the Salar de Ascotán and Biomphalaria costata (Biese, 1951) from the Salar de Carcote, previously synonymized with Biomphalaria andecola (d'Orbigny, 1835) and B. peregrina, respectively, are distinct taxa. Molecular data did not resolve the relationship of Biomphalaria aymara Valdovinos & Stuardo, 1991 from the Isluga swamps to other Biomphalaria species, but confirm that the populations from the Lauca and Huasco basins may represent a distinct undescribed species of Biomphalaria from the Southern Altiplano. Snails examined for trematodes were found to be positive in some Altiplano localities. © 2012 The Linnean Society of London, Zoological Journal of the Linnean Society, 2012, 165 , 795–808.     

Effects of Plagiorchis elegans (Digenea: Plagiorchiidae) infection of Biomphalaria glabrata (Pulmonata: Planorbidae) on a challenge infection with Schistosoma mansoni (Digenea: Schistosomatidae)

Prior exposure of Biomphalaria glabrata to the eggs of an incompatible digenean, Plagiorchis elegans, rendered this snail host less suitable to a compatible species, Schistosoma mansoni. Although P. elegans failed to develop patent infections in B. glabrata, it reduced the production of S. mansoni cercariae by 88%. Concomitantly, host attributes such as reproduction, growth, and survival were compromised. The effect of P. elegans infection was most severe among snails that, in addition, had developed patent schistosome infections. Although few S. mansoni cercariae were produced, egg production by B. glabrata was only 4% of control values. Furthermore, no doubly infected snails survived for more than 3 wk after patency, whereas controls experienced no mortality during the same time period. The above effects were attributable to the establishment and persistence of P. elegans sporocysts in the tissues of the incompatible snail host. Their indirect antagonistic interaction with thelarval stages of S. mansoni may be mediated, in part, through their long-term stimulation of the host's internal defense mechanisms. These findings are discussed with a view to use P. elegans and other plagiorchiid digeneans as agents in the biological control of snails and snail-borne diseases.     

Phylogenetic analysis of eleven species of Biomphalaria Preston, 1910 (Gastropoda: Planorbidae) based on comparisons of allozymes

Freshwater snails in the genus Biomphalaria transmit Schistosoma mansoni in Africa, South America and the Caribbean region. Although considerable attention has been given to the identification of species, little is known of evolutionary relationships among the species. A phylogenetic analysis of 25 populations representing 11 species was performed on 25 enzyme loci examined using starch gel electrophoresis. A phylogenetic analysis of the individual populations produced 60 trees of equal length. The 60 trees have a consistency index value of 75.9% and a retention index value of 76.5%. The phylogenetic analysis provided strong support for the monophyly of Biomphalaria with either 14 or 15 synapomorphies uniting all of the species included and separating them from the outgroup, two species of Helisoma. Four nominal species represented by multiple populations formed monophyletic groups. Populations of B. sudanica, B. choanomphala, and B. alexandrina were interspersed. Ten arrangements were obtained for the populations of these three species. A variety of ingroup taxa were used to root the trees, and all provided support for the use of Helisoma species as an outgroup. In all of the trees obtained, the African species together formed a monophyletic group. In none of the trees obtained did the neotropical species form a monophyletic group. A constrained analysis requiring the monophyly of the neotropical species as well as the African species resulted in 90 trees just two steps longer than the shortest trees. Analysis of the species from either hemisphere alone resulted in decreased resolution, as measured by an increase in the number of trees obtained. This finding suggests that further comparisons of species from the two hemispheres will be of considerable value. Finally, two species which are resistant to infection with S. mansoni were included among the eleven studied. Neither of these species formed the sister group to all of the other species included, indicating that susceptibility is the plesiomorphic state, and that resistance is derived. Similarly, in none of the trees obtained did the two resistant species fall out as sister taxa, indicating that resistance arose independently twice.     

Parental size and environmental conditions affect egg capsule production by Nassarius reticulatus (Linnaeus 1758) (Gastropoda: Nassariidae)

In shallow coastal habitats scavenging netted whelks Nassarius reticulatus attached egg capsules to the stipes of red algae Chondrus crispus and occasionally on Furcellaria lumbricalis and Plumaria plumose. In the laboratory egg capsules were laid on aquaria sides and lids by individuals ≥ 21 mm shell length. Larger size classes produced more egg capsules and spawned over a longer period and in doing so partitioned less energy into shell growth. Large netted whelks (25-28.9 mm) produced larger capsules which contained significantly more and larger eggs than those produced by smaller individuals (21-24.9 mm). Egg capsule production continued throughout the year by regularly fed N. reticulatus held at ambient seawater temperatures. Egg production increased in the spring and summer with peak production during June (15 °C), decreased between August and October and resumed again during the winter (November to February at ∼ 7 °C). During the summer (15-16 °C) egg capsules were smaller and contained smaller eggs than those deposited during the winter (7-10 °C), although the number of eggs · capsule⁻¹ was similar. Enforced food limitation reduced the number and size of the egg capsules, the number and size of eggs produced · female⁻¹ and the duration of the breeding period. Hatching success of N. reticulatus egg capsules was high (95%) even at winter seawater temperatures (11-8.5 °C) and the duration of embryonic development was fastest between 15 and 17.5 °C.     

Usnic Acid Potassium Salt: An Alternative for the Control of Biomphalaria glabrata (Say, 1818)

In Brazil, the snail Biomphalaria glabrata is the most important vector of schistosomiasis due to its wide geographical distribution, high infection rate and efficient disease transmission. Among the methods of schistosomiasis control, the World Health Organization recommends the use of synthetic molluscicides, such as niclosamide. However, different substances of natural origin have been tested as alternatives for the control or eradication of mollusks. The literature describes the antitumor, antimicrobial and antiviral properties of usnic acid as well as other important activities of common interest between medicine and the environment. However, usnic acid has a low degree of water solubility, which can be a limiting factor for its use, especially in aquatic environments, since the organic solvents commonly used to solubilize this substance can have toxic effects on aquatic biota. Thus, the aim of the present study was to test the potassium salt of usnic acid (potassium usnate) with regard to molluscicidal activity and toxicity to brine shrimp (Artemia salina). To obtain potassium usnate, usnic acid was extracted with diethyl ether isolated and purified from the lichen Cladonia substellata. Biological assays were performed with embryos and adult snails of B. glabrata exposed for 24 h to the usnate solution solubilized in dechlorinated water at 2.5; 5 and 10 µg/ml for embryos, 0.5; 0.9; 1;5 and 10 µg/ml for mollusks and 0.5; 1; 5; 10 µg/ml for A. salina. The lowest lethal concentration for the embryos and adult snails was 10 and 1 µg/ml, respectively. No toxicity to A. salina was found. The results show that modified usnic acid has increased solubility (100%) without losing its biological activity and may be a viable alternative for the control of B. glabrata.     

Ecology of Biomphalaria (Gastropoda: Planorbidae) in Lake Albert, Western Uganda: snail distributions, infection with schistosomes and temporal associations with environmental dynamics

In Lake Albert, an ecological study was conducted, between June 2000 and May 2003, which assessed snail population dynamics, parasite infection patterns and interplay of environmental factors upon Biomphalaria. Monthly sampling surveys were conducted at 29 sites monitoring populations of Biomphalaria stanleyi and Biomphalaria sudanica. Altogether, a total of 21,715 B. stanleyi and 8452 B. sudanica were collected during the period. Both species could be found infected with Schistosoma mansoni although infection prevalence was significantly higher (p < 0.01**) in B. stanleyi (4.4%) than in B. sudanica (3.5%). Each species occupied slightly different aquatic niches with B. stanleyi preferring deeper water habitats whilst B. sudanica was found along the shoreline in shallower water. B. stanleyi was more widely distributed among the sampling locations (19 sites) than B. sudanica (10 sites). Of the four villages included in the study area, snails from sites near Piida and Bugoigo villages had the highest schistosome infection rates, presumably attributable to the closer proximity of people with intestinal schistosomiasis. After inspection of cross-correlation plots which identified most suitable time lags, snail density dynamics could be associated with seasonal variations inclusive of: air temperature, rainfall, lake level, water temperature, water conductivity and water pH. These temporal observations better reveal the relationship between snail populations and environmental factors, providing important information concerning the relative roles of B. stanleyi and B. sudanica in transmission of S. mansoni and development of integrated strategies for disease control around Lake Albert.