Land snails can trap trematode cercariae in their shell: Encapsulation as a general response against parasites?

Terrestrial gastropods are hosts of a wide variety of metazoan parasites and can respond to parasite exposure in various ways. One of these defence mechanisms, the ability to trap parasites in the host shell, was previously thought to apply only against nematodes. During a field survey along an urbanisation gradient, we found that the shell of Cornu aspersum and Cepaea nemoralis can contain encapsulated trematode cercariae, with prevalences of 7% and 1%, respectively over the entire sample, and up to 47% at the local population level. To our knowledge, this is the first case study unambiguously showing that land snails can trap non-nematode parasites in their shell at non-negligible prevalences. Shell-encapsulation could be a more general defence mechanism than previously described, and more studies are needed to understand its importance and variability.

Title: Les gastéropodes terrestres peuvent piéger les cercaires de trématodes dans leur coquille : l'encapsulation comme réponse générale contre les parasites ? Abstract: Les gastéropodes terrestres sont les hôtes d'une grande variété de métazoaires parasites, et peuvent répondre de plusieurs façons à l'exposition parasitaire. L'un de ces mécanismes de défense, la capacité à piéger le parasite dans la coquille, semblait ne concerner que les nématodes. Lors d'un suivi de terrain le long d'un gradient d'urbanisation, nous avons observé que la coquille de Cornu aspersum et de Cepaea nemoralis pouvait contenir des cercaires de trématodes encapsulées, à des prévalences respectives de 7 % et 1 % sur l'ensemble de l'échantillon, et pouvant atteindre 47 % au niveau d'une population locale. À notre connaissance, ceci est la première étude montrant sans ambiguïté que les escargots terrestres peuvent piéger de façon significative des parasites autres que les nématodes dans leur coquille. L'encapsulation des parasites dans la coquille pourrait donc être un mécanisme de défense plus général que précédemment décrit, plus d'études étant nécessaires pour comprendre son importance et sa variabilité.

Dispersal syndromes in challenging environments: A cross-species experiment.

Dispersal is a central biological process tightly integrated into life-histories, morphology, physiology and behaviour. Such associations, or syndromes, are anticipated to impact the eco-evolutionary dynamics of spatially structured populations, and cascade into ecosystem processes. As for dispersal on its own, these syndromes are likely neither fixed nor random, but conditional on the experienced environment. We experimentally studied how dispersal propensity varies with individuals' phenotype and local environmental harshness using 15 species ranging from protists to vertebrates. We reveal a general phenotypic dispersal syndrome across studied species, with dispersers being larger, more active and having a marked locomotion-oriented morphology and a strengthening of the link between dispersal and some phenotypic traits with environmental harshness. Our proof-of-concept metacommunity model further reveals cascading effects of context-dependent syndromes on the local and regional organisation of functional diversity. Our study opens new avenues to advance our understanding of the functioning of spatially structured populations, communities and ecosystems.

Brachylaima spp. (Trematoda) parasitizing Cornu aspersum (Gastropoda) in France with potential risk of human consumption.

The edible land snail Cornu aspersum, native to the Mediterranean coastlines of North Africa, is widely distributed on most continents and often invasive in areas where introduction is recent. This species could contribute to the geographic spread of parasites as demonstrated for Brachylaima spp. These cosmopolitan trematodes may represent a threat to human health, like in Australia where Brachylaima cribbi infects humans. In this study, we demonstrate for the first time the occurrence of Brachylaima spp. in two French populations of C. aspersum, Thorigné-Fouillard (Ille-et-Vilaine), and Arçais (Deux-Sèvres), with an overall prevalence of 10.4% (Thorigné-Fouillard) and 73.3% (Arçais), respectively and a metacercarial intensity on average three times higher in Thorigné-Fouillard (37) than in Arçais (11). Cornu aspersum may act as a first and second intermediate host, as demonstrated in Arçais. The morphometrics of metacercariae, particularly the great body length about 2 mm, discriminate our Brachylaima species from those already described in C. aspersum (B. cribbi in Australia, and B. aspersae, B. llobregatensis and B. mascomai in Europe). Molecular analysis, based on 28S and COI, suggests the occurrence of two species in our study, one of which is probably Brachylaima mesostoma, an intestinal parasite of passeriform birds described in Central Europe. We underline the need for further research to identify species of Brachylaima in France and measure the health hazard of consuming field-collected snails.

TITLE: Brachylaima spp. (Trematoda) parasitant Cornu aspersum (Gastropoda) en France et risque potentiel pour la consommation humaine. ABSTRACT: L'escargot Cornu aspersum, originaire des côtes méditerranéennes d'Afrique du Nord, est largement répandu sur la plupart des continents et souvent invasif dans ses aires d'introduction récente. Cette espèce peut contribuer à l'expansion géographique de parasites comme démontré pour Brachylaima spp. Ces trématodes cosmopolites peuvent représenter une menace pour la santé comme en Australie où Brachylaima cribbi parasite l'espèce humaine. Dans cette étude, nous démontrons pour la première fois la présence de Brachylaima spp. dans deux populations françaises de C. aspersum, à Thorigné-Fouillard (Ille-et-Vilaine) et Arçais (Deux-Sèvres), avec une prévalence totale de 10.4 % (Thorigné-Fouillard) et 73.3 % (Arçais), et une intensité de métacercaires en moyenne trois fois plus importante à Thorigné-Fouillard (37) qu'à Arçais (11). Cornu aspersum peut jouer le rôle de premier et de second hôte intermédiaire, comme démontré à Arçais. La morphométrie des métacercaires, en particulier la grande taille corporelle d'environ 2 mm de long, suggèrent qu'il ne s'agit pas d'espèces de Brachylaima déjà décrites chez C. aspersum (B. cribbi en Australie et B. aspersae, B. llobregatensis et B. mascomai en Europe). L'analyse moléculaire, basée sur 28S et COI, suggère la présence de deux espèces dans notre étude, dont l'une est probablement Brachylaima mesostoma, un parasite intestinal d'oiseaux passeriformes décrit en Europe Centrale. Nous soulignons le besoin de recherche future pour identifier les espèces de Brachylaima présentes en France et mesurer le risque associé à la consommation d'escargots terrestres prélevés en milieu naturel.

Dispersers are more likely to follow mucus trails in the land snail Cornu aspersum.

Dispersal, movement leading to gene flow, is a fundamental but costly life history trait. The use of indirect social information may help mitigate these costs, yet we often know little about the proximate sources of such information, and how dispersers and residents may differ in their information use. Terrestrial molluscs, which have a high cost of movement and obligatorily leave information potentially exploitable by conspecifics during movement (through mucus trails), are a good model to investigate links between dispersal costs and information use. We studied whether dispersers and residents differed in their trail-following propensity in the snail Cornu aspersum. Dispersers followed mucus trails more frequently than expected by chance, contrary to non-dispersers. Trail-following by dispersers may reduce dispersal costs by reducing energy expenditure and helping snails find existing habitat or resource patches. Finally, we point that ignoring the potential for collective dispersal provided by trail-following may hinder our understanding of the demographic and genetic consequences of dispersal.

Bottom-up and top-down control of dispersal across major organismal groups.

Ecology and evolution unfold in spatially structured communities, where dispersal links dynamics across scales. Because dispersal is multicausal, identifying general drivers remains challenging. In a coordinated distributed experiment spanning organisms from protozoa to vertebrates, we tested whether two fundamental determinants of local dynamics, top-down and bottom-up control, generally explain active dispersal. We show that both factors consistently increased emigration rates and use metacommunity modelling to highlight consequences on local and regional dynamics.

Refining the biogeographical scenario of the land snail Cornu aspersum aspersum: Natural spatial expansion and human-mediated dispersal in the Mediterranean basin.

The land snail Cornu aspersum aspersum, native to the Mediterranean region, has been the subject of several anatomical and molecular studies leading to recognize two divergent lineages, named "East" and "West" according to their geographical distribution in North Africa. The first biogeographical scenario proposed the role of Oligocene paleogeographic events and Quaternary glacial refugia to explain spatial patterns of genetic variation. The aim of this study was to refine this scenario using molecular and morphometric data from 169 populations sampled across Mediterranean islands and continents. The two previously described lineages no longer correspond to distinct biogeographical entities. Phylogenetic relationships reveal the existence of seven clades, do not support the Tyrrhenian vicariance hypothesis, and suggest that C. a. aspersum most likely originates from North Africa. We found two contrasted patterns with the seven clades defining spatially well-structured populations in the southern Mediterranean whereas one clade is distributed across the basin. High genetic diversities and rates of endemism in North Africa support the role of this region for the diversification of C. a. aspersum. In referring to divergence times previously estimated, we suggest allopatric differentiation due to geological changes of the Atlas system and multiple refugial areas during Pleistocene glaciations. The new biogeographical scenario implies an initial range expansion from North Africa to the Iberian Peninsula and the peri-Tyrrhenian regions through land bridges connections during the Messinian Salinity Crisis and Pleistocene glaciations. Historical events appear to have also structured morphometric variation but recent dispersal events favored the emergence of secondary contacts between clades. Southern Mediterranean clades are limited to their initial distribution and populations of the recent clade would have rapidly recolonized the whole Mediterranean in the Holocene due to greater adaptive potential and the influence of human transportations.

Conservation at a slow pace: terrestrial gastropods facing fast-changing climate.

The climate is changing rapidly, and terrestrial ectotherms are expected to be particularly vulnerable to changes in temperature and water regime, but also to an increase in extreme weather events in temperate regions. Physiological responses of terrestrial gastropods to climate change are poorly studied. This is surprising, because they are of biodiversity significance among litter-dwelling species, playing important roles in ecosystem function, with numerous species being listed as endangered and requiring efficient conservation management. Through a summary of our ecophysiological work on snail and slug species, we gained some insights into physiological and behavioural responses to climate change that we can organize into the following four threat categories. (i) Winter temperature and snow cover. Terrestrial gastropods use different strategies to survive sub-zero temperatures in buffered refuges, such as the litter or the soil. Absence of the insulating snow cover exposes species to high variability in temperature. The extent of specific cold tolerance might influence the potential of local extinction, but also of invasion. (ii) Drought and high temperature. Physiological responses involve high-cost processes that protect against heat and dehydration. Some species decrease activity periods, thereby reducing foraging and reproduction time. Related costs and physiological limits are expected to increase mortality. (iii) Extreme events. Although some terrestrial gastropod communities can have a good resilience to fire, storms and flooding, an increase in the frequency of those events might lead to community impoverishment. (iv) Habitat loss and fragmentation. Given that terrestrial gastropods are poorly mobile, landscape alteration generally results in an increased risk of local extinction, but responses are highly variable between species, requiring studies at the population level. There is a great need for studies involving non-invasive methods on the plasticity of physiological and behavioural responses and the ability for local adaptation, considering the spatiotemporally heterogeneous climatic landscape, to allow efficient management of ecosystems and conservation of biodiversity.

Density-dependence across dispersal stages in a hermaphrodite land snail: insights from discrete choice models.

Dispersal movements, i.e. movements leading to gene flow, are key behaviours with important, but only partially understood, consequences for the dynamics and evolution of populations. In particular, density-dependent dispersal has been widely described, yet how it is determined by the interaction with individual traits, and whether density effects differ between the three steps of dispersal (departure, transience, and settlement), remains largely unknown. Using a semi-natural landscape, we studied dispersal choices of Cornu aspersum land snails, a species in which negative effects of crowding are well documented, and analysed them using dispersal discrete choice models, a new method allowing the analysis of dispersal decisions by explicitly considering the characteristics of all available alternatives and their interaction with individual traits. Subadults were more dispersive than adults, confirming existing results. In addition, departure and settlement were both density dependent: snails avoided crowded patches at both ends of the dispersal process, and subadults were more reluctant to settle into crowded patches than adults. Moreover, we found support for carry-over effects of release density on subsequent settlement decisions: snails from crowded contexts were more sensitive to density in their subsequent immigration choices. The fact that settlement decisions were informed indicates that costs of prospecting are not as important as previously thought in snails, and/or that snails use alternative ways to collect information, such as indirect social information (e.g. trail following). The observed density-dependent dispersal dynamics may play an important role in the ability of C. aspersum to successfully colonise frequently human-disturbed habitats around the world.

Movement propensity and ability correlate with ecological specialization in European land snails: comparative analysis of a dispersal syndrome.

Intra- and interspecific differences in movement behaviour play an important role in the ecology and evolution of animals, particularly in fragmented landscapes. As a consequence of rarer and generally more fragmented habitat, and because dispersal tends to disrupt benefits brought by local adaptation, theory predicts that mobility and dispersal should be counter-selected in specialists. Using experimental data and phylogenetic comparative tools, we analysed movement propensity and capacity, as well as dispersal-related phenotypic traits, in controlled conditions in 20 species of European land snails from the Helicoidea superfamily. Costs of movement in terrestrial gastropods are among the highest in animals, which make them a potentially powerful model to test these predictions. Habitat specialists were indeed less likely to cross a boundary between a familiar and an unfamiliar substrate than generalists. They also had smaller feet, after accounting for size. Furthermore, exploring specialists were slower than generalists and had more tortuous trajectories, leading them to stay closer to the familiar patch. Movement traits were generally evolutionary labile, but some were constrained by body size, a phylogenetically conserved trait. High specialization and low-dispersal ability are two traits often considered to increase species vulnerability to fragmentation, climate changes and extinction. This study confirms they should not be considered separately, due to their integration in a dispersal syndrome. Therefore, specialist species face double penalty under habitat loss and other environmental changes, making them more vulnerable to extinction and contributing to the biotic homogenization of communities.

Study of narrow band millimeter-wave potential interactions with endoplasmic reticulum stress sensor genes.

The main purpose of this article is to study potential biological effects of low-power millimeter waves (MMWs) on endoplasmic reticulum (ER), an organelle sensitive to a wide variety of environmental insults and involved in a number of pathologies. We considered exposure frequencies around 60 GHz in the context of their near-future applications in wireless communication systems. Radiations within this frequency range are strongly absorbed by oxygen molecules, and biological species have never been exposed to such radiations in natural environmental conditions. A set of five discrete frequencies has been selected; three of them coincide with oxygen spectral lines (59.16, 60.43, and 61.15 GHz) and two frequencies correspond to the spectral line overlap regions (59.87 and 60.83 GHz). Moreover, we used a microwave spectroscopy approach to select eight frequencies corresponding to the spectral lines of various molecular groups within 59-61 GHz frequency range. The human glial cell line, U-251 MG, was exposed or sham-exposed for 24 h with a peak incident power density of 0.14 mW/cm(2). The average specific absorption rate (SAR) within the cell monolayer ranges from 2.64 +/- 0.08 to 3.3 +/- 0.1 W/kg depending on the location of the exposed well. We analyzed by quantitative reverse transcription-polymerase chain reaction (RT-PCR) the level of expression of two endogenous ER-stress biomarkers, namely, the chaperones BiP/GRP78 and ORP150/GRP170. It was found that exposure to low-power MMW does not significantly modify the mRNA levels of these stress-sensitive genes suggesting that ER homeostasis is not altered by low-power MMW at the considered frequencies.

Influence of temperature acclimation and gut content on the supercooling ability of the land snail Cornu aspersum.

The invasive land snail Cornu aspersum possesses a low ability to supercool (c. -5 degrees C in winter) and survives only minimal ice formation in its body fluids, what may limit its expansion to colder environments. In the present study, we investigated the influence of acclimation and starvation on its supercooling ability. During eight weeks, individuals were maintained at 20 degrees C, fed or starved, or placed at 5 degrees C, directly or with progressive acclimation to cold and shorter photoperiod. Temperature of crystallisation of whole individual (Tc(I)) and hemolymph (Tc(H)), mass data and gut content were recorded every two weeks. Hemolymphatic glucose and glycerol were measured at the end of experiment and occurrence of intestinal ice-nucleating agents (INA) was researched. Acclimation had no effect on Tc(I) but stimulated purging of the gut. Starvation induced a slight decrease of Tc(I) whereas a high quantity of alimentary particles in the digestive tract limited the supercooling ability. Glucose and glycerol were not synthesized in cold conditions. Mean Tc(H) was low (c. -17 degrees C), some INA being present in hemolymph of fed animals. Intestinal content of starved individuals exhibited a mean Tc of c. -6 degrees C, decreasing to c. -12 degrees after heating, suggesting the presence of organic INA.

Supercooling ability is surprisingly invariable in eggs of the land snail Cantareus aspersus.

From an ontogenetic point of view, invertebrate eggs are generally the most freezing intolerant stage of a species. Development state, water content and acclimation may affect their supercooling ability. In this study, we measured fresh mass, water content and temperature of crystallisation (T(c)) of eggs of the edible land snail Cantareus aspersus, depending on its form ("aspersa"vs. "maxima"), incubation temperature (20, 12 and 7 degrees C) and physiological age (as part of the complete development). We also tested their tolerance to freezing. Despite a high number of individual observations (n=759) and significant differences of fresh mass and water content between both subspecies, no effect of origin, incubation temperature or development state has been found in this study. T(c) remained constant whatever the condition, with an overall mean of -5.40+/-0.24 degrees C (mean+/-SD). We suggest that fresh mass is important, a high water content and a constantly wet surface confer to land snail eggs a poor ability to supercool. Moreover, the presence of ice nucleating agents at the egg surface (microorganisms present in the soil, calcium carbonate crystals of the egg shell) might induce freezing. Thus, considering the present results, to delay hatching by cryopreservation of eggs does not seem possible.

Supercooling ability in two populations of the land snail Helix pomatia (Gastropoda: Helicidae) and ice-nucleating activity of gut bacteria.

The land snail Helix pomatia (Gastropoda: Helicidae) is widely distributed in Northern and Central Europe where it may experience subzero temperatures during winter months. Its supercooling ability was studied in two populations of H. pomatia. One population originated from Southern Sweden (Gotaland) and the other from Central France (Auvergne). In the experimental design, they were acclimated, over 2 weeks, to artificial winter conditions (hibernation, T=5 degrees C). The Swedish snails showed a rather limited supercooling ability (temperature of crystallization, T(c)=-6.4+/-0.8 degrees C), significantly greater, however, than the supercooling capacity of the population from France (T(c)=-4.6+/-1.4 degrees C). In artificial spring conditions (3 months of hibernation followed by a progressive acclimation, over 2 weeks, to activity at T=20 degrees C), both populations exhibited a similar high T(c) (-2.0+/-1.0 degrees C). The lower T(c) of hibernating Swedish snails could be due to a greater loss of body water, accompanied by a higher concentration of solutes in the hemolymph. In both populations, the variation in hemolymph osmolality measured between hibernating (250-270 mOsm kg(-1)) and active (165-215 mOsm kg(-1)) snails may be explained by the variation in body water mass and did not suggest the production of colligative cryoprotectants. Moreover, the three bacterial strains, Buttiauxella sp., Kluyvera sp., and Tatumella sp. (Enterobacteriaceae) which were isolated from fed snails, but absent in starved snails, did not show any ice-nucleating activity at temperatures higher than -9 degrees C. Only the strain Kluyvera sp. initiated nucleation at -9 degrees C. This strain, therefore, is a weak, also termed a Type III or Class C ice-nucleating active bacterium, but with no influence on the supercooling ability of individual snails. In summary, fluctuations in body water mass of hibernating snail populations, triggering changes in osmolyte concentration, rather than the presence of endogenous ice-nucleating-active bacteria, accounts for fluctuations in their T(c).

Cold hardiness abilities vary with the size of the land snail Cornu aspersum.

The land snail Cornu aspersum (syn. Helix aspersa) living in Brittany (France) can be considered partially freezing tolerant as it possesses a low ability to supercool and a limited capacity to bear freezing of its body tissues. The absence of a marked cold hardiness strategy permits the emphasis of the role of parameters such as individual size or water mass (W(M)) contained by the organism. Adult snails (shell diameter 30-32 mm) had a supercooling ability, about 1-1.5 degrees C lower than that of immatures (shell diameter 12-20 mm) and survived longer to an exposure to -5 degrees C, with an Lt(50) comprised between 6.0 and 9.8 h against 2.6 to 4.2 h for immature snails. This better ability to bear freezing was explained by the faster dynamic of body ice formation observed in small individuals, which attained ice lethal quantity more rapidly. At the species level, large snails will then tend to be more tolerant to freezing and small ones to be freezing avoidant, a statement also observable at the phylum level.

Effects of a freezing event during hibernation on further survival, reproduction and growth in the partially freezing tolerant land snail Helix aspersa muller (Gastropoda: helicidae).

Tolerance of ectothermic animals to freezing is often estimated by assessing survival a few days after the treatment. However, in the long term, ice formation in the body tissues can affect survival, as well as reproductive capability and growth. The land snail Helix aspersa survives only short durations with ice in its tissues, to a lethal limit of 40 to 60 % of its body water frozen. Adult and immature snails were treated during their winter dormancy period to a freezing event above this limit; their survival was observed both in the short and long term, as well as their ability to reproduce (adults) and grow (immature snails). Treated snails were compared with a control group, which was not frozen. No difference appeared in the survival, reproduction and growth of control and frozen snails. This study confirms partial freezing tolerance in this population of Helix aspersa.

The effect of antibiotic treatment on the supercooling ability of the land snail Helix aspersa (Gastropoda: Pulmonata).

The land snail Helix aspersa is a partially freezing tolerant species whose supercooling ability is limited to ca. -3 to -5 degrees C. One hundred adult snails were subjected to the following two experimental conditions: (i) a starved group, provided with water; (ii) an antibiotic-treated group that was provided with a solution containing a mixture of two antibiotics. The antibiotic group exhibited a T(c) significantly lower than the starved group (-3.94 +/- 1.32 degrees C, n = 40 and -3.07 +/- 0.99, n = 30, t test, p < 0.005). This study showed that bacteria of the gut are likely to elevate animal supercooling points. It is also the first report in which a possible ice-nucleating activity of the gut microflora in a land snail has been suggested by the action of antibiotics on the T(c).