Stay in shape: Assessing the adaptive potential of shell morphology and its sensitivity to temperature in the invasive New Zealand mud snail Potamopyrgus antipodarum through phenotypic plasticity and natural selection in Europe.

Climate change may force organisms to adapt genetically or plastically to new environmental conditions. Invasive species show remarkable potential for rapid adaptation. The ovoviviparous New Zealand mud snail (NZMS), Potamopyrgus antipodarum, has successfully established across Europe with two clonally reproducing mitochondrial lineages since its arrival in the first half of the 19th century. Its remarkable variation in shell morphology was shown to be fitness relevant. We investigated the effects of temperature on shell morphology across 11 populations from Germany and the Iberian Peninsula in a common garden across three temperatures. We analyzed size and shape using geometric morphometrics. For both, we compared reaction norms and estimated heritabilities. For size, the interaction of temperature and haplotype explained about 50% of the total variance. We also observed more genotype by environment interactions indicating a higher degree of population differentiation than in shape. Across the three temperatures, size followed the expectations of the temperature-size rule, with individuals growing larger in cold environments. Changes in shape may have compensated for changes in size affecting space for brooding embryos. Heritability estimates were relatively high. As indicated by the very low coefficients of variation for clonal repeatability (CV A ), they can probably not be compared in absolute terms. However, they showed some sensitivity to temperature, in haplotype t more so than in z, which was only found in Portugal. The low CV A values indicate that genetic variation among European populations is still restricted with a low potential to react to selection. A considerable fraction of the genetic variation was due to differences between the clonal lineages. The NZMS has apparently not been long enough in Europe to accumulate significant genetic variation relevant for morphological adaptation. As temperature is obviously not the sole factor influencing shell morphology, their interaction will probably not be a factor limiting population persistence under a warming climate in Europe.

Synthesis and Polyelectrolyte Functionalization of Hollow Fiber Membranes Formed by Solvent Transfer Induced Phase Separation.

Ultrafiltration membranes are important porous materials to produce freshwater in an increasingly water-scarce world. A recent approach to generate porous membranes is solvent transfer induced phase separation (STrIPS). During STrIPS, the interplay of liquid-liquid phase separation and nanoparticle self-assembly results in hollow fibers with small surface pores, ideal structures for applications as filtration membranes. However, the underlying mechanisms of the membrane formation are still poorly understood, limiting the control over structure and properties. To address this knowledge gap, we study the nonequilibrium dynamics of hollow fiber structure evolution. Confocal microscopy reveals the distribution of nanoparticles and monomers during STrIPS. Diffusion simulations are combined with measurements of the interfacial elasticity to investigate the effect of the solvent concentration on nanoparticle stabilization. Furthermore, we demonstrate the separation performance of the membrane during ultrafiltration. To this end, polyelectrolyte multilayers are deposited on the membrane, leading to tunable pores that enable the removal of dextran molecules of different molecular weights (>360 kDa, >60 kDa, >18 kDa) from a feed water stream. The resulting understanding of STrIPS and the simplicity of the synthesis process open avenues to design novel membranes for advanced separation applications.

Nanostructured, Fluid-Bicontinuous Gels for Continuous-Flow Liquid-Liquid Extraction.

Fluid-bicontinuous gels are unique materials that allow two distinct fluids to interact through a percolating, rigid scaffold. Current restrictions for their use are the large fluid-channel sizes (>5 µm), limiting the fluid-fluid interaction surface-area, and the inability to flow liquids through the channels. In this work a scalable synthesis route of nanoparticle stabilized fluid-bicontinuous gels with channels sizes below 500 nm and specific surface areas of 2 m2 cm-3 is introduced. Moreover, it is demonstrated that liquids can be pumped through the fluid-bicontinuous gels via electroosmosis. The fast liquid flow in the fluid-bicontinuous gel facilitates their use for molecular separations in continuous-flow liquid-liquid extraction. Together with the high surface areas, liquid flow through fluid-bicontinuous gels enhances their potential as highly permeable porous materials with possible uses as microreaction media, fuel-cell components, and separation membranes.

Centrifugal Assembly of Helical Bijel Fibers for pH Responsive Composite Hydrogels.

In microfluidics, centrifugal forces are important for centrifugal microfluidic chips and curved microchannels. Here, an unrecognized use of the centrifugal effect in microfluidics is introduced. The assembly of helical soft matter fibers in a rotating microcapillary is investigated. During assembly, the fibers undergo phase separation, generating particle stabilized bicontinuous interfacially jammed emulsions gels. This process is accompanied by a transition of the fiber density over time. As a result, the direction of the centrifugal force in the rotating microcapillary changes. The authors analyze this effect systematically with high-speed video microscopy and complementary computer simulations. The resulting understanding enables the control of the helical fiber assembly into microropes. These microropes can be converted into pH responsive hydrogels that swell and shrink with potential applications in tissue engineering, soft robotics, controlled release, and sensing. More generally, the knowledge gained from this work shows that centrifugal forces potentially enable directed self-assembly or separation of colloids, biological cells, and emulsions in microfluidics.

Inference of DNA methylation patterns in molluscs.

Mollusca are the second largest and arguably most diverse phylum of the animal kingdom. This is in sharp contrast to our very limited knowledge concerning epigenetic mechanisms including DNA methylation in this invertebrate group. Here, we inferred DNA methylation patterns by analysing the normalized dinucleotide CG content in protein-coding sequences and identified DNA methyltransferases (DNMT1 and 3) in published transcriptomes and genomes of 140 species across all eight classes of molluscs. Given the evolutionary age and morphological diversity of molluscs, we expected to find evidence for diverse methylation patterns. Our inferences suggest that molluscs possess substantial levels of DNA methylation in gene bodies as a rule. Yet, we found deviations from this general picture with regard to (i) the CpG observed/expected distributions indicating a reduction in DNA methylation in certain groups and (ii) the completeness of the DNMT toolkit. Reductions were evident in Caudofoveata, Solenogastres, Polyplacophora, Monoplacophora, as well as Scaphopoda. Heterobranchia and Oegopsida were remarkable as they lacked DNMT3, usually responsible for de novo methylation, yet showed signs of DNA methylation. Our survey may serve as guidance for direct empirical analyses of DNA methylation in molluscs. This article is part of the Theo Murphy meeting issue 'Molluscan genomics: broad insights and future directions for a neglected phylum'.

Stabilizing liquid drops in nonequilibrium shapes by the interfacial crosslinking of nanoparticles.

Droplets are spherical due to the principle of interfacial energy minimization. Here, we show that nonequilibrium droplet shapes can be stabilized via the interfacial self-assembly and crosslinking of nanoparticles. This principle allows for the stability of practically infinitely long liquid tubules and monodisperse cylindrical droplets. Droplets of oil-in-water are elongated via gravitational or hydrodynamic forces at a reduced interfacial tension. Silica nanoparticles self-assemble and cross-link on the interface triggered by the synergistic surface modification with hexyltrimethylammonium- and trivalent lanthanum-cations. The droplet length dependence is described by a scaling relationship and the rate of nanoparticle deposition on the droplets is estimated. Our approach potentially enables the 3D-printing of Newtonian Fluids, broadening the array of material options for additive manufacturing techniques.

A layover in Europe: Reconstructing the invasion route of asexual lineages of a New Zealand snail to North America.

Non-native invasive species are threatening ecosystems and biodiversity worldwide. High genetic variation is thought to be a critical factor for invasion success. Accordingly, the global invasion of a few clonal lineages of the gastropod Potamopyrgus antipodarum is thus both puzzling and has the potential to help illuminate why some invasions succeed while others fail. Here, we used SNP markers and a geographically broad sampling scheme (N = 1617) including native New Zealand populations and invasive North American and European populations to provide the first widescale population genetic assessment of the relationships between and among native and invasive P. antipodarum. We used a combination of traditional and Bayesian molecular analyses to demonstrate that New Zealand populations harbour very high diversity relative to the invasive populations and are the source of the two main European genetic lineages. One of these two European lineages was in turn the source of at least one of the two main North American genetic clusters of invasive P. antipodarum, located in Lake Ontario. The other widespread North American group had a more complex origin that included the other European lineage and two New Zealand clusters. Altogether, our analyses suggest that just a small handful of clonal lineages of P. antipodarum were responsible for invasion across continents. Our findings provide critical information for prevention of additional invasions and control of existing invasive populations and are of broader relevance towards understanding the establishment and evolution of asexual populations and the forces driving biological invasion.

Controlling Surfactant Adsorption on Highly Charged Nanoparticles to Stabilize Bijels.

Bicontinuous particle-stabilized emulsions (bijels) are networks of interpenetrating oil/water channels with applications in catalysis, tissue engineering, and energy storage. Bijels can be generated by arresting solvent transfer induced phase separation (STrIPS) via interfacial jamming of nanoparticles. However, until now, STrIPS bijels have only been formed with silica nanoparticles of low surface charge densities, limiting their potential applications in catalysis and fluid transport. Here, we show how strongly charged silica nanoparticles can stabilize bijels. To this end, we carry out a systematic study employing dynamic light scattering, zeta potential, acid/base titrations, turbidimetry, surface tension, and confocal microscopy. We find that moderating the adsorption of oppositely charged surfactants on the particles is crucial to facilitate particle dispersibility in the bijel casting mixture and bijel stabilization. Our results potentially introduce a general understanding for bijel fabrication with different inorganic nanoparticle materials of variable charge density.

Subspecies in the Sarus Crane Antigone antigone revisited; with particular reference to the Australian population.

Subspecies are often less well-defined than species but have become one of the basic units for legal protection. Evidence for the erection or synonymy of subspecies therefore needs to be founded on the best science available. Here we show that there is clear genetic disjunction in the Sarus Crane Antigone antigone, where previously the variation had appeared to be clinal. Based on a total sample of 76 individuals, analysis of 10 microsatellite loci from 67 samples and 49 sequences from the mitochondrial control region, this research establishes that the Australian Sarus Crane A. a. gillae differs significantly from both A. a. antigone (South Asia) and A. a. sharpii (Myanmar and Indochina). A single sample from the extinct Philippine subspecies A. a luzonica clustered with A. a. gillae, hinting at the potential for a more recent separation between them than from A. a. antigone and A. a. sharpii, even though A. a. sharpii is closer geographically. The results demonstrate that failure to detect subspecies through initial genetic profiling does not mean discontinuities are absent and has significance for other cases where subspecies are dismissed based on partial genetic evidence. It could also be potentially important for sourcing birds for reintroduction to the Philippines.

Designing bijels formed by solvent transfer induced phase separation with functional nanoparticles.

Bicontinuous interfacially jammed emulsion gels (bijels) formed via solvent transfer induced phase separation (STrIPS) are new soft materials with potential applications in separations, healthcare, or catalysis. To facilitate their applications, means to fabricate STrIPS bijels with nanoparticles of various surface chemistries are needed. Here, we investigate the formation of STrIPS bijels with nanoparticles of different wettabilities, ranging from partially hydrophobic to extremely hydrophilic. To this end, the surface wettability of silica nanoparticles is tailored by functionalization with ligands bearing either hydrophobic or hydrophilic terminal groups. We show that partially hydrophobic particles with acrylate groups can impart short-term stability to STrIPS bijels on their own. However, to enable long-term stability, the use of cationic surfactants is needed. Partially hydrophobic particles require short chain surfactants for morphological stability while glycerol-functionalized hydrophilic particles require double chain cationic surfactants. Variation of the surfactant concentration results in various STrIPS bijel morphologies with controllable domain sizes. Last, we show that functional groups on the nanoparticles facilitate interfacial cross-linking for the purposes of reinforcing STrIPS bijels. Our research lays the foundation for the use of a wide variety of solid particles, irrespective of their surface wettabilities, to fabricate bijels with potential applications in Pickering interfacial catalysis and as cross-flow microreactors.

Bicontinuous Interfacially Jammed Emulsion Gels (bijels) as Media for Enabling Enzymatic Reactive Separation of a Highly Water Insoluble Substrate.

Although enzymes are efficient catalysts capable of converting various substrates into desired products with high specificity under mild conditions, their effectiveness as catalysts is substantially reduced when substrates are poorly water-soluble. In this study, to expedite the enzymatic conversion of a hydrophobic substrate, we use a bicontinuous interfacially jammed emulsion gel (bijel) which provides large interfacial area between two immiscible liquids: oil and water. Using lipase-catalyzed hydrolysis of tributyrin as a model reaction in a batch mode, we show that bijels can be used as media to enable enzymatic reaction. The bijel system gives a four-fold increase in the initial reaction rate in comparison to a stirred biphasic medium. Our results demonstrate that bijels are powerful biphasic reaction media to accelerate enzymatic reactions with various hydrophobic reagents. This work also demonstrates that bijels can potentially be used as reaction media to enable continuous reactive separations.

Templating Interfacial Nanoparticle Assemblies via in Situ Techniques.

In situ surface modification of nanoparticles has a rich industrial history, but in recent years, it has also received increased attention in the field of directed self-assembly. In situ techniques rely on components within a Pickering emulsion system, such as amphiphiles that act as hydrophobizers or ionic species that screen charges, to drive the interfacial assembly of particles. Instead of stepwise procedures to chemically tune the particle wettability, in situ methods use elements already present within the system to alter the nanoparticle interfacial behavior, often depending on Coulombic interactions to simplify operations. The surface modifications are not contingent on specific chemical reactions, which further enables a multitude of possible nanoparticles to be used within a given system. In recent studies, in situ methods have been combined with external means of shaping the interface to produce materials with high interfacial areas and complex geometries. These systems have facilely tunable properties, enabling their use in an extensive array of applications. In this feature article, in honor of the late Prof. Helmuth Möhwald, we review how in situ techniques have influenced the development of soft, advanced materials, covering the fundamental interfacial phenomena with an outlook on materials science.

Testing the adaptive value of gastropod shell morphology to flow: a multidisciplinary approach based on morphometrics, computational fluid dynamics and a flow tank experiment.

A major question in stream ecology is how invertebrates cope with flow. In aquatic gastropods, typically, larger and more globular shells with larger apertures are found in lotic (flowing water) versus lentic (stagnant water) habitats. This has been hypothetically linked to a larger foot, and thus attachment area, which has been suggested to be an adaptation against risk of dislodgement by current. Empirical evidence for this is scarce. Furthermore, these previous studies did not discuss the unavoidable increase in drag forces experienced by the snails as a consequence of the increased cross sectional area. Here, using Potamopyrgus antipodarum as a study model, we integrated computational fluid dynamics simulations and a flow tank experiment with living snails to test whether 1) globular shell morphs are an adaptation against dislodgement through lift rather than drag forces, and 2) dislocation velocity is positively linked to foot size, and that the latter can be predicted by shell morphology. The drag forces experienced by the shells were always stronger compared to the lift and lateral forces. Drag and lift forces increased with shell height but not with globularity. Rotating the shells out of the flow direction increased the drag forces, but decreased lift. Our hypothesis that the controversial presence of globular shells in lotic environments could be explained by an adaptation against lift rather than drag forces was rejected. The foot size was only predicted by the size of the shell, not by shell shape or aperture size, showing that the assumed aperture/foot area correlation should be used with caution and cannot be generalized for all aquatic gastropod species. Finally, shell morphology and foot size were not related to the dislodgement speed in our flow tank experiment. We conclude that other traits must play a major role in decreasing dislodgement risk in stream gastropods, e.g., specific behaviours or pedal mucus stickiness. Although we did not find globular shells to be adaptations for reducing dislodgement risk, we cannot rule out that they are still flow-related adaptations. For instance, globular shells are more crush-resistant and therefore perhaps adaptive in terms of diminishing damage caused by tumbling after dislodgement or against lotic crush-type predators.

Robust Bijels for Reactive Separation via Silica-Reinforced Nanoparticle Layers.

Pickering emulsions have been successfully used as media for catalysis and separation. However, simultaneous reaction and separation cannot be performed in a continuous mode in these systems, because reagents cannot be readily loaded into or recovered from the dispersed phase. Bicontinuous interfacially jammed emulsion gels (bijels), in which the oil and water phases are continuous throughout the structure, have potential as media for simultaneous reaction and separation in a continuous mode. In this work, we take a major step toward realizing this vision by demonstrating the ability of bijels to be used in reactive separation performed in a batch fashion. To perform effectively, bijels must maintain their morphology and interfacial mass transfer properties during reaction. To strengthen the bijels, we modify the solvent transfer-induced phase separation (STRIPS) method to make bijels resistant to mechanical stresses and prevent detachment of nanoparticles from the oil/water interface due to pH changes by chemically fusing the interfacial nanoparticles. The reinforced bijel is successfully tested in base-catalyzed hydrolysis of esters and remains robust under these challenging conditions.

Shaping nanoparticle fingerprints at the interface of cholesteric droplets.

The ordering of nanoparticles into predetermined configurations is of importance to the design of advanced technologies. Here, we balance the interfacial energy of nanoparticles against the elastic energy of cholesteric liquid crystals to dynamically shape nanoparticle assemblies at a fluid interface. By adjusting the concentration of surfactant that plays the dual role of tuning the degree of nanoparticle hydrophobicity and altering the molecular anchoring of liquid crystals, we pattern nanoparticles at the interface of cholesteric liquid crystal emulsions. In this system, interfacial assembly is tempered by elastic patterns that arise from the geometric frustration of confined cholesterics. Patterns are tunable by varying both surfactant and chiral dopant concentrations. Adjusting the particle hydrophobicity more finely by regulating the surfactant concentration and solution pH further modifies the rigidity of assemblies, giving rise to surprising assembly dynamics dictated by the underlying elasticity of the cholesteric. Because particle assembly occurs at the interface with the desired structures exposed to the surrounding water solution, we demonstrate that particles can be readily cross-linked and manipulated, forming structures that retain their shape under external perturbations. This study serves as a foundation for better understanding inter-nanoparticle interactions at interfaces by tempering their assembly with elasticity and for creating materials with chemical heterogeneity and linear, periodic structures, essential for optical and energy applications.

Adaptive phenotypic plasticity in a clonal invader.

Organisms featuring wide trait variability and occurring in a wide range of habitats, such as the ovoviviparous New Zealand freshwater snail Potamopyrgus antipodarum, are ideal models to study adaptation. Since the mid-19th century, P. antipodarum, characterized by extremely variable shell morphology, has successfully invaded aquatic areas on four continents. Because these obligately and wholly asexual invasive populations harbor low genetic diversity compared to mixed sexual/asexual populations in the native range, we hypothesized that (1) this phenotypic variation in the invasive range might be adaptive with respect to colonization of novel habitats, and (2) that at least some of the variation might be caused by phenotypic plasticity. We surveyed 425 snails from 21 localities across northwest Europe to attempt to disentangle genetic and environmental effects on shell morphology. We analyzed brood size as proxy for fitness and shell geometric morphometrics, while controlling for genetic background. Our survey revealed 10 SNP genotypes nested into two mtDNA haplotypes and indicated that mainly lineage drove variation in shell shape but not size. Physicochemical parameters affected both shell shape and size and the interaction of these traits with brood size. In particular, stronger stream flow rates were associated with larger shells. Our measurements of brood size suggested that relatively larger slender snails with relatively large apertures were better adapted to strong flow than counterparts with broader shells and relatively small apertures. In conclusion, the apparent potential to modify shell morphology plays likely a key role in the invasive success of P. antipodarum; the two main components of shell morphology, namely shape and size, being differentially controlled, the former mainly genetically and the latter predominantly by phenotypic plasticity.

Variable Molecular Markers for the Order Mantophasmatodea (Insecta).

The recently discovered insect order Mantophasmatodea currently comprises 19 Southern African species. These mainly occur in allopatry, have high levels of color polymorphism and communicate via species- and gender-specific vibratory signals. High levels of interspecific morphological conservatism mean that cryptic species are likely to be uncovered. These aspects of Mantophasmatodean biology make them an ideal group in which to investigate population divergence due to habitat-specific adaptation, sexual selection, and potentially sensory speciation. Lack of appropriate genetic markers has thus far rendered such studies unfeasible. To address this need, the first microsatellite loci for this order were developed. Fifty polymorphic loci were designed specifically for Karoophasma biedouwense (Austrophasmatidae), out of which 23 were labeled and tested for amplification across the order using 2-3 individuals from 10 species, representing all 4 currently known families. A Bayesian mitochondrially encoded cytochrome c oxidase I (COI) topology was reconstructed and divergence dates within the order were estimated for the first time. Amplification success and levels of polymorphism were compared with genetic divergence and time since divergence. In agreement with studies on vertebrate taxa, both amplification and variability were negatively correlated with distance (temporal and genetic). The high number of informative loci will offer sufficient resolution for both broad level population genetic analysis and individual based pedigree or parentage analyses for most species in Austrophasmatidae, with at least some loci available for the other families. This resource will facilitate research into the evolutionary biology of this understudied but fascinating group.

One-Step Generation of Salt-Responsive Polyelectrolyte Microcapsules via Surfactant-Organized Nanoscale Interfacial Complexation in Emulsions (SO NICE).

Polyelectrolyte microcapsules are versatile compartments for encapsulation, protection, and controlled/triggered release of active agents. Conventional methods of polyelectrolyte microcapsule preparation require multiple steps or do not allow for efficient encapsulation of active agents in the lumen of the microcapsule. In this work, we present the fabrication of hollow polyelectrolyte microcapsules with a salt-responsive property based on surfactant organized nanoscale interfacial complexation in emulsions (SO NICE). In SO NICE, polyelectrolyte microcapsules are templated by water-in-oil-in-water (W/O/W) double emulsions. One polyelectrolyte is dissolved in the inner water droplet of the W/O/W double emulsions, whereas the second polyelectrolyte is dissolved in the organic phase by hydrophobic ion paring with an oppositely charged hydrophobic surfactant. Interfacial complexation of the two polyelectrolytes generates a few hundred-nanometer thick film at the inner water-oil interface of the W/O/W double emulsions. SO NICE microcapsules can be triggered to release their cargo by increasing the ionic strength of the solution, which is a hallmark of polyelectrolyte-based microcapsules. By enabling dissolution and interfacial complexation of polyelectrolytes in organic solvents, SO NICE widens the pallet of polymers that can be used to generate functional polyelectrolyte microcapsules with high encapsulation efficiency for applications in encapsulation and controlled/triggered release.

Multifunctional nanocomposite hollow fiber membranes by solvent transfer induced phase separation.

The decoration of porous membranes with a dense layer of nanoparticles imparts useful functionality and can enhance membrane separation and anti-fouling properties. However, manufacturing of nanoparticle-coated membranes requires multiple steps and tedious processing. Here, we introduce a facile single-step method in which bicontinuous interfacially jammed emulsions are used to form nanoparticle-functionalized hollow fiber membranes. The resulting nanocomposite membranes prepared via solvent transfer-induced phase separation and photopolymerization have exceptionally high nanoparticle loadings (up to 50 wt% silica nanoparticles) and feature densely packed nanoparticles uniformly distributed over the entire membrane surfaces. These structurally well-defined, asymmetric membranes facilitate control over membrane flux and selectivity, enable the formation of stimuli responsive hydrogel nanocomposite membranes, and can be easily modified to introduce antifouling features. This approach forms a foundation for the formation of advanced nanocomposite membranes comprising diverse building blocks with potential applications in water treatment, industrial separations and as catalytic membrane reactors.

Snails in the desert: Species diversification of Theba (Gastropoda: Helicidae) along the Atlantic coast of NW Africa.

The spatial subdivision of species often plays a pivotal role in speciation. Across their entire range, species are rarely panmictic and crucial consequences of spatial subdivision are (1) random genetic drift including historical factors, (2) uniform selection, and (3) divergent selection. Each of these consequences may result in geographic variation and eventually reproductive isolation, but their relative importance in speciation is still unclear. In this study, we used a combination of genetic, morphological, and climatic data to obtain a comprehensive picture of differentiation among three closely related, parapatrically distributed taxa of the land snail genus Theba occurring along the Atlantic coasts of South Morocco and Western Sahara. We conducted Mantel and partial Mantel tests to relate phenotypic and genotypic variation of these species to geography and/or climate. As null hypothesis for an evolutionary scenario, we assumed nonadaptive speciation and expected a pattern of isolation by distance among taxa. Rejection of the null hypothesis would indicate isolation by environment due to adaptation. Generally, genetic drift plays an important role but is rarely considered as sole driver of speciation. It is the combination of drift and selection that predominantly drives speciation. This study, however, provides a potential example, in which nonadaptive speciation, that is, genetic drift, is apparently the main driver of shaping the diversity of Theba in NW Africa. Restriction of gene flow between populations caused by geographic isolation probably has played an important role. Climate oscillations during the Plio- and Pleistocene may have led to repeated ecological changes in NW Africa and disruptions of habitats promoting differentiation by geographic isolation. The inferred evolutionary scenario, however, did not fully explain the incongruence between the AFLP- and mtDNA-tree topologies. This incongruence might indicate past hybridization among the studied Theba forms.