Hierarchical looping results in extreme extensibility of silk fibre composites produced by Southern house spiders (Kukulcania hibernalis).

Spider silk is a tough and versatile biological material combining high tensile strength and extensibility through nanocomposite structure and its nonlinear elastic behaviour. Notably, spiders rarely use single silk fibres in isolation, but instead process them into more complex composites, such as silk fibre bundles, sheets and anchorages, involving a combination of spinneret, leg and body movements. While the material properties of single silk fibres have been extensively studied, the mechanical properties of silk composites and meta-structures are poorly understood and exhibit a hereto largely untapped potential for the bio-inspired design of novel fabrics with outstanding mechanical properties. In this study, we report on the tensile mechanics of the adhesive capture threads of the Southern house spider (Kukulcania hibernalis), which exhibit extreme extensibility, surpassing that of the viscid capture threads of orb weavers by up to tenfold. By combining high-resolution mechanical testing, microscopy and in silico experiments based on a hierarchical modified version of the Fibre Bundle Model, we demonstrate that extreme extensibility is based on a hierarchical loops-on-loops structure combining linear and coiled elements. The stepwise unravelling of the loops leads to the repeated fracture of the connected linear fibres, delaying terminal failure and enhancing energy absorption. This principle could be used to achieve tailored fabrics and materials that are able to sustain high deformation without failure.

Robustness of antiadhesion between nanofibers and surfaces covered with nanoripples of varying spatial period.

Since nanofibers have a high surface-to-volume ratio, van der Waals forces render them attracted to virtually any surface. The high ratio provides significant advantages for applications in drug delivery, wound healing, tissue regeneration, and filtration. Cribellate spiders integrate thousands of nanofibers into their capture threads as an adhesive to immobilize their prey. These spiders have antiadhesive nanoripples on the calamistrum, a comb-like structure on their hindmost legs, and are thus an ideal model for investigating how nanofiber adhesion can be reduced. We found that these nanoripples had similar spacing in the cribellate species Uloborus plumipes, Amaurobius similis, and Menneus superciliosus, independent of phylogenetic relation and size. Ripple spacing on other body parts (i.e., cuticle, claws, and spinnerets), however, was less homogeneous. To investigate whether a specific distance between the ripples determines antiadhesion, we fabricated nanorippled foils by nanosecond UV laser processing. We varied the spatial periods of the nanoripples in the range ~203-613 nm. Using two different pulse numbers resulted in ripples of different heights. The antiadhesion was measured for all surfaces, showing that the effect is robust against alterations across the whole range of spatial periods tested. Motivated by these results, we fabricated irregular surface nanoripples with spacing in the range ~130-480 nm, which showed the same antiadhesive behavior. The tested surfaces may be useful in tools for handling nanofibers such as spoolers for single nanofibers, conveyor belts for producing endless nanofiber nonwoven, and cylindrical tools for fabricating tubular nanofiber nonwoven. Engineered fibers such as carbon nanotubes represent a further candidate application area.

Physico-chemical properties of functionally adhesive spider silk nanofibres.

Currently, synthetic fibre production focuses primarily on high performance materials. For high performance fibrous materials, such as silks, this involves interpreting the structure-function relationship and downsizing to a smaller scale to then harness those properties within synthetic products. Spiders create an array of fibres that range in size from the micrometre to nanometre scale. At about 20 nm diameter spider cribellate silk, the smallest of these silks, is too small to contain any of the typical secondary protein structures of other spider silks, let alone a hierarchical skin-core-type structure. Here, we performed a multitude of investigations to elucidate the structure of cribellate spider silk. These confirmed our hypothesis that, unlike all other types of spider silk, it has a disordered molecular structure. Alanine and glycine, the two amino acids predominantly found in other spider silks, were much less abundant and did not form the usual α-helices and ß-sheet secondary structural arrangements. Correspondingly, we characterized the cribellate silk nanofibre to be very compliant. This characterization matches its function as a dry adhesive within the capture threads of cribellate spiders. Our results imply that at extremely small scales there may be a limit reached below which a silk will lose its structural, but not functional, integrity. Nano-sized fibres, such as cribellate silk, thus offer a new opportunity for inspiring the creation of novel scaled-down functional adhesives and nano meta-materials.

Laser-processed antiadhesive bionic combs for handling nanofibers inspired by nanostructures on the legs of cribellate spiders.

Nanofibers are drawing the attention of engineers and scientists because their large surface-to-volume ratio is favorable for applications in medicine, filter technology, textile industry, lithium-air batteries, and optical sensors. However, when transferring nanofibers to a technical product in the form of a random network of fibers, referred to as nonwoven fabric, the stickiness of the freshly produced and thus fragile nanofiber nonwoven remains a problem. This is mainly because nanofibers strongly adhere to any surface because of van der Waals forces. In nature, there are animals that are actually able to efficiently produce, process, and handle nanofibers, namely cribellate spiders. For that, the spiders use the calamistrum, a comb-like structure of modified setae on the metatarsus of the hindmost (fourth) legs, to which the 10-30 nm thick silk nanofibers do not stick due to a special fingerprint-like surface nanostructure. In this work, we present a theoretical model of the interaction of linear nanofibers with a sinusoidally corrugated surface. This model allows for a prediction of the adhesive interaction and, thus, the design of a suitable surface structure to prevent sticking of an artificially nonwoven of nanofibers. According to the theoretical prediction, a technical analogon of the nanoripples was produced by ultrashort pulse laser processing on different technically relevant metal surfaces in the form of so-called laser-induced periodic surface structures (LIPSS). Subsequently, by means of a newly established peel-off test, the adhesion of an electrospun polyamide fiber-based nonwoven was quantified on such LIPSS-covered aluminium alloy, steel, and titanium alloy samples, as well as on polished (flat) control samples as reference and, additionally, on samples with randomly rough surfaces. The latter revealed that the adhesion of electrospun nanofiber nonwoven is significantly lowered on the nanostructured surfaces compared with the polished surfaces.

Change of mechanical characteristics in spider silk capture threads after contact with prey.

Most spiders rely on specialized capture threads to subdue prey. Cribellate spiders use capture threads, whose adhesion is based on thousands of nanofibers instead of specialized glue. The nanofibers adhere due to van der Waals and hygroscopic forces, but the adhesion is strengthened by an interaction with the cuticular hydrocarbons (CHCs) covering almost all insects. The interaction between CHCs and cribellate threads becomes visible through migration of the CHCs into the thread even far beyond the point of contact. In this study, we were able to show that the migrated CHCs not only influence adhesion but also change the mechanical characteristics of the thread. While adhesion, extensibility and total energy decreased in threads treated with CHCs from different insects, we observed an increasing force required to break threads. Such mechanical changes could be beneficial for the spider: Upon the first impact of the insect in the web, it is important to absorb all the energy without breaking. Afterwards, a reduction in extensibility could cause the insect to stay closer to the web and thus become additionally entangled in neighboring threads. An increased tensile force would additionally ensure that for insects already in the web, it is even harder to free themselves. Taken together, all these changes make it unlikely that cribellate spiders reuse their capture threads, if not reacting rapidly and removing the prey insect before the CHCs can spread across the thread. STATEMENT OF SIGNIFICANCE: Cribellate spiders use capture threads that, unlike other spiders, consist of nanofibers and do not rely glue. Instead, prey adheres mainly because their surface compounds, so-called cuticular hydrocarbons (CHCs), interact with the thread, this way generating strong adhesion forces. Previous studies on biomechanics and adhesion of cribellate threads only dealt with artificial surfaces, neglecting any interaction with surface compounds. This study examines the dramatical mechanical changes of a cribellate thread after interaction with prey CHCs, showing modifications of the thread's extensibility, tensile force and total energy. Our results highlight the importance of studying mechanical properties of silk not only in an artificial context, but also in real life.

Insect cuticular hydrocarbon composition influences their interaction with spider capture threads.

Insects represent the main prey of spiders, and spiders and insects co-diversified in evolutionary history. One of the main features characterizing spiders is their web as a trap to capture prey. Phylogenetically, the cribellate thread is one of the earliest thread types that was specialized to capture prey. In contrast to other capture threads, it lacks adhesive glue and consists of nanofibres, which do not only adhere to insects via van der Waals forces but also interact with the insects' cuticular hydrocarbon (CHC) layer, thus enhancing adhesion. The CHC layer consists of multiple hydrocarbon types and is highly diverse between species. In this study, we show that CHC interaction with cribellate capture threads is affected by CHC composition of the insect. We studied the interaction in detail for four insect species with different CHC profiles and observed a differential migration of CHCs into the thread. The migration depends on the molecular structure of the hydrocarbon types as well as their viscosity, influenced by the ambient temperature during the interaction. As a consequence, adhesion forces to CHC layers differ depending on their chemical composition. Our results match predictions based on biophysical properties of hydrocarbons, and show that cribellate spiders can exert selection pressure on the CHC composition of their insect prey.

Ambient Climate Influences Anti-Adhesion between Biomimetic Structured Foil and Nanofibers.

Due to their uniquely high surface-to-volume ratio, nanofibers are a desired material for various technical applications. However, this surface-to-volume ratio also makes processing difficult as van der Waals forces cause nanofibers to adhere to virtually any surface. The cribellate spider Uloborus plumipes represents a biomimetic paragon for this problem: these spiders integrate thousands of nanofibers into their adhesive capture threads. A comb on their hindmost legs, termed calamistrum, enables the spiders to process the nanofibers without adhering to them. This anti-adhesion is due to a rippled nanotopography on the calamistrum. Via laser-induced periodic surface structures (LIPSS), these nanostructures can be recreated on artificial surfaces, mimicking the non-stickiness of the calamistrum. In order to advance the technical implementation of these biomimetic structured foils, we investigated how climatic conditions influence the anti-adhesive performance of our surfaces. Although anti-adhesion worked well at low and high humidity, technical implementations should nevertheless be air-conditioned to regulate temperature: we observed no pronounced anti-adhesive effect at temperatures above 30 °C. This alteration between anti-adhesion and adhesion could be deployed as a temperature-sensitive switch, allowing to swap between sticking and not sticking to nanofibers. This would make handling even easier.

Phylogenetic and morphological influence on habitat choice in moisture-harvesting horned lizards (Phrynosoma spp.).

In previous studies, the superhydrophilic skin of moisture-harvesting lizards has been linked to the morphological traits of the lizards' integument, that is, the occurrence of honeycomb-shaped microstructures. Interestingly, these structures can also cover the skin of lizards inhabiting wet habitats. We therefore tested the influence of the microstructures' main features on the habitat choice and wettability in the genus Phrynosoma. The genus Phrynosoma comprises moisture-harvesting species as well as nonspecialists. Lizards of this genus inhabit large areas of North America with diverse climatic conditions. Remarkably, the differences in the manifestation of microstructures are just as versatile as their surroundings. The phylogeny of the lizards as well as the depth of their ventral microstructures, though independent of each other, correlated with the precipitation in their respective habitat. All other morphological traits, as well as the skin's wettability itself, could not predict the habitat of Phrynosoma species. Hence, it is unlikely that the microstructure influences the wettability, at least directly. Hence, we presume an indirect influence for the following reasons: (a) As the ventral side cannot get wet by rain, but the belly could easily interact with a wet surface, the microstructure might facilitate water absorption from wet soil following precipitation. (b) We found the number of dorsal microstructures to be linked to the occurrence of silt in the habitat. In our study, we observed scales being heavily contaminated, most likely with a mixture of dead skin (after shedding) and silt. As many lizards burrow themselves or even shovel sand onto their backs, deploying the substrate might be a mechanism to increase the skin's wettability.

Robust substrate anchorages of silk lines with extensible nano-fibres.

Living systems are built of multiscale-composites: materials formed of components with different properties that are assembled in complex micro- and nano-structures. Such biological multiscale-composites often show outstanding physical properties that are unachieved by artificial materials. A major scientific goal is thus to understand the assembly processes and the relationship between structure and function in order to reproduce them in a new generation of biomimetic high-performance materials. Here, we tested how the assembly of spider silk nano-fibres (i.e. glue coated 0.5 µm thick fibres produced by so-called piriform glands) into different micro-structures correlates with mechanical performance by empirically and numerically exploring the mechanical behaviour of line anchors in an orb weaver, a hunting spider and two ancient web builders. We demonstrate that the anchors of orb weavers exhibit outstanding mechanical robustness with minimal material use by the indirect attachment of the silk line to the substrate through a soft domain ('bridge'). This principle can be used to design new artificial high-performance attachment systems.

Biocompatible Micron-Scale Silk Fibers Fabricated by Microfluidic Wet Spinning.

For successful material deployment in tissue engineering, the material itself, its mechanical properties, and the microscopic geometry of the product are of particular interest. While silk is a widely applied protein-based tissue engineering material with strong mechanical properties, the size and shape of artificially spun silk fibers are limited by existing processes. This study adjusts a microfluidic spinneret to manufacture micron-sized wet-spun fibers with three different materials enabling diverse geometries for tissue engineering applications. The spinneret is direct laser written (DLW) inside a microfluidic polydimethylsiloxane (PDMS) chip using two-photon lithography, applying a novel surface treatment that enables a tight print-channel sealing. Alginate, polyacrylonitrile, and silk fibers with diameters down to 1 µm are spun, while the spinneret geometry controls the shape of the silk fiber, and the spinning process tailors the mechanical property. Cell-cultivation experiments affirm bio-compatibility and showcase an interplay between the cell-sized fibers and cells. The presented spinning process pushes the boundaries of fiber fabrication toward smaller diameters and more complex shapes with increased surface-to-volume ratio and will substantially contribute to future tailored tissue engineering materials for healthcare applications.

Cribellate thread production as model for spider's spinneret kinematics.

Spider silk attracts researchers from the most diverse fields, such as material science or medicine. However, still little is known about silk aside from its molecular structure and material strength. Spiders produce many different silks and even join several silk types to one functional unit. In cribellate spiders, a complex multi-fibre system with up to six different silks affects the adherence to the prey. The assembly of these cribellate capture threads influences the mechanical properties as each fibre type absorbs forces specifically. For the interplay of fibres, spinnerets have to move spatially and come into contact with each other at specific points in time. However, spinneret kinematics are not well described though highly sophisticated movements are performed which are in no way inferior to the movements of other flexible appendages. We describe here the kinematics for the spinnerets involved in the cribellate spinning process of the grey house spider, Badumna longinqua, as an example of spinneret kinematics in general. With this information, we set a basis for understanding spinneret kinematics in other spinning processes of spiders and additionally provide inspiration for biomimetic multiple fibre spinning.

Adhesion of spider cribellate silk enhanced in high humidity by mechanical plasticization of the underlying fiber.

The disruptive nature of water presents a significant challenge when designing synthetic adhesives that maintain functionality in wet conditions. However, many animal adhesives can withstand high humidity or underwater conditions, and some are even enhanced by them. An understudied mechanism in such systems is the influence of material plasticization by water to induce adhesive work through deformation. Cribellate silk is a dry adhesive used by particular spiders to capture moving prey. It presents as a candidate for testing the water plasticization model as it can remain functional at high humidity despite lacking an aqueous component. We performed herein tensile and adhesion tests on cribellate threads from the spider, Hickmania troglodytes; a spider that lives within wet cave environments. We found that the work of adhesion of its cribellate threads increased as the axial fibre deformed during pull-off experiments. This effect was enhanced when the silk was wetted and as spider body size increased. Dry threads on the other hand were stiff with low adhesion. We rationalized our experiments by a series of scaling law models. We concluded that these cribellate threads operate best when the nanofibrils and axial fibers both contribute to adhesion. Design of future synthetic materials could draw inspiration from how water facilitates, rather than diminishes, cribellate silk adhesion.

Uncoiling springs promote mechanical functionality of spider cribellate silk.

Composites, both natural and synthetic, achieve novel functionality by combining two or more constituent materials. For example, the earliest adhesive silk in spider webs - cribellate silk - is composed of stiff axial fibers and coiled fibers surrounded by hundreds of sticky cribellate nanofibrils. Yet, little is known of how fiber types interact to enable capture of insect prey with cribellate silk. To understand the roles of each constituent fiber during prey capture, we compared the tensile performance of native-state and manipulated threads produced by the cribellate spider Psechrus clavis, and the adhesion of native threads along a smooth surface and hairy bee thorax. We found that the coiled fiber increases the work to fracture of the entire cribellate thread by up to 20-fold. We also found that the axial fiber breaks multiple times during deformation, an unexpected observation that indicates: (i) the axial fiber continues to contribute work even after breakage, and (ii) the cribellate nanofibrils may perform a previously unidentified role as a binder material that distributes forces throughout the thread. Work of adhesion increased on surfaces with more surface structures (hairy bee thorax) corresponding to increased deformation of the coiled fiber. Together, our observations highlight how the synergistic interactions among the constituents of this natural composite adhesive enhance functionality. These highly extensible threads may serve to expose additional cribellate nanofibrils to form attachment points with prey substrata while also immobilizing prey as they sink into the web due to gravity.

Small behavioral adaptations enable more effective prey capture by producing 3D-structured spider threads.

Spiders are known for producing specialized fibers. The radial orb-web, for example, contains tough silk used for the web frame and the capture spiral consists of elastic silk, able to stretch when prey impacts the web. In concert, silk proteins and web geometry affects the spider's ability to capture prey. Both factors have received considerable research attention, but next to no attention has been paid to the influence of fiber processing on web performance. Cribellate spiders produce a complex fiber alignment as their capture threads. With a temporally controlled spinneret movement, they connect different fibers at specific points to each other. One of the most complex capture threads is produced by the southern house spider, Kukulcania hibernalis (Filistatidae). In contrast to the so far characterized linear threads of other cribellate spiders, K. hibernalis spins capture threads in a zigzag pattern due to a slightly altered spinneret movement. The resulting more complex fiber alignment increased the thread's overall ability to restrain prey, probably by increasing the adhesion area as well as its extensibility. Kukulcania hibernalis' cribellate silk perfectly illustrates the impact of small behavioral differences on the thread assembly and, thus, of silk functionality.

Same Principles but Different Purposes: Passive Fluid Handling throughout the Animal Kingdom.

Everything on earth is subject to physical laws, thus they influence all facets of living creatures. Although these laws restrain animals in many ways, some animals have developed a way to use physical phenomena in their favor to conserve energy. Many animals, which have to handle fluids, for example, have evolved passive mechanisms by adapting their wettability or using capillary forces for rapid fluid spreading. In distinct animals, a similar selection pressure always favors a convergent development. However, when assessing the biological tasks of passive fluid handling mechanisms, their diversity is rather surprising. Besides the well-described handling of water to facilitate drinking in arid regions, observed in, e.g., several lizards, other animals like a special flat bug have developed a similar mechanism for a completely different task and fluid: Instead of water, these bugs passively transport an oily defense secretion to a region close to their head where it finally evaporates. And again some spiders use capillary forces to capture prey, by sucking in the viscous waxy cuticle of their prey with their nanofibrous threads. This review highlights the similarities and differences in the deployed mechanisms of passive fluid handling across the animal kingdom. Besides including well-studied animals to point out different mechanisms in general, we stretch over to not as extensively studied species for which similar mechanisms are described for different tasks. Thus, we provide an extensive overview of animals for which passive fluid handling is described so far as well as for future inspiration.

Mechanoreceptive sensillum fields at the tarsal tip of insect legs.

Groups of mechanoreceptive sensilla form small sensory fields on the ventral rim of the most distal tarsomeres in insects. Within these fields two or three sensilla are located closely together. Anterior and posterior fields are found in all three pairs of legs with only a few exceptions. The composition, exact location, and morphology of the fields were studied in representative species of several insect orders using light and scanning electron microscopy. There was no obvious correlation between field morphology and insect phylogenetic relationships.

The Calamistrum of the Feather-Legged Spider Uloborus plumipes Investigated by Focused Ion Beam and Scanning Electron Microscopy (FIB-SEM) Tomography.

Spiders are natural specialists in fiber processing. In particular, cribellate spiders manifest this ability as they produce a wool of nanofibers to capture prey. During its production they deploy a sophisticated movement of their spinnerets to darn in the fibers as well as a comb-like row of setae, termed calamistrum, on the metatarsus which plays a key role in nanofiber processing. In comparison to the elaborate nanofiber extraction and handling process by the spider's calamistrum, the human endeavors of spinning and handling of artificial nanofibers is still a primitive technical process. An implementation of biomimetics in spinning technology could lead to new materials and applications. Despite the general progress in related fields of nanoscience, the expected leap forward in spinning technology depends on a better understanding of the specific shapes and surfaces that control the forces at the nanoscale and that are involved in the mechanical processing of the nanofibers, respectively. In this study, the authors investigated the morphology of the calamistrum of the cribellate spider Uloborus plumipes. Focused ion beam and scanning electron microscopy tomography provided a good image contrast and the best trade-off between investigation volume and spatial resolution. A comprehensive three-dimensional model is presented and the putative role of the calamistrum in nanofiber processing is discussed.

Vibratory movements in contests between females of the feather-legged spider (Uloborus plumipes).

Females of the feather-legged spider Uloborus plumipes invade, and compete for, each other's orb webs. In the context of these competitive interactions the question arose how the spiders communicate. Since substrate-borne vibrations are the most important component of the sensory environment of web-building spiders, we investigated vibratory movements that might serve as signals of communication. Three behaviors were found to be associated with female-female contests and to cause propagating vibrations in the spider webs: thread pulling, abdominal trembling, and web shaking. While thread pulling and abdominal trembling were also observed when prey insects were caught in the webs, web shaking occurred only in response to the presence of a competing conspecific. Caused by flexing of the first legs and a vigorous rotary movement of the opisthosoma, web shaking creates a short burst of strong oscillations of the orb web. This behavior always elicited a behavioral reaction by the competitor and may serve as an intraspecific signal in the mutual assessment of competing spiders. We suggest that web shaking communicates resource holding potential in U. plumipes.

Adhesion enhancement of cribellate capture threads by epicuticular waxes of the insect prey sheds new light on spider web evolution.

To survive, web-building spiders rely on their capture threads to restrain prey. Many species use special adhesives for this task, and again the majority of those species cover their threads with viscoelastic glue droplets. Cribellate spiders, by contrast, use a wool of nanofibres as adhesive. Previous studies hypothesized that prey is restrained by van der Waals' forces and entrapment in the nanofibres. A large discrepancy when comparing the adhesive force on artificial surfaces versus prey implied that the real mechanism was still elusive. We observed that insect prey's epicuticular waxes infiltrate the wool of nanofibres, probably induced by capillary forces. The fibre-reinforced composite thus formed led to an adhesion between prey and thread eight times stronger than that between thread and wax-free surfaces. Thus, cribellate spiders employ the originally protective coating of their insect prey as a fatal component of their adhesive and the insect promotes its own capture. We suggest an evolutionary arms race with prey changing the properties of their cuticular waxes to escape the cribellate capture threads that eventually favoured spider threads with viscous glue.

Nanofibre production in spiders without electric charge.

Technical nanofibre production is linked to high voltage, because nanofibres are typically produced by electrospinning. In contrast, spiders have evolved a way to produce nanofibres without high voltage. These spiders are called cribellate spiders and produce nanofibres within their capture thread production. It is suggested that their nanofibres become frictionally charged when brushed over a continuous area on the calamistrum, a comb-like structure at the metatarsus of the fourth leg. Although there are indications that electrostatic charges are involved in the formation of the thread structure, final proof is missing. We proposed three requirements to validate this hypothesis: (1) the removal of any charge during or after thread production has an influence on the structure of the thread; (2) the characteristic structure of the thread can be regenerated by charging; and (3) the thread is attracted to or repelled from differently charged objects. None of these three requirements were proven true. Furthermore, mathematical calculations reveal that even at low charges, the calculated structural assembly of the thread does not match the observed reality. Electrostatic forces are therefore not involved in the production of cribellate capture threads.