More than one way to smell ashore - Evolution of the olfactory pathway in terrestrial malacostracan crustaceans.

Crustaceans provide a fascinating opportunity for studying adaptations to a terrestrial lifestyle because within this group, the conquest of land has occurred at least ten times convergently. The evolutionary transition from water to land demands various morphological and physiological adaptations of tissues and organs including the sensory and nervous system. In this review, we aim to compare the brain architecture between selected terrestrial and closely related marine representatives of the crustacean taxa Amphipoda, Isopoda, Brachyura, and Anomala with an emphasis on the elements of the olfactory pathway including receptor molecules. Our comparison of neuroanatomical structures between terrestrial members and their close aquatic relatives suggests that during the convergent evolution of terrestrial life-styles, the elements of the olfactory pathway were subject to different morphological transformations. In terrestrial anomalans (Coenobitidae), the elements of the primary olfactory pathway (antennules and olfactory lobes) are in general considerably enlarged whereas they are smaller in terrestrial brachyurans compared to their aquatic relatives. Studies on the repertoire of receptor molecules in Coenobitidae do not point to specific terrestrial adaptations but suggest that perireceptor events - processes in the receptor environment before the stimuli bind - may play an important role for aerial olfaction in this group. In terrestrial members of amphipods (Amphipoda: Talitridae) as well as of isopods (Isopoda: Oniscidea), however, the antennules and olfactory sensilla (aesthetascs) are largely reduced and miniaturized. Consequently, their primary olfactory processing centers are suggested to have been lost during the evolution of a life on land. Nevertheless, in terrestrial Peracarida, the (second) antennae as well as their associated tritocerebral processing structures are presumed to compensate for this loss or rather considerable reduction of the (deutocerebral) primary olfactory pathway. We conclude that after the evolutionary transition from water to land, it is not trivial for arthropods to establish aerial olfaction. If we consider insects as an ingroup of Crustacea, then the Coenobitidae and Insecta may be seen as the most successful crustacean representatives in this respect.

Friction reduction mechanism of the cuticle surface in the sandhopper talitrus saltator (Amphipoda, talitridae).

In many cases, strong friction reduction is critical for success of both living organisms and engineering systems. Some arthropods exhibit good antifriction abilities in their specific living environments and have inspired many innovations for solving industry challenges. However, the current literature mainly focused on terrestrial insects, such as beetles, grasshoppers and katydids. The antifriction mechanisms in amphibious arthropods are still unknown, even if their surfaces are optimized for both air and water environments. Herein the tribological properties of the cuticle surface of the sandhopper Talitrus saltator were studied using a universal microtribometer. Further investigations were developed to identify the microstructural, compositional, wettability, and mechanical properties of the sandhopper shell cuticles. It was found that increasing normal force can significantly reduce the coefficient of friction of the shell cuticle, especially for the alive and rewet sandhopper shells. The shell consists of bottle-like nano-caves in its exocuticle, nano-tubes in its mesocuticle, and gauze-like multilayers in its endocuticle. Under physiological conditions, glycoprotein-like fluid fillings exist in both the bottle-like caves and the nano-tubes below and cover on the shell surface. More importantly, a new antifriction mechanism of lubricant-squeezing nano-porous system was established for the sandhopper shell. This work can deepen our understanding in antifriction surfaces of amphibiotic crustaceans, and provide a potential approach to resolve the friction challenge in micro-machines, especially for the applications under aqueous condition. STATEMENT OF SIGNIFICANCE: Friction regulation is one of the critical mechanisms for animal locomotion in natural environments. However, not much is known about the mechanism of amphibious arthropods to reduce friction between their body and diverse environments, particularly achieving adaption under both air and aqueous conditions. We quantitatively study the microstructural, compositional and mechanical properties of the sandhopper (Talitrus saltator) shell cuticle and tribological behaviors under different conditions. Our results reveal the nano-porous system with fluid fillings for the sandhopper's shell and demonstrate the potential antifriction mechanism of this amphibious animal. We anticipate this work will inspire some effective antifriction designs for micro-machines, especially for their applications in complex environment like human body.

Comparative analysis of the antennae of three amphipod species with different lifestyles.

Crustaceans detect chemical stimuli in the environment with aesthetasc sensilla, which are located on their 1st antennae. With the transition to other environments, chemoreception faces physical challenges. To provide a deeper understanding of the relation between the morphology of olfactory organs and different lifestyles, we studied the peripheral olfactory system of three amphipod species, the marine Gammarus salinus, the blind subterranean freshwater species Niphargus puteanus, and the terrestrial Cryptorchestia garbinii. We compared the 1st and 2nd antennae of these species with respect to length and presence of aesthetascs and other sensilla. The females of N. puteanus reveal the longest 1st antennae in relation to body size. G. salinus shows the largest aesthetascs and the same relative length of the 1st antennae as male N. puteanus. C. garbinii has very short 1st antennae and reduced (putative) aesthetascs. Our findings show that the compensation of vision loss by olfaction cannot be generally assumed in animals from dark environments. Furthermore, the behaviour of C. garbinii indicates a chemosensory ability, despite the reduction of the 1st antennae. A comparison with other terrestrial crustaceans suggests that the loss of the olfactory sense on the 1st antennae in C. garbinii might be compensated with chemoreception by the 2nd antennae.

An aluminum shield enables the amphipod Hirondellea gigas to inhabit deep-sea environments.

The amphipod Hirondellea gigas inhabits the deepest regions of the oceans in extreme high-pressure conditions. However, the mechanisms by which this amphipod adapts to its high-pressure environment remain unknown. In this study, we investigated the elemental content of the exoskeleton of H. gigas specimens captured from the deepest points of the Mariana Trench. The H. gigas exoskeleton contained aluminum, as well as a major amount of calcium carbonate. Unlike other (accumulated) metals, aluminum was distributed on the surface of the exoskeleton. To investigate how H. gigas obtains aluminum, we conducted a metabolome analysis and found that gluconic acid/gluconolactone was capable of extracting metals from the sediment under the habitat conditions of H. gigas. The extracted aluminum ions are transformed into the gel state of aluminum hydroxide in alkaline seawater, and this gel covers the body to protect the amphipod. This aluminum gel is a good material for adaptation to such high-pressure environments.

Nanostructures and Monolayers of Spheres Reduce Surface Reflections in Hyperiid Amphipods.

Transparent zooplankton and nekton are often nearly invisible when viewed under ambient light in the pelagic zone [1-3]. However, in this environment, where the light field is directional (and thus likely to cause reflections), and under the bioluminescent searchlights of potential predators, animals may be revealed by reflections from their body surface [4-7]. We investigated the cuticle surfaces of seven species of hyperiids (Crustacea; Amphipoda) using scanning electron microscopy and found two undocumented features that may reduce reflectance. We found that the legs of Cystisoma spp. (n = 5) are covered with an ordered array of nanoprotuberances 200 ± 20 nm SD in height that function optically as a gradient refractive index material [6, 8, 9]. Additionally, we observed that Cystisoma and six other species of hyperiids are covered with a monolayer of homogenous spheres (diameters ranging from 52 ± 7 nm SD on Cystisoma spp. to 320 ± 15 nm SD on Phronima spp.). Optical modeling using effective medium theory and transfer matrix methods demonstrated that both the nanoprotuberances and the monolayers reduce reflectance by as much as 100-fold, depending on the wavelength and angle of the incident light and the thickness of the gradient layer. Even though we only consider surface reflectance and not internal light scattering, our study demonstrates that these nanoprotuberances and spheres can improve crypsis in a featureless habitat where the smallest reflection can render an animal vulnerable to visual predation.

A unique apposition compound eye in the mesopelagic hyperiid amphipod Paraphronima gracilis.

The mesopelagic habitat is a vast space that lacks physical landmarks and is structured by depth, light penetration, and horizontal currents. Solar illumination is visible in the upper 1,000 m of the ocean, becoming dimmer and spectrally filtered with depth-generating a nearly monochromatic blue light field. The struggle to perceive dim downwelling light and bioluminescent sources and the need to remain unseen generate contrasting selective pressures on the eyes of mesopelagic inhabitants. Hyperiid amphipods are cosmopolitan members of the mesopelagic fauna with at least ten different eye configurations across the family-ranging from absent eyes in deep-living species to four enlarged eyes in mesopelagic individuals. The hyperiid amphipod Paraphronima gracilis has a pair of bi-lobed apposition compound eyes, each with a large upward-looking portion and a small lateral-looking portion. The most unusual feature of the P. gracilis eye is that its upward-looking portion is resolved into a discontinuous retina with 12 distinct groups, each serving one transverse row of continuously spaced facets. On the basis of eye morphology, we estimated spatial acuity (2.5° ± 0.11°, SEM; n = 25) and optical sensitivity (30 ± 3.4 µm(2) ⋅ sr, SEM; n = 25). Microspectrophotometry showed that spectral sensitivity of the eye peaked at 516 nm (±3.9 nm, SEM; n = 6), significantly offset from the peak of downwelling irradiance in the mesopelagic realm (480 nm). Modeling of spatial summation within the linear retinal groups showed that it boosts sensitivity with less cost to spatial acuity than more typical configurations.

A 'NanoSuit' surface shield successfully protects organisms in high vacuum: observations on living organisms in an FE-SEM.

Although extremely useful for a wide range of investigations, the field emission scanning electron microscope (FE-SEM) has not allowed researchers to observe living organisms. However, we have recently reported that a simple surface modification consisting of a thin extra layer, termed 'NanoSuit', can keep organisms alive in the high vacuum (10(-5) to 10(-7) Pa) of the SEM. This paper further explores the protective properties of the NanoSuit surface-shield. We found that a NanoSuit formed with the optimum concentration of Tween 20 faithfully preserves the integrity of an organism's surface without interfering with SEM imaging. We also found that electrostatic charging was absent as long as the organisms were alive, even if they had not been coated with electrically conducting materials. This result suggests that living organisms possess their own electrical conductors and/or rely on certain properties of the surface to inhibit charging. The NanoSuit seems to prolong the charge-free condition and increase survival time under vacuum. These findings should encourage the development of more sophisticated observation methods for studying living organisms in an FE-SEM.

Intertidal caprellids (Crustacea: Amphipoda) of the Persian Gulf and the Gulf of Oman, with description of three new species.

In the present contribution, intertidal caprellids of the Persian Gulf and the Gulf of Oman were studied. Materials were collected, from 40 localities, with a variety of habitats, along the Iranian coasts. Three new species are described including; Metaprotella macoranicus sp. nov, Monoliropus kazemii sp. nov, Pseudaeginella hormozensis sp. nov and Pseudocaprel-linapambanensis is reported from studied area. Metaprotella macoranicus sp. nov., is characterized with clear suture between head and pereonite 1, second peduncle article larger than third ones in first antenna, tuft and three long setae in abdominal appendages and reduced mid-dorsal projections in pereonite 3. Monoliropus kazemii sp. nov., possesses biarticulate abdominal appendage, straight second male gnathopod and setal formula with two apical and two medial setae. The main diagnostic character of Pseudaeginella hormozensis sp. nov., is reduced dorsal projection, bearing mid-lateral projection on pereonite 2-5 and robust pereopod 5 with grasping spines on propodus.

A novel marine silk.

The discovery of a novel silk production system in a marine amphipod provides insights into the wider potential of natural silks. The tube-building corophioid amphipod Crassicorophium bonellii produces from its legs fibrous, adhesive underwater threads that combine barnacle cement biology with aspects of spider silk thread extrusion spinning. We characterised the filamentous silk as a mixture of mucopolysaccharides and protein deriving from glands representing two distinct types. The carbohydrate and protein silk secretion is dominated by complex ß-sheet structures and a high content of charged amino acid residues. The filamentous secretion product exits the gland through a pore near the tip of the secretory leg after having moved through a duct, which subdivides into several small ductules all terminating in a spindle-shaped chamber. This chamber communicates with the exterior and may be considered the silk reservoir and processing/mixing space, in which the silk is mechanically and potentially chemically altered and becomes fibrous. We assert that further study of this probably independently evolved, marine arthropod silk processing and secretion system can provide not only important insights into the more complex arachnid and insect silks but also into crustacean adhesion cements.

Optical diagnostic test of stress conditions of aquatic organisms.

Global climate change has become a dire reality and its impact is expected to rise dramatically in the near future. Combined with the day-to-day human activities the climatic changes heavily affect the environment. In particular, a global temperature increase accompanied by a number of anthropogenic chemicals falling within the freshwater ecosystem results in a dramatic enhancement of the overall stress for most aquatic organisms. This leads to a significant shift in the species inventory and potential breakdown of the water ecosystem with severe consequences for local economies and water supply. In order to understand and predict the influence of climatic changes on the physiological and biochemical processes that take place in living aquatic organisms we explore the application of optical spectroscopy for monitoring and quantitative assessment of antioxidant enzymes activity in benthic amphipods of Lake Baikal. We demonstrate that the changes of the enzymes activity in Baikal amphipods undergoing thermal and/or hypoxia stress can be observed and documented by UV and optical spectroscopy both in vivo and in vitro.

Occurrence of the ectoparasite Isocyamus deltobranchium (Amphipoda: Cyamidae) on cetaceans from Atlantic waters.

The cyamids collected from a True's beaked whale (Mesoplodon mirus), a common dolphin (Delphinus delphis), and a harbour porpoise (Phocoena phocoena) stranded in Galicia (NW Spain) were identified as Isocyamus deltobranchium, extending the range of this species to the Atlantic Ocean. The three cetacean species examined are new hosts for this parasite. The microscopic examination of female specimens reveals that certain important taxonomic characters can be present or absent depending on the maturation stage, an aspect which should be considered in future studies describing cyamid species.

Crustacean-acanthocephalan interaction and host cell-mediated immunity: parasite encapsulation and melanization.

Host-parasite interactions of Pomphorhynchus laevis (Müller, 1776) in naturally infected amphipod, Echinogammarus stammeri (Karaman), from the Brenta River (northern Italy) are described. A fully developed acanthocephalan larva occupies a large portion of an amphipod's haemocoelic space; thus, the parasite frequently induces displacement of host digestive tract and other internal organs. However, no apparent damage to the host's internal structures was observed. Within the haemocoel of E. stammeri, each larva of P. laevis is surrounded with a membranous layer, formed by microvilli, which maintains intimate contact with the amphipod's internal organs and haemocytes. Three types of circulatory haemocytes were identified based upon their distinct appearance: hyaline cell, semi-granular cell and granular cell. Echinogammarus stammeri haemocytes surrounded acanthocephalan larvae and in some instances a partially and/or totally melanized P. laevis larva was noticed. Interestingly, no melanized larvae were found in E. stammeri parasitized with other acanthocephalans namely Echinorhynchus truttae (Schrank, 1788), Polymorphus minutus (Goeze, 1782) and Acanthocephalus clavula (Dujardin, 1845).

Cuticle differentiation in the embryo of the amphipod crustacean Parhyale hawaiensis.

The arthropod cuticle is a multilayered extracellular matrix produced by the epidermis during embryogenesis and moulting. Molecularly and histologically, cuticle differentiation has been extensively investigated in the embryo of the insect Drosophila melanogaster. To learn about the evolution of cuticle differentiation, we have studied the histology of cuticle differentiation during embryogenesis of the amphipod crustacean Parhyale hawaiensis, which had a common ancestor with Drosophila about 510 million years ago. The establishment of the layers of the Parhyale juvenile cuticle is largely governed by mechanisms observed in Drosophila, e.g. as in Drosophila, the synthesis and arrangement of chitin in the inner procuticle are separate processes. A major difference between the cuticle of Parhyale and Drosophila concerns the restructuring of the Parhyale dorsal epicuticle after deposition. In contrast to the uniform cuticle of the Drosophila larva, the Parhyale cuticle is subdivided into two regions, the ventral and the dorsal cuticles. Remarkably, the boundary between the ventral and dorsal cuticles is sharp suggesting active extracellular regionalisation. The present analysis of Parhyale cuticle differentiation should allow the characterisation of the cuticle-producing and -organising factors of Parhyale (by comparison with the branchiopod crustacean Daphnia pulex) in order to contribute to the elucidation of fundamental questions relevant to extracellular matrix organisation and differentiation.

Comparative electron microscopic study of the stomach of Orchestia cavimana and Arcitalitrus sylvaticus (crustacea: Amphipoda).

The functional morphology of stomachs of the European semiterrestrial amphipod Orchestia cavimana and of the Australian terrestrial species Arcitalitrus sylvaticus was studied by electron microscopy. The stomach of the two amphipod species is divided longitudinally into a spacious dorsal food channel and two ventral filtration channels. Additionally, a prominent helically oriented circulation channel is situated on each lateral side of the stomach, forming a semicircular channel separated from the food channel by spines. The food channel conveys coarse food particles directly into the midgut through a funnel. The filtration channels receive fine material filtered through primary and secondary filters. Material forced through the secondary filters by the pressure of the laterally located inferolateralia eventually reaches the openings of the midgut glands. Washing of filters and soaking of ingested food items with enzymes probably is achieved by a forward stream of digestive juice from the midgut glands and conveyed through the circulatory channels. The specializations of the stomach of the two species of Amphipoda investigated are described and compared to the pertinent structures of Mysidacea and Isopoda.