Mollusk Shells Research Articles

Possible roles of Wnt in the shell growth of the pond snail Lymnaea stagnalis

Although the mechanisms of molluscan shell growth have been studied using mathematical models, little is known about the molecular basis underpinning shell morphogenesis. Here, we performed Wnt activation experiments to elucidate the potential roles of Wnt signaling in the shell growth of Lymnaea stagnalis. In general, we observed following three types of shell malformations in both dose- and developmental stage-dependent manners: (i) cap-shaped shell, (ii) cap-shaped shell with hydropic soft tissues, and (iii) compressed shell with a smaller number of coiling. We analyzed the morphologies of these malformed shells using the growing tube model, revealing that the compressed malformations show significantly larger values for T (torsion), with no significant changes in the values for the remaining parameters E (expansion) and C (curvature). We also found that cap-shaped malformations have significantly larger values for E, suggesting that the effects of BIO on shell formation may change during growth. Since the changes in T and/or E parameter values can greatly alter the shell morphologies from a planispiral or a cap-shaped one to various three-dimensional helices, changes in shell developmental processes possibly controlled by Wnt signaling may account for at least a part of the evolution of diverse shell forms in molluscs.

χ-Conotoxins are an evolutionary innovation of mollusk-hunting cone snails as a counter-adaptation to prey defense.

Mollusk-hunting (molluscivorous) cone snails belong to a monophyletic group in Conus, a genus of venomous marine snails. The molluscivorous lineage evolved from ancestral worm-hunting (vermivorous) snails ∼18 million years ago. To enable the shift to a molluscivorous lifestyle, molluscivorous cone snails must solve biological problems encountered when hunting other gastropods, namely: 1) preventing prey escape and 2) overcoming the formidable defense of the prey in the form of the molluscan shell, a problem unique to molluscivorous Conus. Here we show that χ-conotoxins, peptides exclusively expressed in the venoms of molluscivorous Conus, provide solutions to the above problems. Injecting χ-conotoxins into the gastropod mollusk Aplysia californica results in impaired locomotion and uncoordinated hyperactivity. Impaired locomotion impedes escape and a hyperactive snail will likely emerge from its shell, negating the protection the shell provides. Thus, χ-conotoxins are an evolutionary innovation that accompanied the emergence of molluscivory in Conus and provide solutions to problems posed by hunting other snails.

Tracing the origin of raw materials used for the production of ancient ceramics: a case study of multi-period archaeological sites in the Turopolje area (Continental Croatia)

The selection of raw materials for ancient pottery production was influenced by a variety of technological and cultural factors, underscoring the importance of characterising these materials to understand the technology of ancient societies. This research examines ancient ceramics from two multi-period archaeological sites: Staro Čiče-Gradišće (Neolithic, Copper Age, Copper/Early Bronze Age, Early Bronze Age, Late Bronze Age, Roman period, and Late Mediaeval period) and Kurilovec-Belinščica (Early Bronze Age and Middle/Late Bronze Age), alongside potential clayey raw materials collected near these settlements. Using a multi-analytical approach (optical microscopy, X-ray powder diffraction, mass and emission spectrometry, and laser granulometry), the research aims to determine the characteristics of the paste recipes (raw clay and tempers) and to examine the type and provenience of the raw materials used for ancient pottery production over different periods of the past (from the Neolithic to the Late Mediaeval period). The results showed that ancient potters preferred to use moderately plastic, sandy clay, while Bronze Age potters often used highly plastic clay. Potters utilised various non-plastic tempering materials, such as sands and gravels, grog, and mollusc shells, with their choice being influenced by the need to enhance technological properties as well as by regional and culturally determined pottery traditions. Most of the ceramics are of local origin, made from easily available raw materials that represent flood sediments of the nearby Sava River. However, non-local materials were detected in Neolithic samples, indicating the presence of exchange networks among those communities.

To date or not to date? A comparison of different 14C pretreatment methods applied to archeological marine shells from Vale Boi (Portugal)

Abstract Mollusk shells are often found in archeological sites, given their great preservation potential and high value as a multipurpose resource, and they can often be the only available materials useful for radiocarbon (14C) dating. However, dates obtained from shells are often regarded as less reliable compared to those from bones, wood, or charcoals due to different factors (e.g., Isotope fractionation, reservoir effects etc.). The standard acid etching pretreatment for mollusk shells is the most used in many 14C laboratories, although another protocol known as CarDS (Carbonate Density Separation) was introduced just over a decade ago. We compare these two methods with two newly proposed methods for intracellular organic matrix extraction. We applied all four methods to samples selected from different archeological layers of the well-known Upper Paleolithic site of Vale Boi, rich in mollusk specimens throughout the stratigraphic sequence. Here we compare our results to previous dates to determine which of these pretreatment methods results in the most reliable 14C dates. Based on the results of this study, all four methods gave inconsistent ages compared to previous dates and the stratigraphic attribution.

Analysis of fauna of mollusks, pollen assemblages and sedimentary environments in the Taganrog Bay of the Sea of Azov

The paper presents the results of a comprehensive study of well sections drilled on the coastal spits of the Taganrog Gulf of the Sea of Azov and bottom sediment columns selected in the water area. A detailed analysis of the early malacological communities, an important participant in the sedimentary process, made it possible to trace changes in their living conditions in the Holocene. Based on determining the absolute age (14C) from the valves of mollusk shells buried in situ, it is shown that at the beginning of the Phanagorian regression, in the area of the distal part of the Dolgaya Spit, there was a sea reservoir with an Azov-Black Sea complex of reference species. Based on the results of palynological analysis, changes in the natural conditions of the Azov region over the past few thousand years have been reconstructed and arid and humid climatic phases have been identified.

Nacre‐Mimetic Multi‐Mechanical Synergistic Ceramic Aerogels with Interfacial Bridging and Stress Delocalization

AbstractElastic ceramic aerogels are exceptionally suitable for thermal insulating applications, but lacking multi‐mechanical synergistic high strength remains a deficiency. Inspired by the robust protective shells of mollusks in nature, which feature distinct hierarchical ordered structures, exhibiting strong and tough mechanical properties, a system construction and structural governing strategy is proposed to allow the nanostructure to assemble strong and flexible ceramic aerogels. Through the design of a hierarchical multi‐arched structure configurated binary‐soldering‐assemble interfacial topology interlocking, concentrated stress is effectively discrete. This biomimetic strategy enables the effective dissipation of mechanical energy, resulting in ceramic aerogels with exemplary compressive resilience, bendability, tensile resistance, and their strength exceeding that of other nanofibrous aerogels by up to tenfold. Additionally, the flexible aerogels also show remarkable thermal resistance from −196 to 1100 °C and superior thermal insulation capabilities (39.72 mW m−1 K−1), making them more suitable for the applications.

Microbial mediated hardening of a Central Mediterranean upper shelf seafloor during the MIS 5.5; a possible post global warming scenario?

Higher sea level and sea surface temperature than at present have been inferred for the Last Interglacial (MIS 5.5, 135–116 ka), making it a good analogue for modeling the climate response of the environment to global warming in the near future. With the aim of predicting the possible evolution of some central Mediterranean shallow sandy seabeds, a MIS 5.5 biocalcarenite was investigated (Gulf of Taranto, Italy). Limited post-depositional diagenesis affected this deposit, which represents an infralittoral sandy seabed, with local vegetation and relatively high energy conditions. Medium-coarse sand sized bioclasts, with a negligible quantity of siliciclastic, compose the sediment that also hosts large shells of mollusks. Micritic cements are widespread, often in continuum with the micritized part of the shells, showing non-isopachous aphanitic and filamentous rims, aphanitic micro-mounds, vacuolar peloidal menisci, and aphanitic pore-filling matrix. All these cements consist of submicrometer anhedral or nanospheroidal crystals of low-Mg calcite, mixed with a smaller amount of irregular plate-like crystals of saponite. Micritic cements are also rich in mineralized filamentous, tubular, and subspherical bacterial bodies. This highlights the occurrence of an epilithic and endolithic microbial community forming a biofilm that stabilized the mobile sediment as consequence of the microbial mediated early cement precipitation. This led the synsedimentary hardening of some parts the mobile sandy substrate, the settlement of sessile taxa - such as Spondylus gaederopus, oysters, serpulids and barnacles, together with endofaunal organisms. Early micritic cementation is common in modern tropical climate, whereas is substantially absent in the modern Mediterranean. Consequently, its presence in the MIS 5.5 deposit confirms warmer sea water temperature compared to today (estimated at ca +2 °C) and suggests that similar hardening of mobile substrates may occur in the near future as a response to global warming.

Neuropeptides regulate shell growth in the Mediterranean mussel (Mytilus galloprovincialis)

In bivalves, which are molluscs enclosed in a biomineralized shell, a diversity of neuropeptide precursors has been described but their involvement in shell growth has been largely neglected. Here, using a symmetric marine bivalve, the Mediterranean mussel (Mytilus galloprovincialis), we uncover a role for the neuroendocrine system and neuropeptides in shell production. We demonstrate that the mantle is rich in neuropeptide precursors and that a complex network of neuropeptide-secreting fibres innervates the mantle edge a region highly involved in shell growth. We show that shell damage and shell repair significantly modify neuropeptide gene expression in the mantle edge and the nervous ganglia (cerebropleural ganglia, CPG). When the CPG nerve commissure was severed, shell production was impaired after shell damage, and modified neuropeptide gene expression, the spatial organization of nerve fibres in the ganglia and mantle and biomineralization enzyme activity in the mantle edge. Injection of CALCIa and CALCIIa peptides rescued the impaired shell repair phenotype providing further support for their role in biomineralization. We propose that the regulatory mechanisms identified are likely to be conserved across bivalves and other shelled molluscs since they all share a similar nervous system, a common mantle biomineralization toolbox, and shell structure.

Land snail Δ47 thermometry using cultured and European natural populations of Clausilia pumila, Succinella oblonga and Trochulus hispidus

Clumped isotope composition (Δ47) of aragonitic shells of terrestrial molluscs allow the quantitative reconstructions of continental paleotemperatures from loess-paleosol records of the last 2.5 million years, but it needs testing the method under controlled laboratory conditions and using natural populations. In this study the stable carbon, oxygen and clumped isotope compositions of cultured individuals and field-collected specimens of three species (Clausilia pumila, Succinella oblonga and Trochulus hispidus), widespread in Quaternary loess deposits in Europe, were measured in the Debrecen (ATOMKI) and ETH Zürich laboratories. Our data of cultured snails demonstrate that shells of the three species record Δ47 signatures close to equilibrium conditions with mostly positive (C. pumila), both positive/negative (T. hispidus), and mostly negative (S. oblonga) offsets on the order of 0.0158 ± 0.0105 ‰ (mean offset±1 SD) compared to the Anderson et al. (2021) unified calibration. Calcification temperatures (TΔ47) derived from C. pumila shells were found to be in excellent agreement with chamber temperatures of 12, 18 and 24 °C, especially in the ATOMKI laboratory (mean T offset: 2.1 ± 2.2 °C, 1 SD), while the other two species exhibited larger discrepancies on the order of 5–8 °C (mean offset). Single shell-based mean TΔ47 values of field-collected molluscs indicated growth temperatures spanning values from the frost-free period to summer season, or even above summer maxima. The sites of natural populations, characterized by different climate conditions, could be grouped using the aridity index (AI) with values of 0.96–1.06 representing mean temperature of the frost-free period (TFFP), and AI ranging either from 0.85 to 0.91 or 1.14 to 1.51 reflecting mean summer season or July temperatures (TJJA/TJ). Multi-shell TΔ47, obtained by homogenizing the material of 3–5 shells of C. pumila and T. hispidus, reflect TJJA in most cases. We observed a dependence of the shell oxygen isotope compositions of the three species on oceanity/continentality with more negative/positive values characteristic for more continental/oceanic sites, likely due to moisture source distance and depletion of rainfall in 18O towards the continental interior. In addition, deviation of body fluid δ18O values (derived from δ18Oshell and Δ47shell) from seasonal precipitation δ18O is also controlled by an additional temperature effect, being independent of the climate zone of the collection site, but co-varying with the site-specific difference between single shell TΔ47 and TJJA.

CALCIUM CHANGES IN Bulimulus tenuissimus (MOLLUSCA) EXPERIMENTALLY INFECTED WITH LARVAL Angiostrongylus cantonensis (NEMATODA)

Only in 2018 Bulimulus tenuissimus was incriminated as an intermediate host of Angiostrongylus cantonensis, the causative agent of infections in domestic, wild animals and humans, causing neural angiostrongyliasis or eosinophilic meningitis in humans. The present study aimed to infect the mollusk B. tenuissimus, using L1 larvae of A. cantonensis, under laboratory conditions, and to analyse the changes that occurred in the calcium metabolism of the host mollusk. The infection caused hypercalcification in the shell of B. tenuissimus infected with A. cantonensis, with a 242% increase in the concentration of CaCO3, at the end of the prepatent period, in the shells of infected mollusks and hypercalcemia, especially at the end of the third week of infection (+18.51%) in relation to the control group. The interference of the parasitic nematode in the calcium metabolism of the host mollusk was evidenced, with reductions in tissue deposits and elevation of calcemia and calcium content in the shell.

Palaeoenvironmental implications of Late Quaternary bioerosion traces in central Patagonia (Southern Atlantic, Argentina)

ABSTRACTBioerosion is a valuable tool for inferring palaeoenvironmental and palaeoclimatic changes over time and across different regions. However, studies of bioerosion traces are scarce in the Southern Hemisphere. Most ichnological studies within Argentina are concentrated in San Jorge Gulf (Patagonia, Argentina) and little is known about deposits located north of the Gulf. Here, we focus on bioerosion traces on Quaternary mollusc shells. Samples were collected from Quaternary marine deposits at the Bahía Vera–Cabo Raso sites in northern San Jorge Gulf. To resolve age discrepancies reported in the literature, we use amino acid racemization and radiocarbon dating to confirm the presence of beach ridge deposits from Marine Isotope Stage (MIS) 5 and MIS 1. Fourteen ichnotaxa are recorded in the study area. Additionally, distinct variations in the pattern of bioerosion across different ages are observed, indicating that environmental changes occurred in the northern San Jorge Gulf between the MIS 5 interglacial and the Holocene. This reinforces the hypothesis that there is an association between bioerosion, productivity and circulation in the Southern Atlantic Ocean.

Shellfishing, sea levels, and the earliest Native American villages (5000–3800 yrs. BP) of the South Atlantic Coast of the U.S

Shell ring archaeological sites are one of the most visible site types along the lower South Atlantic Coast of the United States. These cultural sites are large, circular to arcuate piles of mollusk shells with some reaching over three meters in elevation and over 100 m in diameter. They are comprised largely of mollusk shells (e.g., Eastern oyster, Crassostrea virginica), but also contain early pottery, nonhuman faunal remains, and other artifacts. Our work establishes that they represent the earliest widespread Native American villages occupied year-round in the Eastern Woodlands of North America. Significantly, our results from sea-level modelling and isotope geochemistry on mollusks establish that the inhabitants of these earliest villages (ca. 5000–3800 yrs. BP) lived within a fluctuating coastal environment, harvested certain resources year-round, and targeted diverse habitats across the estuaries. Both the growth and decline of these earliest villages are associated with a concomitant rise and lowering of sea level that impacted the productivity of the oyster reef fishery along the South Atlantic Coast. Despite these large-scale environmental changes, this research indicates that Native American fishing villages persisted along the coast for over 1000 years.

New Polymer Biocomposites Based on Biopoly(Ethylene Terephthalate) and Waste Mollusc Shells

Currently, scientific studies have are focusing on environmentally friendly solutions, such as the effective use of waste in new green polymeric materials according to circular economy. Waste valorization is the main driving force for upcoming academic research. In this study, the impact of mussel particle size on reinforced biopoly(terphtalate ethylene) (bPET) is investigated. The waste filler was modified using NaOH. The filler content was 10 wt% and the same for all samples. The strength properties of the materials were determined in static tensile, bending and impact tests. The wetting angle was also analyzed for the obtained biocomposites. A low-cycle dynamic test was carried out to determine changes in dissipation energy and to observe the development of relaxation processes. This present study proves that preparation of new biocomposites based on waste mussels is an effective option in waste management.

Shell-bearing Bivalvia and Gastropoda from a rocky shore in the Central Caribbean of Colombia

This study documents the natural accumulation of mollusk shells on an elevated rocky shore carved into the calcareous rocks of the La Popa formation at Punta Roca (Atlántico), covering an estimated area of 0.35 km2. Hydrodynamic transport and differential exposure to environmental conditions are the primary factors contributing to shell deposition. A total of 58 mollusk species were identified, including 30 bivalves and 28 gastropods. The Veneridae was the most prominent, with the highest number of species (7) and shells comprising 64.22 % of the total. Key species contributing to the accumulation include the bivalves Anomalocardia cf. flexuosa, Polymesoda cf. arctata, Leukoma pectorina, Tivela mactroides, Crassostrea rhizophorae, and the gastropod Vitta virginea. These species are primarily characteristic of estuarine environments, with the exception of T. mactroides. Notably, the first recorded occurrence of the non-native gastropod Naria turdus (Cypraeidae) on the Colombian Caribbean coast was documented. The accumulations exhibit varying degrees of abrasion, fragmentation, encrustation, and bioerosion, with some shells appearing almost intact, indicating differences in origin and transport mechanisms. The extraction of shells for ornamental purposes poses a significant challenge to this natural accumulation.

ХОЗЯЙСТВЕННАЯ АДАПТАЦИЯ НАСЕЛЕНИЯ ПАЛЕОМЕТАЛЛА НА ПАМЯТНИКЕ КЛЕРК-5 В ПРИМОРЬЕ

The article examines the events of the Early Palaeometal epoch, when the multidirectional migration of the farmers from the continental regions of Dongbei to the territory of Primorye took place. At the multi-layered site Klerk 5 in Boysman Bay a habitation horizon left by people about 3,200 years ago was examined. It included a hearth, two units of mollusk shell deposits, pottery, stone tools and a variety of ecofacts. Based on the analysis of the paleogeographical situation, the composition and location of artifacts and ecofacts, the authors reconstruct individual aspects of the life support system and lifestyle of thePaleometal epoch population.

Mechanical Characteristics of Bivalve (Mollusc) Shells from the East Coast of India: Implications and Their Significance on the Survival Strategy

Due to natural selection, an increase in biotic and abiotic factors develops an adaptive response in bivalves (molluscs) for survival. Morphological variation in a bivalve is strongly correlated with the changes in the structural and compositional properties of a shell. Therefore, the mechanical characteristics of a shell reflect its morpho-functional adaptation against predation and ecological stresses. An initial investigation into the primary mechanical attributes of predominant bivalve shells from the east coast of India, Tegillarca granosa, Meretrix meretrix, Sunetta meroe, Mactra turgida and Donax scortum, has been documented in this study. The quantitative assessment of the centreline thickness, Vickers hardness and density variation of the shells were measured and correlated along with the preliminary investigation of the microstructural patterns. The results from our study are in agreement with the most vulnerable region of a shell, located just below the umbonal zone, which has been verified through predatory boreholes in other studies. Our data shows a high correlation between shell hardness and density, which could also be the potential properties to consider in detail for future studies, apart from the shell thickness, which could provide a broader perspective on the stability of bivalve shells. Different microstructural patterns observed in the species could be phylogenetically driven based on their life modes.

Industrial manufacturing method and characterization of Ayurvedic marine drugs: mother pearl, cowry, coral and pearl

Introduction: Ayurvedic marine drugs derived from mollusc shells and coral are regularly used by Ayurvedic physicians to treat several disease conditions like acid peptic disease, irritable bowel syndrome, osteoporosis, etc. However, standard operating procedures for manufacturing these drugs and their complete characterization have not been published in the Ayurvedic Formulary and Ayurvedic Pharmacopeia of India to date. Methods: Present study describes the traditional manufacturing process and thorough characterization using classical and advanced analytical tools. The raw materials characters, in-process parameters, and finished product specifications have been elaborated to develop monographs. Especially, the identity and purity of raw coral and pearl were checked by Raman Spectroscopy and Energy Dispersive X-ray Fluorescence analysis. Results: In the finished product analysis, the X-Ray Diffraction study revealed that incineration after trituration with Aloe barbadensis leaf pulp or rose water converted the aragonite phase of calcium carbonate into calcite phase in mother pearl, cowry, and pearl while the calcite form of raw coral was retained. The prominent bands around 1390, 870, and 712 cm−1 detected by Fourier Transform-Infrared Spectroscopy and mass loss between 39–44% (w/w) revealed by thermogravimetric analysis confirmed the carbonate form of these calcium-based drugs. The finished products were very fine grayish-white powders constituted by irregularly shaped nano-micro particulate calcium carbonate exhibiting particle size between 600 nm (D10 value) to 1.2 µm (D90 value). Conclusion: The quality control and assurance achieved in this study may be further utilized by the pharmaceutical industries to manufacture quality marine drugs and conduct efficacy studies.

Application of Marine Mollusk Shells (Meretrix lusoria) as Low-Cost Biosorbent for Removing Cd2+ and Pb2+ Ions from Aqueous Solution: Kinetic and Equilibrium Study

The present work aims to evaluate the applicability of mollusk (Meretrix lusoria) shells as a biosorbent for toxic metal ions (Cd2+ and Pb2+) following the batch mode biosorption procedure. Some well-known analytical methods have been used to characterize the biosorbent such as a scanning electron microscope (SEM), an energy dispersive X-ray (EDX), Fourier transform infrared spectroscopy (FTIR), and X-ray diffraction. The mechanism of metal ion biosorption was examined using various analytical techniques. Therefore, an evaluation of operating factors such as contact time, pH, initial concentration of metal ions, biosorbent dose, and temperature was performed. The results obtained in this investigation indicated that the optimum conditions for the biosorption of Cd+2 and Pb+2 ions are as follows: pH = 6; contact times of 90 min; and the 20 mg/L of initial [M2+]. And a biosorbent dosage of 1.0 g/100 mL for each metal ion solution was also determined. The maximum removal efficiency results were 90.6% for Cd+2 and 91.5% for Pb+2 at pH 6.0. The biosorption isotherm was investigated using three forms of linear equilibrium (Freundlich, Langmuir, and Temkin models). Kinetic studies were also conducted to determine the equilibrium time for the biosorption of the studied metals utilizing the pseudo-second-order, pseudo-first-order, and intraparticle diffusion model. The data indicate that the biosorption kinetics of Cd2+ and Pb2+ follow the pseudo-second-order models. According to the present study, it can be identified that the shell of Meretrix lusoria is a suitable biosorbent for Cd2+ and Pb2+ ions and can contribute to their removal from environmentally polluted water.

Synthesis and Characterization of Nano-Crystallite Triple Super Phosphate (TSP) from Marine Mollusk Waste: Babylonia japonica, Oliva sayana, and Conasprella bermudensis.

The study aims to synthesize nano-crystallite TSP using renewable, low-cost, waste marine mollusk from three different species such as Babylonia japonica, Oliva sayana, and Conasprella bermudensis. The molar ratio of phosphate to calcium in triple superphosphate [TSP, Ca(H2PO4)2.H2O] significantly impacts its properties and fertilizer performance, in this case, we kept the ratio to 2. Raw TSP has a high phosphate content and lower calcium content. The synthesized TSP was analyzed using various techniques including TGA, XRD, EDX, FT-IR, and SEM. The study utilized multiple XRD model equations to analyze crystallite size ( ), with all models except the Liner straight-line method providing higher estimates for synthesized TSP. Furthermore, the values for stress (2×107 to 4×107 N/m2), strain (4×10-4 to 9×10-4), as well as energy density (4.54×103 to 16.27×103 J/m3) were also calculated for the synthesized product. However, the preferential growth calculation indicates that (010), (021), and (020) planes are the most thermodynamically stable planes for the growth of the synthesized TSP. Apart from that, FTIR result confirms that CaO, -OH, as well as PO4 3- functional groups are present in the synthesized products. This research suggests that marine mollusks can be utilized as a calcium precursor for P-fertilizer and 60 % phosphoric acid, thereby reducing production costs by eliminating additional dehydrating. Additionally, waste marine mollusk shells could be utilized as an alternative to the production of phosphate-based fertilizer.

Evolution of chitin-synthase in molluscs and their response to ocean acidification

Chitin-synthase (CHS) is found in most eukaryotes and has a complex evolutionary history. Research into CHS has mainly been in the context of biomineralization of mollusc shells an area of high interest due to the consequences of ocean acidification. Exploration of CHS at the genomic level in molluscs, the evolution of isoforms, their tissue distribution, and response to environmental challenges are largely unknown. Exploiting the extensive molecular resources for mollusc species it is revealed that bivalves possess the largest number of CHS genes (12–22) reported to date in eukaryotes. The evolutionary tree constructed at the class level of molluscs indicates four CHS Type II isoforms (A-D) probably existed in the most recent common ancestor, and Type II-A (Type II-A-1/Type II-A-2) and Type II-C (Type II-C-1/Type II-C-2) underwent further differentiation. Non-specific loss of CHS isoforms occurred at the class level, and in some Type II (B-D groups) isoforms the myosin head domain, which is associated with shell formation, was not preserved and highly species-specific tissue expression of CHS isoforms occurred. These observations strongly support the idea of CHS functional diversification with shell biomineralization being one of several important functions. Analysis of transcriptome data uncovered the species-specific potential of CHS isoforms in shell formation and a species-specific response to ocean acidification (OA). The impact of OA was not CHS isoform-dependent although in Mytilus, Type I-B and Type II-D gene expression was down-regulated in both M. galloprovincialis and M. coruscus. In summary, during CHS evolution the gene family expanded in bivalves generating a large diversity of isoforms with different structures and with a ubiquitous tissue distribution suggesting that chitin is involved in many biological functions. These findings provide insight into CHS evolution in molluscs and lay the foundation for research into their function and response to environmental changes.