Insights on long-term ecosystem changes from stable isotopes in historical squid beaks.

BACKGROUND: Assessing the historical dynamics of key food web components is crucial to understand how climate change impacts the structure of Arctic marine ecosystems. Most retrospective stable isotopic studies to date assessed potential ecosystem shifts in the Arctic using vertebrate top predators and filter-feeding invertebrates as proxies. However, due to long life histories and specific ecologies, ecosystem shifts are not always detectable when using these taxa. Moreover, there are currently no retrospective stable isotopic studies on various other ecological and taxonomic groups of Arctic biota. To test whether climate-driven shifts in marine ecosystems are reflected in the ecology of short-living mesopredators, ontogenetic changes in stable isotope signatures in chitinous hard body structures were analysed in two abundant squids (Gonatus fabricii and Todarodes sagittatus) from the low latitude Arctic and adjacent waters, collected between 1844 and 2023. RESULTS: We detected a temporal increase in diet and habitat-use generalism (= opportunistic choice rather than specialization), trophic position and niche width in G. fabricii from the low latitude Arctic waters. These shifts in trophic ecology matched with the Atlantification of the Arctic ecosystems, which includes increased generalization of food webs and higher primary production, and the influx of boreal species from the North Atlantic as a result of climate change. The Atlantification is especially marked since the late 1990s/early 2000s. The temporal patterns we found in G. fabricii's trophic ecology were largely unreported in previous Arctic retrospective isotopic ecology studies. Accordingly, T. sagittatus that occur nowadays in the high latitude North Atlantic have a more generalist diet than in the XIXth century. CONCLUSIONS: Our results suggest that abundant opportunistic mesopredators with short life cycles (such as squids) are good candidates for retrospective ecology studies in the marine ecosystems, and to identify ecosystem shifts driven by climate change. Enhanced generalization of Arctic food webs is reflected in increased diet generalism and niche width in squids, while increased abundance of boreal piscivorous fishes is reflected in squids' increased trophic position. These findings support opportunism and adaptability in squids, which renders them as potential winners of short-term shifts in Arctic ecosystems.

Stem-Cell-Regenerative and Protective Effects of Squid (Symplectoteuthis oualaniensis) Skin Collagen Peptides against H2O2-Induced Fibroblast Injury.

Recently, there has been a growing interest in collagen peptides derived from marine sources for their notable ability to protect skin cells against apoptosis induced by oxidants. Therefore, the current study aimed to investigate the fundamental properties of collagen peptides, including their physicochemical, thermal, structural, stem-cell-regenerative, and skin-cell-protective effects, in comparison to commercial collagen peptides. The acid-soluble (ASC) and pepsin-soluble (PSC) collagens exhibited three distinct bands on SDS-PAGE, namely α (α1 and α2), ß, and γ chains, confirming a type I pattern. The thermal profiles obtained from TG and DSC analyses confirmed the denaturation of PSC and ASC at temperatures ranging from 51.94 to 56.4 °C and from 52.07 to 56.53 °C, respectively. The purified collagen peptides were analyzed using SDS-PAGE and MALDI-TOF mass spectrometry, revealing a mass range of 900-15,000 Da. Furthermore, the de novo peptide sequence analysis confirmed the presence of the Gly-X-Y repeating sequence in collagen peptides. Collagen peptide treatments significantly enhanced HFF-1 cell proliferation and migration compared to the control group. ELISA results confirmed the potential interactions between collagen peptides and HFF-1 cells through α2ß1, α10ß1, and α11ß1 integrin receptors. Notably, collagen peptide treatment effectively restored the proliferation of HFF-1 cells damaged by H2O2. Consequently, the advantageous characteristics of squid skin collagen peptides highlight their promising role in regenerative medicine.

Peracetic acid treatment of squid eggs infected with parasitic copepod (Ikanecator primus gen. et sp. nov.).

Having been successfully bred in semi-intensive and intensive aquaculture systems, oval squids of the Sepioteuthis lessoniana species complex are emerging as promising candidates for research and industry. Nevertheless, information about pathogens and diseases that may affect squid aquaculture remains sparse. In this study, we identify new parasitic copepod species that causes squid mortality and decreases squid hatching rates, and we also offer a solution to eliminate the pathogen during incubation of squid eggs. The newly discovered copepod Ikanecator primus gen. et sp. nov. was identified on oval squid eggs for the first time using both morphological and molecular diagnostic markers. In the genomes of the copepod and associated microbiome, we identified multiple genes for enzymes involved in cephalopod eggshell degradation in genomes of the copepod and associated microbiome. Furthermore, we conducted experiments to assess efficacy of peracetic acid in inhibiting the I. primus gen. et sp. nov. both in vitro and in vivo using immersion treatment. We established that a 2-min exposure to a concentration of 250 µl/L of peracetic acid containing product (PAA-product; 35 mg/L PAA and 15 mg/L H2O2) inhibited the development of nauplii in vitro. All parasites exposed to a concentration of 500 µl/L of PAA-product (70 mg/L PAA and 30 mg/L H2O2) were eliminated within two minutes. On top of this, the immersion treatment with 500 µl/L of PAA-product (70 mg/L PAA and 30 mg/L H2O2) improved survival of squid embryos and increased size of squid hatchlings compared with control and the immersion treatment with 125 µl/L of PAA-product (17.5 mg/L PAA and 7.5 mg/L H2O2) and the immersion treatment with 250 µl/L of PAA-product (35 mg/L PAA and 15 mg/L H2O2). These findings suggest that PAA holds a great potential as inhibitor and controller of parasitic copepod infections and for overall health management in cephalopod culture.

Transcriptomic responses in the nervous system and correlated behavioural changes of a cephalopod exposed to ocean acidification.

BACKGROUND: The nervous system is central to coordinating behavioural responses to environmental change, likely including ocean acidification (OA). However, a clear understanding of neurobiological responses to OA is lacking, especially for marine invertebrates. RESULTS: We evaluated the transcriptomic response of the central nervous system (CNS) and eyes of the two-toned pygmy squid (Idiosepius pygmaeus) to OA conditions, using a de novo transcriptome assembly created with long read PacBio ISO-sequencing data. We then correlated patterns of gene expression with CO2 treatment levels and OA-affected behaviours in the same individuals. OA induced transcriptomic responses within the nervous system related to various different types of neurotransmission, neuroplasticity, immune function and oxidative stress. These molecular changes may contribute to OA-induced behavioural changes, as suggested by correlations among gene expression profiles, CO2 treatment and OA-affected behaviours. CONCLUSIONS: This study provides the first molecular insights into the neurobiological effects of OA on a cephalopod and correlates molecular changes with whole animal behavioural responses, helping to bridge the gaps in our knowledge between environmental change and animal responses.

Lipidomics profile and volatile compounds of squids (Illex argentinus, Ommastrephes Bartram and Dosidicus gigas) in the Argentine, North Pacific Ocean, Equator and Peru by UPLC-triple TOF-MS and HS-SPME-GC-O-MS.

Comprehensive lipid and volatile compound analyses were performed with squids collected from four varied geographical locations to discriminate the regional characteristics. A total of 1442 lipid molecules and 110 volatiles were detected in the squid muscle samples. There were significant differences in the lipid profiles between Argentine squid (Illex argentinus, AGT), North Pacific Ocean squid (Ommastrephes Bartram, NPO), Equatorial squid (Dosidicus gigas, EQ), and Peruvian squid (Dosidicus gigas, PR) muscle. Phosphatidylcholines (14.64%), triacylglycerols (12.42%), and ceramides (10.97%) were the main lipid components. The contents of polyunsaturated fatty acid in phospholipids and in glycerolipids were 30.35-52.05% and 18.11-25.15%, respectively. The volatiles in squids exhibited significant regional variation; 1-pentanol and 1-octanol, 2-ethyl-1-hexanol and terpinen-4-ol, 2,7-ethyl-1-hexanol, 3-methy-1-butanol and 2-propyl-1-pentanol were identified as characteristic flavor compounds in AGT, NPO, EQ, and PR, respectively. Sphingomyelin, phosphatidylserine, phosphatidylethanolamine, and ceramide were strongly correlated with volatiles in squid muscle. Our study is a reference for the lipid nutritional value and flavor compounds of squids.

Isolation, characterization, and application of bacteriophage on Vibrio parahaemolyticus biofilm to control seafood contamination.

OBJECTIVE: This study intended to isolate a Vibrio-particular phage from the natural environment, analyse its characteristics and genome sequence, and investigate its reduction effect on V. parahaemolyticus biofilm as a biocontrol agent in squid and mackerel. METHODS: Among 21 phages, phage CAU_VPP01, isolated from beach mud, was chosen for further experiments based on host range and EOP tests. When examining the reduction effect of phage CAU_VPP01 against Vibrio parahaemolyticus biofilms on surfaces (stainless steel [SS] and polyethylene terephthalate [PET]) and food surfaces (squid and mackerel). RESULTS: The phage showed the most excellent reduction effect at a multiplicity-of-infection (MOI) 10. Three-dimensional images acquired with confocal laser scanning microscopy (CLSM) analysis were quantified using COMSTAT, which showed that biomass, average thickness, and roughness coefficient decreased when treated with the phage. Colour and texture analysis confirmed that the quality of squid and mackerel was maintained after the phage treatment. Finally, a comparison of gene expression levels determined by qRT-PCR analysis showed that the phage treatment induced a decrease in the gene expression of flaA, vp0962, andluxS, as examples. CONCLUSION: This study indicated that Vibrio-specific phage CAU_VPP01 effectively controlled V. parahaemolyticus biofilms under various conditions and confirmed that the isolated phage could possibly be used as an effective biocontrol weapon in the seafood manufacturing industry.

Corrected and republished from: "Vibrio fischeri Possesses Xds and Dns Nucleases That Differentially Influence Phosphate Scavenging, Aggregation, Competence, and Symbiotic Colonization of Squid".

Cells of Vibrio fischeri colonize the light organ of Euprymna scolopes, providing the squid bioluminescence in exchange for nutrients and protection. The bacteria encounter DNA-rich mucus throughout their transition to a symbiotic lifestyle, leading us to hypothesize a role for nuclease activity in the colonization process. In support of this, we detected abundant extracellular nuclease activity in growing cells of V. fischeri. To discover the gene(s) responsible for this activity, we screened a V. fischeri transposon mutant library for nuclease-deficient strains. Interestingly, only one strain, whose transposon insertion mapped to nuclease gene VF_1451, showed a complete loss of nuclease activity in our screens. A database search revealed that VF_1451 is homologous to the nuclease-encoding gene xds in Vibrio cholerae. However, V. fischeri strains lacking xds eventually revealed slight nuclease activity on plates upon prolonged incubation. This led us to hypothesize that a second secreted nuclease, identified through a database search as VF_0437, a homolog of V. cholerae dns, might be responsible for the residual nuclease activity. Here, we show that Xds and/or Dns are involved in essential aspects of V. fischeri biology, including natural transformation, aggregation, and phosphate scavenging. Furthermore, strains lacking either nuclease were outcompeted by the wild type for squid colonization. Understanding the specific role of nuclease activity in the squid colonization process represents an intriguing area of future research.IMPORTANCEFrom soil and water to host-associated secretions such as mucus, environments that bacteria inhabit are awash in DNA. Extracellular DNA (eDNA) is a nutritious resource that microbes dedicate significant energy to exploit. Calcium binds eDNA to promote cell-cell aggregation and horizontal gene transfer. eDNA hydrolysis impacts the construction of and dispersal from biofilms. Strategies in which pathogens use nucleases to avoid phagocytosis or disseminate by degrading host secretions are well-documented; significantly less is known about nucleases in mutualistic associations. This study describes the role of nucleases in the mutualism between Vibrio fischeri and its squid host Euprymna scolopes. We find that nuclease activity is an important determinant of colonization in V. fischeri, broadening our understanding of how microbes establish and maintain beneficial associations.

Males conditionally inseminate at three female body locations according to female mating history and female maturity status in a squid.

In some squids, such as those in the family Loliginidae, upon copulation, females receive and store male-delivered sperm capsules, spermatangia, at two different body locations: the buccal membrane and the distal end of the oviduct. This insemination site dimorphism is associated with alternative reproductive strategies. However, in Loliolus sumatrensis, a species of Loliginidae, the females possess three insemination sites: buccal membrane (BM), basal left IV arm (ARM) and lateral head behind the left eye (EYE), therefore we studied such the unusual phenomena. We developed microsatellite markers and genotyped the paternity of each spermatangium on three sites. We found multiple paternity at every single site and simultaneous usage of all three sites by a few males. The seasonal dynamics of a population in the Seto Inland Sea revealed a set priority for the initial use of insemination sites as BM, followed by ARM and then EYE, whereas the maximum number of stored spermatangia was greater in EYE > ARM > BM. Female maturity status was correlated with the usage pattern of insemination sites but not with the number of stored spermatangia at any insemination site. These results suggest that a male squid inseminates at different locations according to female mating history and female maturity status.

The first chromosomal-level genome assembly and annotation of white suckerfish Remora albescens.

Remora albescens, also known as white suckerfish, recognized for its distinctive suction-cup attachment behavior and medicinal significance. In this study, we produced a high-quality chromosome-level genome assembly of R. albescens through the integration of 23.87 Gb PacBio long reads, 64.54 Gb T7 short reads, and 88.63 Gb Hi-C data. Initially, we constructed a contig-level genome assembly totaling 605.30 Mb with a contig N50 of 23.12 Mb. Subsequently, employing Hi-C technology, approximately 99.68% (603.38 Mb) of the contig-level genome was successfully assigned to 23 pseudo-chromosomes. Through the integration of homologous-based predictions, ab initio predictions, and RNA-sequencing methods, we successfully identified a comprehensive set of 22,445 protein-coding genes. Notably, 96.36% (21,629 genes) of these were effectively annotated with functional information. The genome assembly achieved an estimated completeness of 98.1% according to BUSCO analysis. This work promotes the applicability of the R. albescens genome, laying a solid foundation for future investigations into genomics, biology, and medicinal importance within this species.

Exploiting specific properties of squid pens for the preparation of oligochitosan hydrochloride.

Herein, we describe in first the application of squid pens for the preparation of pharmaceutical-grade oligochitosan hydrochloride with the physicochemical characteristics corresponding with the requirements of the European Pharmacopoeia. It is shown that the use of specific properties of squid pens as a source of parent chitosan allows preparing the product with a high yield at relatively moderate process conditions used for squid pens treatments and chitosan depolymerization.

Lighting the way: how the Vibrio fischeri model microbe reveals the complexity of Earth's "simplest" life forms.

Vibrio (Aliivibrio) fischeri's initial rise to fame derived from its alluring production of blue-green light. Subsequent studies to probe the mechanisms underlying this bioluminescence helped the field discover the phenomenon now known as quorum sensing. Orthologs of quorum-sensing regulators (i.e., LuxR and LuxI) originally identified in V. fischeri were subsequently uncovered in a plethora of bacterial species, and analogous pathways were found in yet others. Over the past three decades, the study of this microbe has greatly expanded to probe the unique role of V. fischeri as the exclusive symbiont of the light organ of the Hawaiian bobtail squid, Euprymna scolopes. Buoyed by this optically amenable host and by persistent and insightful researchers who have applied novel and cross-disciplinary approaches, V. fischeri has developed into a robust model for microbe-host associations. It has contributed to our understanding of how bacteria experience and respond to specific, often fluxing environmental conditions and the mechanisms by which bacteria impact the development of their host. It has also deepened our understanding of numerous microbial processes such as motility and chemotaxis, biofilm formation and dispersal, and bacterial competition, and of the relevance of specific bacterial genes in the context of colonizing an animal host. Parallels in these processes between this symbiont and bacteria studied as pathogens are readily apparent, demonstrating functional conservation across diverse associations and permitting a reinterpretation of "pathogenesis." Collectively, these advances built a foundation for microbiome studies and have positioned V. fischeri to continue to expand the frontiers of our understanding of the microbial world inside animals.

Exploring the potential of chitosan/aragonite biocomposite derived from cuttlebone waste: Elaboration, physicochemical properties and in vitro bioactivity.

Cuttlefish bone biowaste is a potential source of a composite matrix based on chitin and aragonite. In the present work, we propose for the first time the elaboration of biocomposites based on chitosan and aragonite through the valorization of bone waste. The composition of the ventral and dorsal surfaces of bone is well studied by ICP-OES. An extraction process has been applied to the dorsal surface to extract ß-chitin and chitosan with controlled physico-chemical characteristics. In parallel, aragonite isolation was carried out on the ventral side. The freeze-drying method was used to incorporate aragonite into the chitosan polymer to form CHS/ArgS biocomposites. Physicochemical characterizations were performed by FT-IR, SEM, XRD, 1H NMR, TGA/DSC, potentiometry and viscometry. The ICP-OES method was used to evaluate in vitro the bioactivity level of biocomposite in simulated human plasma (SBF), enabling analysis of the interactions between the material and SBF. The results obtained indicate that the CHS/ArgS biocomposite derived from cuttlefish bone exhibits bioactivity, and that chitosan enhances the bioactivity of aragonite. The CHS/ArgS biocomposite showed excellent ability to form an apatite layer on its surface. After three days' immersion, FTIR and SEM analyses confirmed the formation of this layer.

Bacterial growth dynamics in a rhythmic symbiosis.

The symbiotic relationship between the bioluminescent bacterium Vibrio fischeri and the bobtail squid Euprymna scolopes serves as a valuable system to investigate bacterial growth and peptidoglycan (PG) synthesis within animal tissues. To better understand the growth dynamics of V. fischeri in the crypts of the light-emitting organ of its juvenile host, we showed that, after the daily dawn-triggered expulsion of most of the population, the remaining symbionts rapidly proliferate for ∼6 h. At that point the population enters a period of extremely slow growth that continues throughout the night until the next dawn. Further, we found that PG synthesis by the symbionts decreases as they enter the slow-growing stage. Surprisingly, in contrast to the most mature crypts (i.e., Crypt 1) of juvenile animals, most of the symbiont cells in the least mature crypts (i.e., Crypt 3) were not expelled and, instead, remained in the slow-growing state throughout the day, with almost no cell division. Consistent with this observation, the expression of the gene encoding the PG-remodeling enzyme, L,D-transpeptidase (LdtA), was greatest during the slowly growing stage of Crypt 1 but, in contrast, remained continuously high in Crypt 3. Finally, deletion of the ldtA gene resulted in a symbiont that grew and survived normally in culture, but was increasingly defective in competing against its parent strain in the crypts. This result suggests that remodeling of the PG to generate additional 3-3 linkages contributes to the bacterium's fitness in the symbiosis, possibly in response to stresses encountered during the very slow-growing stage.

HOST SWITCHING IN DICYEMIDS (PHYLUM DICYEMIDA).

Dicyemids (phylum Dicyemida) are the most common and most characteristic endosymbionts in the renal sacs of benthic cephalopod molluscs: octopuses and cuttlefishes. Typically, 2 or 3 dicyemid species are found in a single specimen of the host, and most dicyemids have high host specificity. Host-specific parasites are restricted to a limited range of host species by ecological barriers that impede dispersal and successful establishment; therefore, phylogenies of interacting groups are often congruent due to repeated co-speciation. Most frequently, however, host and parasite phylogenies are not congruent, which can be explained by processes such as host switching and other macro-evolutionary events. Here, the history of dicyemids and their host cephalopod associations were studied by comparing their phylogenies. Dicyemid species were collected from 8 decapodiform species and 12 octopodiform species in Japanese waters. Using whole mitochondrial cytochrome c oxidase subunit 1 (COI) sequences, a phylogeny of 37 dicyemid species, including 4 genera representing the family Dicyemidae, was reconstructed. Phylogenetic trees derived from analyses of COI genes consistently suggested that dicyemid species should be separated into 3 major clades and that the most common genera, Dicyema and Dicyemennea, are not monophyletic. Thus, morphological classification does not reflect the phylogenetic relationships of these 2 genera. Divergence (speciation) of dicyemid species seems to have occurred within a single host species. Possible host-switching events may have occurred between the Octopodiformes and Decapodiformes or within the Octopodiformes or the Decapodiformes. Therefore, the mechanism of dicyemid speciation may be a mixture of host switching and intra-host speciation. This is the first study in which the process of dicyemid diversification involving cephalopod hosts has been evaluated with a large number of dicyemid species and genera.

Protein Hydrolysis as a Way to Valorise Squid-Processing Byproducts: Obtaining and Identification of ACE, DPP-IV and PEP Inhibitory Peptides.

The industrial processing of Argentine shortfin squid to obtain rings generates a significant amount of protein-rich waste, including the skin, which is rich in collagen and attached myofibrillar proteins. This waste is generally discarded. In this study, skin was used as a source of proteins that were hydrolysed using Trypsin, Esperase® or Alcalase®, which released peptides with antioxidant potential and, in particular, antihypertensive (ACE inhibition), hypoglycemic (DPP-IV inhibition) and/or nootropic (PEP inhibition) potential. Among the three enzymes tested, Esperase® and Alcalase produced hydrolysates with potent ACE-, DPP-IV- and PEP-inhibiting properties. These hydrolysates underwent chromatography fractionation, and the composition of the most bioactive fractions was analysed using HPLC-MS-MS. The fractions with the highest bioactivity exhibited very low IC50 values (16 and 66 µg/mL for ACE inhibition, 97 µg/mL for DPP-IV inhibition and 55 µg/mL for PEP inhibition) and were mainly derived from the hydrolysate obtained using Esperase®. The presence of Leu at the C-terminal appeared to be crucial for the ACE inhibitory activity of these fractions. The DPP-IV inhibitory activity of peptides seemed to be determined by the presence of Pro or Ala in the second position from the N-terminus, and Gly and/or Pro in the last C-terminal positions. Similarly, the presence of Pro in the peptides present in the best PEP inhibitory fraction seemed to be important in the inhibitory effect. These results demonstrate that the skin of the Argentine shortfin squid is a valuable source of bioactive peptides, suitable for incorporation into human nutrition as nutraceuticals and food supplements.

Thermal accelerated urease-driven hyaluronan-targeted melanin nano-missile for bio-radar detection and chemodrug-free phototherapy.

Polymer-based nanomotors are attracting increasing interest in the biomedical field due to their microscopic size and kinematic properties which support overcoming biological barriers, completing cellular uptake and targeted blasting in limited spaces. However, their applications are limited by the complex viscous physiological environment and lack of sufficient biocompatibility. This manuscript firstly reports a natural melanin nano-missile of MNP@HA-EDA@Urease@AIE PS (MHUA) based on photothermally accelerated urease-driven to achieve chemodrug-free phototherapy. Compared to conventional nano-missiles that only provide driving force, this photothermally accelerated urease-driven nanomotor is independent of chemodrug to maximise biocompatibility, and achieve ideal therapeutic effect through targeted PTT/PDT. In particular, the thermal effect can not only boost the catalytic activity of urease but also achieve ideally anti-tumor effect. In addition, guided by and AIE PS, the nanomotor can generate 1O2 to achieve PDT and be traced in real time serving as an effective fluorescent bio-radar for intracellular self-reporting during cancer treatment. Finally, the targeting ability of MUHA is provided by hyaluronan. Taken together, this MHUA platform provides a simple and effective strategy for target/fluorescence radar detective-guided PTT/PDT combination, and achieves good therapeutic results without chemodrug under thermal accelerated strategy, providing a new idea for the construction of chemodrug-free nanomotor-therapy system.

Squid male alternative reproductive tactics are determined by birth date.

Alternative reproductive tactics (ARTs) are discontinuous phenotypes associated with reproduction, observed in males of many species. Typically, large males adopt a tactic of competing with rivals for mating, while small males adopt a tactic of stealing fertilization opportunities from the large males. The 'birth date hypothesis', proposing that the date of birth influences the determination of each male's reproductive tactic, has been tested only in teleost fish to date. Here, the birth date hypothesis was tested in ARTs of Japanese spear squid Heterololigo bleekeri (consort/sneaker) by analysing statolith growth increments. The birth date significantly differed between consorts (early-hatched) and sneakers (late-hatched). However, no differences were detected in growth history up to 100 days from hatching. Most immature males caught during the reproductive season were larger than sneakers, and their hatch date was similar to that of consorts, suggesting that these immature males had already been following a life-history pathway as a consort. These results indicate that ARTs of H. bleekeri are determined based on their hatch date in early life. This study firstly suggests that the birth date hypothesis applies to aquatic invertebrates, suggesting that the mechanism by which birth date determines the individual phenotype is a phenomenon more common than previously believed.

Microplastics in the Indian and South Atlantic oceans translocate to gills, digestive glands, and muscle of the chokka squid Loligo reynaudii.

Comparative microplastic (MP) data for cephalopods between oceans is scarce. Our aim was to quantify, characterise, and compare MPs in gills, digestive gland, and mantle of chokka squid from the South Atlantic Ocean (SAO) and Indian Ocean (IO) off the coast of South Africa. South African squid had more MPs compared with other studies (means = 2.0 and 0.4 in SAO and IO squid mantle, respectively). Blue fibres were dominant. Identifiable MPs were polyethylene. Despite IO water having higher MP concentrations than the SAO, SAO squid had higher MP concentrations. Dilution by growth is the likely reason for the lower MP concentrations. Fibres were shorter in SAO than IO squid. However, we could not explain why fibre and mantle lengths from both oceans were positively correlated. Squid may not be the best indicator of marine MPs. The characteristics of MPs in squid can be used to track stocks and migrations.

Development of an efficient hemostatic material based on cuttlefish ink nanoparticles loaded in cuttlebone biocomposite.

Traumatic hemorrhage is one of the main causes of mortality in civilian and military accidents. This study aimed to evaluate the effectiveness of cuttlefish bone (cuttlebone, CB) and CB loaded with cuttlefish ink (CB-CFI) nanoparticles for hemorrhage control. CB and CB-CFI were prepared and characterized using different methods. The hemostasis behavior of constructed biocomposites was investigated in vitro and in vivo using a rat model. Results showed that CFI nanoparticles (NPs) are uniformly dispersed throughout the CB surface. CB-CFI10 (10 mg CFI in 1.0 g of CB) showed the best blood clotting performance in both in vitro and in vivo tests. In vitro findings revealed that the blood clotting time of CB, CFI, and CB-CFI10 was found to be 275.4 ± 12.4 s, 229.9 ± 19.9 s, and 144.0 ± 17.5 s, respectively. The bleeding time in rat liver injury treated with CB, CFI, and CB-CFI10 was 158.1 ± 9.2 s, 114.0 ± 5.7 s, and 46.8 ± 2.7 s, respectively. CB-CFI10 composite resulted in more reduction of aPTT (11.31 ± 1.51 s) in comparison with CB (17.34 ± 2.12 s) and CFI (16.79 ± 1.46 s) (p < 0.05). Furthermore, CB and CB-CFI10 exhibited excellent hemocompatibility. The CB and CB-CFI did not show any cytotoxicity on human foreskin fibroblast (HFF) cells. The CB-CFI has a negative surface charge and may activate coagulation factors through direct contact with their components, including CaCO3, chitin, and CFI-NPs with blood. Thus, the superior hemostatic potential, low cost, abundant, simple, and time-saving preparation process make CB-CFI a very favorable hemostatic material for traumatic bleeding control in clinical applications.