Sepia Officinalis Research Articles

Cuttlebone calcification increases during exposure to elevated seawater pCO2 in the cephalopod Sepia officinalis

Changes in seawater carbonate chemistry that accompany ongoing ocean acidification have been found to affect calcification processes in many marine invertebrates. In contrast to the response of most invertebrates, calcification rates increase in the cephalopod Sepia officinalis during long-term exposure to elevated seawater pCO2. The present trial investigated structural changes in the cuttlebones of S. officinalis calcified during 6 weeks of exposure to 615 Pa CO2. Cuttlebone mass increased sevenfold over the course of the growth trail, reaching a mean value of 0.71 ± 0.15 g. Depending on cuttlefish size (mantle lengths 44–56 mm), cuttlebones of CO2-incubated individuals accreted 22–55% more CaCO3 compared to controls at 64 Pa CO2. However, the height of the CO2-exposed cuttlebones was reduced. A decrease in spacing of the cuttlebone lamellae, from 384 ± 26 to 195 ± 38 μm, accounted for the height reduction The greater CaCO3 content of the CO2-incubated cuttlebones can be attributed to an increase in thickness of the lamellar and pillar walls. Particularly, pillar thickness increased from 2.6 ± 0.6 to 4.9 ± 2.2 μm. Interestingly, the incorporation of non-acid-soluble organic matrix (chitin) in the cuttlebones of CO2-exposed individuals was reduced by 30% on average. The apparent robustness of calcification processes in S. officinalis, and other powerful ion regulators such as decapod cructaceans, during exposure to elevated pCO2 is predicated to be closely connected to the increased extracellular [HCO3 −] maintained by these organisms to compensate extracellular pH. The potential negative impact of increased calcification in the cuttlebone of S. officinalis is discussed with regard to its function as a lightweight and highly porous buoyancy regulation device. Further studies working with lower seawater pCO2 values are necessary to evaluate if the observed phenomenon is of ecological relevance.

Correspondence of Collagen on the Heat-Induced Softening of Frozen-Thawed Cuttlefish Sepia officinalis Mantle

Correspondence of Collagen on the Heat-Induced Softening of Frozen-Thawed Cuttlefish Sepia officinalis Mantle

Analysis of Novel Angiotensin I-Converting Enzyme Inhibitory Peptides from Enzymatic Hydrolysates of Cuttlefish (Sepia officinalis) Muscle Proteins

The angiotensin I-converting enzyme (ACE) inhibitory activities of protein hydrolysates prepared from cuttlefish (Sepia officinalis) proteins by treatment with various bacterial proteases were investigated. The hydrolysate generated by the crude enzyme from Bacillus mojavensis A21 displayed the highest ACE inhibitory activity, and the higher inhibition activity (87.11 +/- 0.92% at 2 mg/mL) was obtained with hydrolysis degree of 16%. This hydrolysate was fractionated by size exclusion chromatography on a Sephadex G-25 into eight major fractions (P(1)-P(8)). Fraction P(6), which exhibited the highest ACE inhibitory activity, was then fractionated by reversed-phase high performance liquid chromatography (RP-HPLC). Eleven ACE inhibitory peptides were isolated, and their molecular masses and amino acids sequences were determined using ESI-MS and ESI-MS/MS, respectively. The structures of the most potent peptides were identified as Ala-His-Ser-Tyr, Gly-Asp-Ala-Pro, Ala-Gly-Ser-Pro and Asp-Phe-Gly. The first peptide displayed the highest ACE inhibitory activity with an IC(50) of 11.6 microM. The results of this study suggest that cuttlefish protein hydrolysates are a good source of ACE inhibitory peptides.

Three novel angiotensin I-converting enzyme (ACE) inhibitory peptides from cuttlefish (Sepia officinalis) using digestive proteases

Abstract The angiotensin I-converting enzyme (ACE) inhibitory activities of protein hydrolysates prepared from muscle of cuttlefish ( Sepia officinalis ) by treatment with various digestive proteases were investigated. The most active hydrolysate was obtained with the crude protease extract from the hepatopancreas of cuttlefish (64.47 ± 1.0% at 2 mg of dry weight/ml) with a degree of hydrolysis of 8%. By gel filtration on Sephadex G-25 and RP-HPLC on C18 column, three novel peptides with high ACE-inhibitory activity were purified and their molecular masses and amino acid sequences were determined. The three peptides Val-Tyr-Ala-Pro, Val-Ile-Ile-Phe and Met-Ala-Trp with IC 50 values of 6.1, 8.7 and 16.32 μM, respectively, were novel ACE-inhibitory peptides. Lineweaver–Burk plots suggest that the three purified peptides act as non-competitive inhibitors against ACE. These results suggest that some peptides from cuttlefish could be a beneficial ingredient for nutraceuticals against hypertension.

Localization of ion-regulatory epithelia in embryos and hatchlings of two cephalopods

The tissue distribution and ontogeny of Na(+)/K(+)-ATPase has been examined as an indicator for ion-regulatory epithelia in whole animal sections of embryos and hatchlings of two cephalopod species: the squid Loligo vulgaris and the cuttlefish Sepia officinalis. This is the first report of the immunohistochemical localization of cephalopod Na(+)/K(+)-ATPase with the polyclonal antibody alpha (H-300) raised against the human alpha1-subunit of Na(+)/K(+)-ATPase. Na(+)/K(+)-ATPase immunoreactivity was observed in several tissues (gills, pancreatic appendages, nerves), exclusively located in baso-lateral membranes lining blood sinuses. Furthermore, large single cells in the gill of adult L. vulgaris specimens closely resembled Na(+)/K(+)-ATPase-rich cells described in fish. Immunohistochemical observations indicated that the amount and distribution of Na(+)/K(+)-ATPase in late cuttlefish embryos was similar to that found in juvenile and adult stages. The ion-regulatory epithelia (e.g., gills, excretory organs) of the squid embryos and paralarvae exhibited less differentiation than adults. Na(+)/K(+)-ATPase activities for whole animals were higher in hatchlings of S. officinalis (157.0 +/- 32.4 micromol g (FM) (-1) h(-1)) than in those of L. vulgaris (31.8 +/- 3.3 micromol g (FM) (-1) h(-1)). S. officinalis gills and pancreatic appendages achieved activities of 94.8 +/- 18.5 and 421.8 +/- 102.3 micromol(ATP) g (FM) (-1) h(-1), respectively. High concentrations of Na(+)/K(+)-ATPase in late cephalopod embryos might be important in coping with the challenging abiotic conditions (low pH, high pCO(2)) that these organisms encounter inside their eggs. Our results also suggest a higher sensitivity of squid vs. cuttlefish embryos to environmental acid-base disturbances.

Use of Amphipods as alternative prey to culture cuttlefish ( Sepia officinalis) hatchlings

The effects of feeding two alternative live prey (exclusively caprellids ( Caprella equilibra) or several species of gammarids, mainly Ericthonius brasiliensis, Jassa marmorata and Elasmopus sp.), to cuttlefish hatchlings were compared to feeding mysids ( Mesopodopsis slabberi), which are normally used during the first weeks of the life cycle. Weight (g) and growth rates (GR, % BW d − 1 ) were determined. Cuttlefish hatchlings fed with mysids and gammarids grew faster (6.7 ± 0.4 and 5.7 ± 0.9% BW d − 1 , respectively) compared to caprellids (1.6 ± 0.2% BW d − 1 ). Survival was higher (96.7 ± 5.8%) for hatchlings fed mysids, compared to 83.3 ± 15.3% and 76.7 ± 5.8%, for those fed gammarids and caprellids, respectively. According to the results obtained, gammarids could be used as an alternative prey to mysids, while Caprella equilibra did not deliver appropriate growth rates and should be disregarded as alternative prey for rearing early stages (hatchlings) of Sepia officinalis. This is the first study revealing a successful use of amphipods, mainly gammarids, as alternative prey for cuttlefish hatchlings.

Purification, characterization and analysis of sepia melanin from commercial sepia ink (Sepia officinalis).

At the moment melanins are still enigmatic bio-pigments with structure and determination methods not clearly understood for the international scientific community. Among biopolymers, melanins are unique in many aspects. Some essential biopolymers (proteins, nucleic acids and carbohydrates) are chemically well characterized and can be determined using well established analytical methodologies. On the contrary, no available methods allow us to accurately determine melanin, mostly due to the intrinsic chemical properties of melanins. These pigments, in fact, are insoluble in a broad range of solvents and pH as well as difficult to purify as a result of the heterogeneity in their structural features. Mammalian melanins exist in two chemically distinct forms: the brown to black Eumelanin and the yellow to reddish-brown Pheomelanin. Sepia melanin obtained from Sepia Officinalis consists of more than 98% of Eumelanin and is therefore used as standard material in the analysis of melanic black. Commercial sepia melanin is purified according to an undescribed procedure. In this research, extraction and purification studies were carried out on sepia melanin using a hydrochloric acid (0.5 - 3.0 M) treatment under mechanical or ultrasonic agitation. A high degree purity sepia melanin was obtained and further characterized using Elemental Analysis (EA), UltravioletVisible (UV-VIS) and Infrared (IR) spectroscopy and Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) for metal ion analysis. Finally, quantitative studies of the chemical degradation of Eu polymers to Pyrrole-2,3,5tricarboxylic acid (PTCA) and Pyrrole-2,3-dicarboxylic acid (PDCA) by High Performance Liquid Chromatography (HPLC) were performed. The procedure we developed can be used to obtain a considerably less expensive sepia melanin which could prove important as a standard in future determinations of eumelanin and pheomelanin as well as in a vast field of scientific and industrial applications in such fields as human and veterinary medicine, pharmacology and cosmetics.

Some Biological Aspects of Cuttlebone (Sepia officinalis L., 1758) off the Coastal Area of Karataş (Northeastern Mediterranean -TURKEY)

Some Biological Aspects of Cuttlebone (Sepia officinalis L., 1758) off the Coastal Area of Karataş (Northeastern Mediterranean -TURKEY)

Visual contrast modulates maturation of camouflage body patterning in cuttlefish (Sepia pharaonis).

Camouflage is the primary defense behavior in cephalopods. It is known that cuttlefish immediately after hatching are capable of showing various body patterns for concealing themselves, however recent studies suggest that maturation of camouflage body patterns is faster for cuttlefish (Sepia officinalis) reared in enriched environments than those reared in impoverished environments. Since camouflage patterning in cephalopods is predominately visually driven, this study specifically investigates effects of the rearing background contrast on the maturation of body patterns in cuttlefish (Sepia pharaonis). Newly hatched animals were separated into two cohorts, one reared in a uniform-gray background (low-contrast, or L group) and the other raised in a black/white checkerboard background (high-contrast, or H group). At Weeks 2, 4, 6, 8, 10, and 12, cuttlefish were placed individually either on uniform or checkerboard substrates to examine their body patterns. Animals from both L and H groups appear to show moderate disruptive patterns on the checkerboard and less disruptive on the uniform background at Week 2. Throughout development, however, cuttlefish from the H group showed stronger disruptive patterns than that of the L group on the checkerboard background at Weeks 10 and 12. In interesting findings, cuttlefish from both L and H groups showed similar strength but different disruptive components on the uniform background in later postembryonic stages. These results suggest that the maturation of camouflage body patterns in S. pharaonis is at least in part affected by visual contrast of their rearing backgrounds, although environmental complexity or social interaction is also likely to be involved in this process. This also implies that early visual experience could exert its effect on the seemingly preprogrammed behaviors such as camouflage body patterning in cephalopods.

Mechanical Performance and In Vivo Tests of an Acrylic Bone Cement Filled with Bioactive Sepia Officinalis Cuttlebone

To promote osteointegration, bioactive cuttlebone particles containing collagen were used to fill an acrylic cement, varying filler concentration (0–50 wt%). Cuttlebone was characterized by X-ray diffraction, plasma atomic emission and FT-IR. Mechanical properties of the filled cement were determined following ASTM procedures, included stress–strain, compression, bending, and fracture toughness tests. For in vivo tests, three groups of seven adult healthy rabbits were prepared to make an implant in the parietal bone of each one. For such groups (I–III), the amount of filler in the cement was 0, 10 and 30 wt%, respectively. Mechanical results for the composites complied with norm requirements. However, as mechanical performance for composite with 50 wt% of filler decreased significantly, for the in vivo tests, such composite was excluded. In vivo tests showed that three implants of group I were loosely attached to the parietal bone, whereas all the implants made with cement containing cuttlebone particles (groups II and III) were firmly attached to the parietal bone, indicating osteointegration. These results clearly show the potential of this type of bioactive filler to be used for medical applications.

Optical and Paramagnetic Properties of Size-Controlled Ink Particles Isolated fromSepia officinalis

The optical and paramagnetic properties of size-controlled ink particles isolated from ink sacs of Sepia officinalis were investigated. Topographic images of atomic force microscopy (AFM) revealed that the average heights of the large and small ink particles were 156 nm and 5.3 nm respectively. The ultraviolet-visible (UV-VIS) spectral features of aqueous solutions of ink particles were dependent on particle size. Electron spin resonance (ESR) spectra suggested that the ink particles are highly pure for paramagnetic species and are of reliable quality. These size-controlled ink particles are suitable for a basic study of melanin-related materials.

Cuttlefish dynamic camouflage: responses to substrate choice and integration of multiple visual cues

Prey camouflage is an evolutionary response to predation pressure. Cephalopods have extensive camouflage capabilities and studying them can offer insight into effective camouflage design. Here, we examine whether cuttlefish, Sepia officinalis, show substrate or camouflage pattern preferences. In the first two experiments, cuttlefish were presented with a choice between different artificial substrates or between different natural substrates. First, the ability of cuttlefish to show substrate preference on artificial and natural substrates was established. Next, cuttlefish were offered substrates known to evoke three main camouflage body pattern types these animals show: Uniform or Mottle (function by background matching); or Disruptive. In a third experiment, cuttlefish were presented with conflicting visual cues on their left and right sides to assess their camouflage response. Given a choice between substrates they might encounter in nature, we found no strong substrate preference except when cuttlefish could bury themselves. Additionally, cuttlefish responded to conflicting visual cues with mixed body patterns in both the substrate preference and split substrate experiments. These results suggest that differences in energy costs for different camouflage body patterns may be minor and that pattern mixing and symmetry may play important roles in camouflage.

Edge detection and texture classification by cuttlefish

Cephalopod mollusks including octopus and cuttlefish are adept at adaptive camouflage, varying their appearance to suit the surroundings. This behavior allows unique access into the vision of a non-human species because one can ask how these animals use spatial information to control their coloration pattern. There is particular interest in factors that affect the relative levels of expression of the Mottle and the Disruptive body patterns. Broadly speaking, the Mottle is displayed on continuous patterned surfaces whereas the Disruptive is used on discrete objects such as pebbles. Recent evidence from common cuttlefish, Sepia officinalis, suggests that multiple cues are relevant, including spatial scale, contrast, and depth. We analyze the body pattern responses of juvenile cuttlefish to a range of checkerboard stimuli. Our results suggest that the choice of camouflage pattern is consistent with a simple model of how cuttlefish classify visual textures, according to whether they are Uniform or patterned, and whether the pattern includes visual edges. In particular, cuttlefish appear to detect edges by sensing the relative spatial phases of two spatial frequency components (e.g., fundamental and the third harmonic Fourier component in a square wave). We discuss the relevance of these findings to vision and camouflage in aquatic environments.

Characterization of a novel LFRFamide neuropeptide in the cephalopod Sepia officinalis

From a single LC–MS/MS analysis, a new C-terminally extended RFamide neuropeptide was characterized in Sepia officinalis. The experimental strategy was based on the specific neutral loss associated with RFamide breakdown. Mass losses of 17 Da (C-terminally amide) and 320 Da (RFamide) have been observed for three known peaks of m/ z 581.7 (FLRFamide), 599.8 (FMRFamide), 1096.3 (ALSGDAFLRFamide) and one unknown of m/ z 752.8. The primary sequence of the peptide of m/ z 752.8 was GNLFRFamide. MS/MS analyses revealed that this novel neuropeptide, called sepFRF1, is largely distributed in the central nervous system of cuttlefish of both sexes. Probably transported in the visceral nerve from the subesophageal mass (the peptide was not detected in the hemolymph), this neuropeptide targeted the rectum in agreement with its peripheral distribution. From concentrations as low as 10 −9 M, sepFRF1 increased the frequency, tonus and amplitude of rectal contractions. SepFRF1 is the first RFamide peptide identified in Sepia officinalis that is not derived from the FaRPs precursor. SepFRF1 could belong to a RFamide subfamily identified in gastropods and may be involved in feeding behavior.

Transscleral visible/near-infrared spectroscopy for quantitative assessment of melanin in a uveal melanoma phantom of ex vivo porcine eyes

Optical spectroscopy has been used as a supplement to conventional techniques for analyzing and diagnosing cancer in many human organs. Because ocular tumors may be characterized by their different melanin content, we investigated the feasibility of using transscleral visible/near-infrared spectroscopy (Vis/NIRS) to estimate the quantity of melanin in a novel uveal melanoma phantom of ex vivo porcine eyes. The phantoms were made by injecting a freshly prepared suspension of 15% (wt/vol) gelatin, 10 mg/ml titanium dioxide (TiO 2), and natural melanin, isolated from the ink sac of cuttlefish ( Sepia officinalis), into the suprachoroidal space of 30 enucleated porcine eyes. The melanin concentrations used were 1 mg/ml, 2 mg/ml, and 3 mg/ml, with 10 eyes in each group. After gelation, the size and location of the phantoms were documented by B-scan ultrasonography and transillumination. Vis/NIRS recordings, covering the wavelength region from 550 to 1000 nm, were performed with two optical fibers separated by 6 mm to deliver and collect the light through the sclera. During all measurements, the exact pressure exerted by the fiber probe on the scleral surface was monitored by placing the eye on an electronic scale. Transscleral Vis/NIRS was performed across the phantom inclusion, as well as on the opposite (normal) side of each eye. A total of three consecutive measurements were carried out alternately on each side of the globe. The spectral data were analyzed using partial least squares regression. In the melanin concentration groups of 1 mg/ml ( n = 10), 2 mg/ml ( n = 10), and 3 mg/ml ( n = 10), the largest basal phantom diameters (mean ± SD) were 14.9 ± 1.6 mm, 14.6 ± 1.5 mm, and 14.3 ± 1.0 mm, respectively ( p > 0.05). The largest phantom thicknesses (mean ± SD) were 4.0 ± 0.5 mm, 4.4 ± 0.7 mm, and 4.5 ± 0.5 mm, respectively ( p > 0.05). Statistical regression modeling of the Vis/NIRS data revealed that it was possible to correctly classify the phantoms according to their melanin concentrations in 84.4% of cases. The correct classification rate for phantoms with the lowest (1 mg/ml) and highest (3 mg/ml) melanin concentrations was 99.2%. The study demonstrates that transscleral Vis/NIRS is a feasible and accurate method for predicting the content of melanin in choroidal lesions.

Population identification of common cuttlefish (<em>Sepia officinalis</em>) inferred from genetic, morphometric and cuttlebone chemistry data in the NE Mediterranean Sea

The population structures of the common cuttlefish Sepia officinalis from the north-eastern Mediterranean (Antalya and Iskenderun Bays), Aegean (Izmir Bay) and Marmara Seas were analyzed with mtDNA PCR-RFLP, body morphometry and cuttlebone chemistry. Analysis of a ND 5/6 (Nikotin Amid Adenin Dehidrojenaz-5/6) gene segment of mtDNA revealed seven haplotypes from 120 individuals. No haplotype sharing was observed among sampling sites. The average nucleotide divergence between samples was 0.009390, and the highest genetic divergence (0.015279) was observed between the Iskenderun Bay and Marmara Sea samples. The lowest genetic divergence (0.003786) was between the Aegean Sea and Antalya Bay samples. Highly significant differences (P S. officinalis in Turkish coastal waters.

Effects of increased <i>p</i>CO<sub>2</sub> and temperature on trace element (Ag, Cd and Zn) bioaccumulation in the eggs of the common cuttlefish, <i>Sepia officinalis</i>

Abstract. Cephalopods play a key role in many marine trophic networks and constitute alternative fisheries resources, especially given the ongoing decline in finfish stocks. Along the European coast, the eggs of the cuttlefish Sepia officinalis are characterized by an increasing permeability of the eggshell during development, which leads to selective accumulation of essential and non-essential elements in the embryo. Temperature and pH are two critical factors that affect the metabolism of marine organisms in the coastal shallow waters. In this study, we investigated the effects of pH and temperature through a crossed (3×2; pH 8.1 (pCO2, 400 ppm), 7.85 (900 ppm) and 7.6 (1400 ppm) at 16 and 19°C, respectively) laboratory experiment. Seawater pH showed a strong effect on the egg weight and non-significant impact on the weight of hatchlings at the end of development implying an egg swelling process and embryo growth disturbances. The lower the seawater pH, the more 110 mAg was accumulated in the tissues of hatchlings. The 109Cd concentration factor (CF) decreased with decreasing pH and 65Zn CF reached maximal values pH 7.85, independently of temperature. Our results suggest that pH and temperature affected both the permeability properties of the eggshell and embryonic metabolism. To the best of our knowledge, this is one of the first studies on the consequences of ocean acidification and ocean warming on metal uptake in marine organisms, and our results indicate the need to further evaluate the likely ecotoxicological impact of the global change on the early-life stages of the cuttlefish.

Contrasting accumulation biokinetics and distribution of 241Am, Co, Cs, Mn and Zn during the whole development time of the eggs of the common cuttlefish, Sepia officinalis

Cuttlefish eggs were exposed to 241Am, Co, Cs, Mn and Zn for different periods of time during the 50-d of the embryonic development at 17 °C. Exposures were carried out using background dissolved concentrations of the metals, using the corresponding γ-emitting radiotracers ( 241Am, 57Co, 134Cs, 54Mn and 65Zn). Eggs were then placed in non-contaminating conditions. Experiments allowed assessing 1) the uptake and depuration kinetics of the selected elements and 2) their distribution among the different egg compartments (i.e. eggshell, vitellus, peri-vitelline fluid and embryo). 241Am, Co and Zn were accumulated continuously by the eggs all along the development time, whereas Mn reached saturation after one month of exposure. Interestingly, the uptake kinetics of Cs tightly followed the weight variation of the eggs, mainly because of water influx/outflux. During the first month of the embryonic life, 241Am, Co, Cs, Mn and Zn remained associated with the eggshell, indicating that the latter acted as an efficient shield against their penetration. Nevertheless, from this time onwards, Co, Cs, Mn and Zn accumulated more or less efficiently in the embryo according to the following order: Cs > Zn > Mn > Co. 241Am was the only element tested that did not cross the eggshell all along the exposure time. The depuration kinetics revealed that the retention capacity of the eggs varied during the embryonic life. The contrasting accumulation biokinetics are discussed in terms of chemical and biological processes occurring during the cuttlefish egg development.

Ontogeny of Oxytocin-Like Immunoreactivity in the Cuttlefish, Sepia officinalis, Central Nervous System

In vertebrate species, the neuropeptide oxytocin (OT) has been implicated in neural and behavioral development. Although several OT-like peptides have been characterized in invertebrate species, the ontogenesis of the OT-like system has not yet been described in these species. Thus, the aim of the present study was to perform an immunohistochemical investigation of the spatiotemporal distribution of OT-like elements in the central nervous system (CNS) of a decapod cephalopod mollusc, the cuttlefish, Sepia officinalis, during the first 3 months of postembryonic development. On the day of birth, OT-like immunoreactivity was detected throughout the whole CNS. Some nervous structures (e.g. the magnocellular lobes) exhibited a stained pattern in newborns similar to that reported in our previous study in adult cuttlefish whereas other lobes (e.g. the vertical lobe complex) showed maturation during the first weeks of life. Finally, at the age of 60 days, the general pattern of staining in the CNS was comparable to the adult distribution. The putative roles of the OT-like system with regard to the development of some behaviors in juvenile cuttlefish are discussed. The present study provides a neurochemical basis for the investigation of postnatal development of complex behaviors in cephalopods and suggests, for the first time in an invertebrate species, important organizational effects for the OT-like system in the course of the first weeks of life.

Characterization of a novel SINE superfamily from invertebrates: “Ceph-SINEs” from the genomes of squids and cuttlefish

Five tRNA-derived short interspersed repetitive elements (SINEs), named SepiaSINE, Sepioth-SINE1, Sepioth-SINE2A, Sepioth-SINE2B and OegopSINE, were isolated from the genomes of three decabrachian species [ Sepia officinalis (order Sepiida), Sepiotheuthis lessoniana (suborder Myopsida), and Mastigoteuthis cordiformes (suborder Oegopsida)], by random sequencing and genome screening. In addition, two tRNA-derived SINEs, named IdioSINE1 and IdioSINE2, were further detected from EST (expressed sequence tag) data of Idiosepius paradoxus (order Idiosepiida), using a GenBank FASTA search with a conserved sequence of the SepiaSINE as the query. All the isolated SINEs had a common and unique highly conserved 149-bp sequence in their central structures (Sepioth-SINE2B and IdioSINEs, however, had a continuous 73-bp deletion in the conserved region.), and are therefore grouped as the fourth SINE superfamily “Ceph-SINEs”, following the CORE-SINE, V-SINE, and DeuSINE superfamilies. Our analysis suggested that the central conserved region called the “Ceph-domain” might have originated before the diversification of cephalopods (505 myr ago). A sequence alignment of Sepioth-SINE1, Sepioth-SINE2A, and Sepioth-SINE2B demonstrated that Sepioth-SINE2A has a chimeric structure shared with two other SINEs. The above relationship suggests possible template switching in the central conserved domain during reverse transcription for the birth of Sepioth-SINE2A, providing the possibility that the presence of the conserved domain contributed to yield a variety of SINEs during evolution. Furthermore, the distributions of the isolated SINEs showed that order Sepiida, suborders Oegopsida and Myopsida, and order Idiosepiida have their own independent SINE(s), and suggest that order Sepiida can be largely separated into two groups, with clarification of the phylogenetic relatedness between subfamily Sepioteuthinae and the other loliginid squids.