Mitsuo Sakai

Ethical and welfare considerations when using cephalopods as experimental animals

When using cephalopods as experimental animals, a number of factors, including morality, quality of information derived from experiments, and public perception, drives the motivation to consider welfare issues. Refinement of methods and techniques is a major step in ensuring protection of cephalopod welfare in both laboratory and field studies. To this end, existing literature that provides details of methods used in the collection, handling, maintenance, and culture of a range of cephalopods is a useful starting point when refining and justifying decisions about animal welfare. This review collates recent literature in which authors have used cephalopods as experimental animals, revealing the extent of use and diversity of cephalopod species and techniques. It also highlights several major issues when considering cephalopod welfare; how little is known about disease in cephalopods and its relationship to senescence and also how to define objective endpoints when animals are stressed or dying as a result of the experiment.

World squid fisheries

Abstract Some 290 species of squids comprise the order Teuthida that belongs to the molluscan Class Cephalopoda. Of these, about 30–40 squid species have substantial commercial importance around the world. Squid fisheries make a rather small contribution to world landings from capture fisheries relative to that of fish, but the proportion has increased steadily over the last decade, with some signs of recent leveling off. The present overview describes all substantial squid fisheries around the globe. The main ecological and biological features of exploited stocks, and key aspects of fisheries management are presented for each commercial species of squid worldwide. The history and fishing methods used in squid fisheries are also described. Special attention has been paid to interactions between squid fisheries and marine ecosystems including the effects of fishing gear, the role of squid in ecosystem change induced by overfishing on groundfish, and ecosystem-based fishery management.

Environmental effects on cephalopod population dynamics: implications for management of fisheries.

Cephalopods are a relatively small class of molluscs (~800 species), but they support some large industrial scale fisheries and numerous small-scale, local, artisanal fisheries. For several decades, landings of cephalopods globally have grown against a background of total finfish landings levelling off and then declining. There is now evidence that in recent years, growth in cephalopod landings has declined. The commercially exploited cephalopod species are fast-growing, short-lived ecological opportunists. Annual variability in abundance is strongly influenced by environmental variability, but the underlying causes of the links between environment and population dynamics are poorly understood. Stock assessment models have recently been developed that incorporate environmental processes that drive variability in recruitment, distribution and migration patterns. These models can be expected to improve as more, and better, data are obtained on environmental effects and as techniques for stock identification improve. A key element of future progress will be improved understanding of trophic dynamics at all phases in the cephalopod life cycle. In the meantime, there is no routine stock assessment in many targeted fisheries or in the numerous by-catch fisheries for cephalopods. There is a particular need for a precautionary approach in these cases. Assessment in many fisheries is complicated because cephalopods are ecological opportunists and stocks appear to have benefited from the reduction of key predator by overexploitation. Because of the complexities involved, ecosystem-based fisheries management integrating social, economic and ecological considerations is desirable for cephalopod fisheries. An ecological approach to management is routine in many fisheries, but to be effective, good scientific understanding of the relationships between the environment, trophic dynamics and population dynamics is essential. Fisheries and the ecosystems they depend on can only be managed by regulating the activities of the fishing industry, and this requires understanding the dynamics of the stocks they exploit.

Genetic Isolation between the Western and Eastern Pacific Populations of Pronghorn Spiny Lobster Panulirus penicillatus

The pronghorn spiny lobster, Panulirus penicillatus, is a circumtropical species which has the widest global distribution among all the species of spiny lobster, ranging throughout the entire Indo-Pacific region. Partial nucleotide sequences of mitochondrial DNA COI (1,142–1,207 bp) and 16S rDNA (535–546 bp) regions were determined for adult and phyllosoma larval samples collected from the Eastern Pacific (EP)(Galápagos Islands and its adjacent water), Central Pacific (CP)(Hawaii and Tuamotu) and the Western Pacific (WP)(Japan, Indonesia, Fiji, New Caledonia and Australia). Phylogenetic analyses revealed two distinct large clades corresponding to the geographic origin of samples (EP and CP+WP). No haplotype was shared between the two regional samples, and average nucleotide sequence divergence (Kimuras two parameter distance) between EP and CP+WP samples was 3.8±0.5% for COI and 1.0±0.4% for 16S rDNA, both of which were much larger than those within samples. The present results indicate that the Pacific population of the pronghorn spiny lobster is subdivided into two distinct populations (Eastern Pacific and Central to Western Pacific), with no gene flow between them. Although the pronghorn spiny lobster have long-lived teleplanic larvae, the vast expanse of Pacific Ocean with no islands and no shallow substrate which is known as the East Pacific Barrier appears to have isolated these two populations for a long time (c.a. 1MY).

Identification of ommastrephid squid paralarvae collected in northern Hawaiian waters and phylogenetic implications for the family Ommastrephidae using mtDNA analysis

A total of 110 adult individuals from four ommastrephid (family Ommastrephidae) squid species (Ommastrephes bartramii, Sthenoteuthis oualaniensis, Eucleoteuthis luminosa, and Hyaloteuthis pelagica) were used to obtain diagnostic DNA markers for species identification. Restriction fragment length polymorphism (RFLP) analysis of a partial segment (855 bp) of the mitochondrial DNA cytochrome oxidase I (COI) gene amplified by polymerase chain reaction (PCR) revealed that the restriction profiles of two endonucleases (Alu I and Tsp509 I) were diagnostic for species identification. The restriction assay partially supplemented with nucleotide sequence analysis successfully assigned 69 damaged and morphologically equivocal ommastrephid paralarvae collected in northern Hawaiian waters, identifying 60 O. bartramii, eight S. oualaniensis, and one E. luminosa. The family Ommastrephidae appears to be monophyletic. Although the phylogenetic relationships among genera were not resolved well due to apparent homoplasy and large genetic divergence between species, COI sequence data without transitions provided support for subfamily level relationships.

Interpretation of statolith microstructure in reared hatchling paralarvae of the squid Illex argentinus

To identify sub-daily or aperiodic increments of statolith growth in the ommastrephid squid Illex argentinus, we examined statolith microstructure, especially with regard to the natal ring, where counting of daily growth increments should begin, and the widths of subsequent daily increments. Paralarvae obtained by artificial fertilisation were incubated on board at different temperatures ranging from 11.4 to 25.4°C, and were starved throughout the experiments. We observed statolith growth from newly hatched to 10-day-old paralarvae and used alizarine complexone staining to attempt validation of the growth. The maximum statolith radius (MSR) of newly hatched paralarvae was constant at 21.1 μm across the full range of temperatures, with the exception of 25.4°C. Daily growth of MSR was analysed separately in two phases, the pre-yolk-absorption phase (i.e. yolk sac still present) and the post-yolk-absorption phase. During the pre-yolk-absorption phase, the daily growth rate (DGR, y) of the MSR varied from 3 to 7 μm day–1 depending on rearing temperature (x) and was expressed as y = 0.37x – 1.77. We concluded that the natal ring forms at 21 μm MSR. The initial increment width obtained from the DGR of MSR seems applicable for distinguishing daily rings from sub-daily rings, although this application should be limited to hatchling paralarvae in the pre-yolk-absorption phase.

Diet and sexual maturation of the neon flying squid Ommastrephes bartramii during autumn and spring in the Kuroshio -Oyashio transition region

Diet and sexual maturation were examined in the winter-spring cohort of the neon flying squid, Ommastrephes bartramii, during its southward migration through the Kuroshio - Oyashio transition region of the western North Pacific. The main prey items are micronektonic animals and small pelagic fish, which were abundantly distributed throughout the study area. Among the prey species, 0. bartramii was dependent on the Japanese anchovy Engraulis japonicus, which also migrate from the northern to southern transition region during the winter, and the micronektonic squid Watasenia scintillans throughout the study period. Other common prey items were Gonatopsis borealis, Diaphus theta, and Ceratoscopelus warmingii from October to November or December, and Tarletonbeania taylori after December. These dietary changes can be explained by the difference in the seasonal north-south migration patterns of the predator and prey species. Male sexual maturation progressed throughout the season, and most individuals were fully mature in January. In contrast, most of the females were immature throughout the study period. The feeding strategy of the squid in relation to their seasonal north to south migration and sexual maturation was discussed.

Molecular evidence for synonymy of the genera Moroteuthis and Onykia and identification of their paralarvae from northern Hawaiian waters

It has been claimed that most squid species in the genus Onykia may be immature stages of species in the genus Moroteuthis. To evaluate the generic status of Moroteuthis and Onykia and to identify paralarvae collected in northern Hawaiian waters, we performed morphological investigation and nucleotide sequence analysis of the mitochondrial cytochrome oxidase I (COI) gene. Of 42 Onykia paralarvae (1.8–8.5 mm dorsal mantle length, DML) examined, 41 had a nucleotide sequence identical to that of M. robusta and one (designated Onykia sp. A) could not be assigned to any known Moroteuthis species. Nucleotide sequence diversity estimates based on Kimura’s two-parameter distances between Onykia sp. A and Moroteuthis spp. (0.109–0.150) fell well within the range of congeneric species, suggesting that Onykia sp. A is a member of the genus Moroteuthis. Molecular data supported the conclusion that the genus Moroteuthis is a junior synonym of the genus Onykia. Morphological investigation revealed that paralarvae of O. robusta (= M. robusta) <4.0 mm DML were distinct from other Onykia paralarvae in the chromatophore pattern on the mantle. The key characters for distinguishing O. robusta paralarvae from Onykia sp. A were the number and arrangement of chromatophores on the funnel.

Genetic Diversity and Population Structure of Pronghorn Spiny Lobster Panulirus penicillatus in the Pacific Region

Abstract: Adult specimens of Panulirus penicillatus were collected from localities distributed in Japan: Hachijojima Island, Ryukyu Archipelago (Amamioshima, Okinawajima, Ishigakijima); Taiwan: Taitung; Indonesia: Java Sea and Gebe Island; French Polynesia: Moorea Island; and Ecuador: Isabela Island. Phyllosoma larval specimens were collected from the western sea of the Galápagos Islands. In total, 569–570 nucleotide sequences were analyzed from the mtDNA control region of 480 individuals. Almost all individuals had a unique haplotype. Intra-population haplotype (h) and nucleotide (&pgr;) diversities were high for each locality, ranging h = 0.9996-1.0000 and &pgr; = 0.0255-0.0448. A mismatch distribution of pairwise differences between haplotypes indicated that P. penicillatus does not fit the sudden population expansion model. Analysis of molecular variance (AMOVA) shows a clear genetic difference between two regions (western/central Pacific and eastern Pacific). High gene flow was found within localities in the western/central Pacific region, probably resulting from an extended planktonic larval stage and prevailing ocean currents. In this study we detected genetic structure between eastern Pacific populations and western Pacific populations, indicating that despite the nearly 1-yr larval period for this species, larvae generally do not pass over the East Pacific Barrier.

Evaluation of the 5′ end of the 16S rRNA gene as a DNA barcode marker for the Cephalopoda

The present study seeks to incorporate a highly variable DNA barcode marker, additional to the standard regions of the cytochrome oxidase I gene and the 3′ end of the 16S large ribosomal subunit (16S rRNA), for a more effective species-level identification among cephalopods. Thus, we evaluated whether the 5′ end region of the 16S rRNA gene can be suitable as a DNA barcode marker among these taxa. Using a novel primer set, 28 different species were evaluated based on the pairwise intra- and interspecific distance and neighbor-joining (NJ) analysis. Except for Enteroctopus dofleini, we were able to obtain the sequences for the remaining species which formed highly supported clusters in the NJ tree. Divergence at the interspecific level may indicate high genetic variation of the region being proposed as compared to the standard of 16S rRNA. In addition, two well-supported clades and high levels of divergence within Sthenoteuthis oualaniensis, Loliolus japonica and Sepia pharaonis suggest the occurrence of cryptic species. This study confirms the efficiency of the 5′ end region of the 16S rRNA gene as a DNA barcode marker which can be used along with the standard DNA markers in future studies.