Abstract: 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.

Abstract: Genetic differentiation of the Atlantic swordfish (Xiphias gladius) was investigated by a single nucleotide polymorphism (SNP) at the calmodulin gene (CAM) intron locus. Clearly distinct allele and genotype frequencies were observed between the north (20°N-41°N) and mid-south (10°N-33°S) Atlantic samples. Much lower frequency of A allele (37.5 to 57.1 %) was observed in the north samples (n=160 in total) than in the mid-south samples (83.3-92.6 %; n=354), and homozygote BB was common in the north samples (23.4-31.3 %) but very rare or absent (0-3.9 %) in the mid-south samples. Very strong population subdivision was observed between the two groups (FST = 0.34, P < 0.001), while the allele and genotype frequencies within each ocean basin persisted over time (1990 to 2002 in the north, and 1994 to 2002 in the mid-south). Of two samples from the presumed boundary zone, one (n=18) (14°N, 48°W) presented intermediate frequencies of the A allele (66.7 %) and BB homozygote (11.1 %), while the other (n=23) (10°N-17°N, 28°W-37°W) shared similar frequencies of the A allele (89.1 %) and BB homozygote (4.3 %) with those of the mid-south Atlantic samples. These results indicate that the gene flow and individual migration between the north and mid-south Atlantic populations are considerably restricted and that the current management boundary between the north and south Atlantic swordfish stocks of 5ºN should be reconsidered.

Abstract: 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.

Abstract: To resolve the controversial taxonomic status of two closely-related lanternfishes, Diaphus perspicillatus (Ogilby, 1898) and D. gigas Gilbert, 1913, mitochondrial cytochrome b (cyt b) gene sequences of representative individuals were obtained, and specifically-diagnostic restriction sites investigated. In the 5'-half nucleotide sequences of the cyt b gene (594 bp), a relatively high level of nucleotide substitution between the species (8.2�8.7%), together with very low intraspecific variation (0.86%), was observed, which clearly indicates the two morphs to be specifically distinct. Previous hypotheses regarding gigantism of D. gigas and suspicions regarding the synonymy of the two species are rejected. The nucleotide sequences and practical restriction enzyme assay indicate that any one of four restriction endonucleases (Fok I, Hae III, Nla IV, and Rsa I) can unambiguously discriminate between the two species. Based on specimens identified by molecular analysis, eye diameter and gill raker count were found to be good diagnostic morphological characters.

Abstract: Intra and interspecific nucleotide sequence variation of rDNA first internal transcribed spacer (ITS1) was analysed using all eight species of the genus Thunnus plus two out�group species within the same family, skipjack tuna Katsuwonus pelamis and striped bonito Sarda orientalis. Intraspecific nucleotide sequence variation in ITS1, including intra�genomic variation, was low, ranging from 0·003 to 0·014 [Kimura's two parameter distance (K2P)], whereas variation between species within the genus Thunnus ranged from 0·009 to 0·05. The Atlantic and Pacific northern bluefin tunas Thunnus thynnus thynnus and Thunnus thynnus orientalis, recently proposed to be distinct species, were found to share nearly identical ITS1 sequences (mean K2P = 0·006) well within the range of intraspecific variation. The northern bluefin tuna appeared to be a sister group to albacore Thunnus alalunga, with all other Thunnus species in a distinct clade. The ITS1 phylogeny was consistent with mtDNA phylogeny in clustering the three tropical Thunnus species (T. albacares, T. atlanticus and T. tonggol). Southern bluefin Thunnus maccoyii and bigeye Thunnus obesus tunas showed a closer affinity to this tropical tuna group than to the northern bluefin tuna and albacore. The molecular data supported mitochondrial introgression between species and contradicted morphological subdivision of the genus into two subgenera Neothunnus and Thunnus.

Abstract: A molecular approach was adopted to investigate the natural diets of palinurid and scyllarid lobster phyllosoma larvae. The central domain of the 18S rDNA gene was amplified using nested polymerase chain reaction (PCR) and genomic DNA extracted from the larval hepatopancreas. The resulting 18S rDNA clones were screened using restriction fragment length polymorphism (RFLP) analysis, and then FASTA homology search and phylogenetic analysis were performed on the nucleotide sequences to identify the source of the eukaryotic organisms. The feasibility of this method was confirmed using the laboratory-reared phyllosoma larvae of the Japanese spiny lobster Panulirus japonicus that were fed either with common mussel Mytilus edulis gonads or with Artemia nauplii exclusively. Among the 804 clones isolated from five wild-caught mid- to late-stage phyllosoma larvae (three palinurids and two scyllarids), 0�132 clones per sample possessed restriction profiles distinct from those of the hosts. The Cnidaria and Urochordata DNA were identified from both the palinurid and the scyllarid larvae, which were thought to be prey animals for the mid- to late-stage phyllosoma larvae.

Abstract: To identify lobster phyllosoma larvae of the genus Panulirus occurring in waters adjacent to Japan, genetic variation within and between 10 Indo-Pacific lobster species was investigated using restriction fragment length polymorphism (RFLP) analysis for the 1300-base pair mitochondrial cytochrome oxidase I (COI) gene. RFLP analysis using two endonucleases (AluI and TaqI) enabled discrimination of all species, including the P. longipes complex. The diagnostic DNA markers, supplemented with nucleotide sequence analysis, were applied to 44 mid- to late-stage phyllosoma larvae (7.4 to 27.7 mm in body length) collected in the northwestern Pacific. These samples were unexpectedly variable in species composition, comprising P. japonicus (n = 16), P. longipes bispinosus (21), P. longipes longipes (1), P. ��aka�� (1), and P. penicillatus (5). Comparison of larval size at similar stages revealed that P. l. bispinosus larvae were significantly larger than P. japonicus.

Abstract: A later stage phyllosoma larva of the genus Panulirus was caught in the central Atlantic in October 2000. Nucleotide sequence analysis of mitochondrial 16S rDNA indicated this larva to be of P. echinatus, which has been undescribed to date. Morphological examination indicated that this 8th stage larva appeared to belong to phyllosoma-species group 2, distinguishable from the other Atlantic species of the genus but closely related to an Indo-Pacific species P. penicillatus, corresponding to the results of molecular phylogenetic analysis.

Abstract: Nucleotide sequence analysis of the mitochondrial COI gene was performed to identify mid to final stage phyllosoma larvae of spiny lobsters of the genus Panulirus caught in the south of the Ryukyu Archipelago (northwest Pacific). The identified larvae were subjected to morphological investigation. All 92 larvae caught in May 2003 were late to final stage phyllosomas of P. japonicus, while 174 larvae in November 2004 comprised four species of phyllosoma group 1 (P. femoristriga (n = 1), P. japonicus (4), P. longipes bispinosus (110), and P. l. longipes (7)), one of group 2 (P. penicillatus (43)), and two of group 4 (P. ornatus (8) and P. versicolor (1)). Photo images of representative phyllosoma larvae, substantiated by molecular identification, are presented Morphological investigation indicated that the ratio between the widths of cephalon and thorax (CT ratio) of the P. japonicus phyllosomas was the smallest, and discrimination of P. japonicus larvae from the other species of the �P. japonicus group� may be possible using the CT ratio combined with the smaller body size. Intraspecific variation was observed in the arrangement of subexopodal spines of P. ornatus and P. versicolor phyllosomas, indicating this character is not diagnostic to separate these two species.

Abstract: A sharp genetic break separates Atlantic from Indo-Pacific bigeye tuna (Thunnus obesus) populations, as the frequencies of two major mitochondrial (mt) DNA types (a and b) found in this species are different across the tip of southern Africa. The level of nucleotide divergence between mtDNA types a and b is of the same order as that between reproductively isolated taxa. To further investigate the genetic structure of bigeye tuna over its distribution range and in the contact zone off southern Africa, bigeye tuna samples collected between 1992 and 2001 (including samples from a previous mtDNA survey) were characterized for four nuclear DNA loci and for mtDNA. Nuclear markers did not support the hypothesis that a and b mitochondria characterize sibling species. Significant allele-frequency differences at one intronic locus (GH2) and one microsatellite locus (l208) were found between Atlantic and Indo-Pacific samples, although the level of nuclear genetic differentiation (Weir and Cockerham�s θ=0.025 to 0.042) was much lower than in mtDNA (θ=0.664 to 0.807). Probabilistic Bayesian assignment of individualsto a population confirmed that southern African bigeye tuna samples represent a simple mixture of individuals from Atlantic and Indian stocks that do not interbreed, with a higher contribution from Indian Ocean individuals (about 2/3 vs. 1/3).

Abstract: Genetic population structure of Atlantic and Mediterranean albacore Thunnus alalunga was investigated using nucleotide sequence variations of the glucose�6�phosphate dehydrogenase gene intron (G6PD) and the mitochondrial DNA (mtDNA) D�loop region (Dloop). Restriction analysis using Ase I digestion detected two major restriction types (A and B) at the Dloop locus with strong frequency differences between Atlantic and Mediterranean samples. Thirty�six individuals of 100 Mediterranean albacore were of the B type whereas no B type individuals were found in the Atlantic samples (n = 102). Phylogenetic analysis using nucleotide sequence data of the Dloop locus indicated that the B type lineage recently arose from the ancestral A lineage in the Mediterranean Sea and has not dispersed into the Atlantic Ocean. The frequencies of two alleles (L and S) at the G6PD locus were significantly different between the samples from the Atlantic (L = 0·495) and the Mediterranean (L = 0·725), but no significant heterogeneity was observed between mtDNA�A and �B types of the Mediterranean sample. These molecular data indicate that gene flow between the Atlantic and Mediterranean albacore populations have been considerably restricted and strongly suggest these populations should continue to be treated as two distinct management units.