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The anatomy of the gas bladder of Diodontidae (porcupinefishes) and Tetraodontidae (pufferfishes) was studied on the basis of dissections and magnetic resonance imaging. Among the examined taxa of Tetraodontiformes, only puffers and porcupinefishes possess a thick walled and dorsally U-shaped or crescent-moon-shaped gas bladder. In the tetraodontid genus Lagocephalus the gas bladder is reduced to a rudiment. The species belonging to the genera Canthigaster, Arothron, and some species of Tetraodon differ in the positioning of their crescent-moon-shaped gas bladder. These observations confirm the close relationship of: (i) Diodontidae and Tetraodontidae and (ii) Canthigaster, Arothron, and some species of Tetraodon. The heterogeneity of the genus Tetraodon is supported by the gas bladder morphology, as previously suggested by molecular studies.

Recent studies suggest that egg activation in mammals is triggered by a sperm-specific phospholipase C, PLCzeta. In other vertebrate species such as medaka fish, chickens, and quail, PLCzeta is also expressed as a testis-specific mRNA. Functional studies suggest that PLCzeta plays a similar role as a trigger of egg activation in these species. Here, we report the identification of PLCzeta orthologues in pufferfish species Takifugu rubripes (Fugu) and Tetraodon nigroviridis (Tetraodon). Unexpectedly in these species PLCzeta is expressed not in the testis, but in ovary and brain. Injection of pufferfish PLCzeta copy ribonucleic acid (cRNA) into mouse eggs failed to trigger calcium oscillations, unlike medaka PLCzeta cRNA. Our findings provide the first evidence that PLCzeta may be expressed in the egg, rather than the sperm, in some vertebrate species, and that its mechanism of action and physiologic role at fertilization may differ in different vertebrate species.

Summary The Toll-like receptor (TLR) family is a pathogen-associated molecular pattern recognition receptor family that plays key roles in the activation of innate immune system. As an important member of TLRs, TLR1 is involved in the recognition of triacylated lipoproteins and mycobacterial products by binding to TLR2 to form a heterodimer. Although partial sequences of TLR1 has been predicted in Fugu and Danio by bioinformatics, the molecular cloning and characterization of this gene in fish still remains largely unclear. Here we report the full-length sequence of TLR1 gene in the pufferfish (Tetraodon nigroviridis), providing experimental evidence to its existence in fish. The cloned Tetraodon TLR1 (TnTLR1) cDNA exhibited 2587 bp in length and contains a 122-bp 5′ untranslated region (UTR), a 2391-bp open reading frame and a 74-bp 3′ UTR. Expression of TLR1 transcripts in most selected tissues were detected at lower levels; however, the transcripts in spleen was markedly increased (threefold) after stimulation with lipopolysaccharide, indicating that TLR1 in fish may be involved in the inflammatory responses to bacterial endotoxin or Gram-negative bacteria infection. Gene organization of TnTLR1 is similar to mammalian and avian, and a high degree of conservation of chromosome synteny exists between them. The protein sequence shares 33.7–77.3% identity with known TLR1 members. Phylogenetic analysis grouped TnTLR1 with other fish TLRs on a separated branch, excluded from mammalian and avian TLR1s. TnTLR1 with 796 amino acids has a calculated molecular mass of 90.69 kDa and a theoretical pI of 8.43. Structurally, TnTLR1 has a Toll/interleukin-1 receptor (TIR) domain, a transmembrane domain and leucine-rich repeat domain which are the hallmarks of TLR family. Phylogenetic analysis provided evidence that the TnTLR1 was very close to mammalian TLR1, TLR6 and TLR10 and seemed to be their common ancestor. This report of the full-length TLR1 gene in Tetraodon will provide us further insights into the study of both function and evolution of fish TLRs as a whole.

The Wnt family of growth factors regulate many different aspects of embryonic development. Assembly of the complete mouse and human genome sequences, plus expressed sequence tag surveys have established the existence of 19 Wnt genes in mammalian genomes. However, despite the importance of model vertebrates for studies in developmental biology, the complete complement of Wnt genes has not been established for nonmammalian genomes. Using genome sequences for chicken (Gallus gallus), frog (Xenopus tropicalis), and fish (Danio rerio and Tetraodon nigroviridis), we have analyzed gene synteny to identify the orthologues of all 19 human Wnt genes in these species. We find that, in addition to the 19 Wnts observed in humans, chicken contained an additional Wnt gene, Wnt11b, which is orthologous to frog and zebrafish Wnt11 (silberblick). Frog and fish genomes contained orthologues of the 19 mammalian Wnt genes, plus Wnt11b and several duplicated Wnt genes. Specifically, the Xenopus tropicalis genome contained 24 Wnt genes, including additional copies of Wnt7-related genes (Wnt7c) and 3 recent Wnt duplications (Wnt3, Wnt9b, and Wnt11). The Danio rerio genome contained 27 Wnt genes with additional copies of Wnt2, Wnt2b, Wnt4b, Wnt6, Wnt7a, and Wnt8a. The presence of the additional Wnt11 sequence (Wnt11b) in the genomes of all ancestral vertebrates suggests that this gene has been lost during mammalian evolution. Through these studies, we identified the frog orthologues of the previously uncharacterized Wnt2, Wnt3, Wnt9a, Wnt9b, Wnt10a, and Wnt16 genes and their expression has been characterized during early Xenopus development. Developmental Dynamics 236:1249 –1258, 2007. © 2007 Wiley-Liss, Inc.

Summary In the present study, 25 195 BAC ends for channel catfish (Ictalurus punctatus) were sequenced, generating 20 366 clean BAC-end sequences (BES), with an average read length of 557 bp after trimming. A total of 11 414 601 bp were generated, representing approximately 1.2% of the catfish genome. Based on this survey, the catfish genome was found to be highly AT-rich, with 60.7% A+T and 39.3% G+C. Approximately 12% of the catfish genome consisted of dispersed repetitive elements, with the Tc1/mariner transposons making up the largest percentage by base pair (4.57%). Microsatellites were detected in 17.5% of BES. Catfish BACs were anchored to the zebrafish and Tetraodon genome sequences by BLASTN, generating 16% and 8.2% significant hits (E

To initiate QTL studies in the nonmodel fish Cottus gobio we constructed a genetic map based on 171 microsatellite markers. The mapping panel consisted of F1 intercrosses between two divergent Cottus lineages from the River Rhine System. Basic local alignment search tool (BLAST) searches with the flanking sequences of the microsatellite markers yielded a significant (e

We performed an intensive search on sequence databases to identify orthologues of ASIP and AGRP peptides in a number of different species, revealing a number of genomic fragments coding for the C-terminal part of agouti-related motifs, different from annotated peptide sequences, including one fragment from chicken, two from zebrafish, two from Fugu (Takifugu rubripes), and three from Tetraodon (Tetraodon nigroviridis). We have thus shown for the first time that both AGRP and ASIP genes exist in many species in "lower vertebrates" and were most probably present in early stages of vertebrate evolution.

The highly conserved Groucho/Tle gene family has widespread functions during embryonic development and in adults. For mammalians, four full-length Tle paralogues are known, whereas the whole spectrum of this gene family in fish species has not been analysed yet. Most detailed data exist for medaka, where 3 Tle genes have been described, Tle1, Tle3, and Tle4. We now isolated 3 additional Tle genes from the medaka genome. Sequence analysis identifies these genes as Tle2a, Tle2b, and Tle3b. Database searches of genomic sequences revealed an identical set of Tle paralogues being present in distantly related fish species, indicating duplicated Tle2 and Tle3 genes for the complete teleost lineage. Like the previously analysed medaka Tle genes, the three new genes show a broad expression pattern during embryogenesis. Nevertheless, a detailed comparison of all six Tle genes reveals critical differences in certain aspects of their expression pattern. In particular, we concentrated on the activity of Tle genes during ear development and found Tle2a and Tle2b expressed in this sensory organ.

Steroid dehydrogenases regulate the access of active steroids to their receptors. In particular, 11beta-hydroxysteroid dehydrogenase-type 1 (11beta-HSD1) and 11beta-HSD2 regulate the levels of glucocorticoids, such as cortisol, and 17beta-HSD1 and 17beta-HSD2 regulate the levels of androgens and estrogens. Human 11beta-HSD1 and 11beta-HSD2 are distant homologs, with less than 25% amino acid sequence identity, as are human 17beta-HSD1 and 17beta-HSD2. In contrast, human 11beta-HSD2 and 17beta-HSD2 are close homologs, with about 43% sequence identity. Until recently, deciphering early events in the evolution of 11beta-HSD2 and 17beta-HSD2 was difficult because only mammalian sequences were available. The completely sequenced Takifugu, Tetraodon and medaka genomes and the almost completed zebrafish genome provide an opportunity to investigate the evolution of 11beta-HSD2, 17beta-HSD2, and 11beta-HSD1. Unexpectedly, a search of the Takifugu, Tetraodon and medaka genomes only found an ortholog to 11beta-HSD2 and none to 17beta-HSD2, while the zebrafish genome contains orthologs of both enzymes. This suggests that 17beta-HSD2 was lost in teleosts after the divergence of zebrafish and medaka. Also unexpectedly, searches with 11beta-HSD1 only identified several fish 11beta-HSD3s, as well as an ortholog in Ciona, indicating that 11beta-HSD3 is the ancestor of 11beta-HSD1.

BACKGROUND In order to understand the evolutionary significance of single triploids among the mostly diploid Poecilia formosa we have developed a simple, noninvasive technique for DNA content and ploidy determination. METHODS From dorsal fin clips of 14 different fish species single cell suspensions were obtained by chopping the material in 2.1% citric acid/0.5% Tween20, passing it through a 0.6-gauge needle and incubating it for 20 min at room temperature (RT) with gentle agitation. After overnight fixation in 70% ethanol, the cells were treated with 1ml 0.5% pepsin/0.1 M HCl for 15 min at RT before adding DAPI to a final volume of 2 ml. The cells were stained for 1–3 h and then analyzed by flow cytometry. RESULTS We obtained good measurements with CVs ranging from 1.23% to 3.36%. The poeciliid species measured contain from 1.6 to 2.0 pg/nucleus, Oryzias latipes (Medaka) exhibits a nuclear DNA content of 2.2 pg, Danio rerio (zebrafish) 4.6 pg, Tetraodon fluviatilis (freshwater fugu) 0.70 pg. All values except zebrafish are in good agreement with the literature. CONCLUSIONS The identification of living specimens of different ploidy for breeding experiments, behavioral studies and tissue transplantations is now made possible. With slight modifications the method can be extended to a field technique, providing therefore a useful tool for a variety of researchers. Cytometry 39:91–95, 2000 © 2000 Wiley-Liss, Inc.