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Using comparative cytogenetic techniques, we characterized the chromosomes of fishes from the family Bovichtidae, the basal lineage of the largely Antarctic suborder Notothenioidei. We focused on three Sub-Antarctic species of the genus Bovichtus that differ greatly in their circumpolar distributions: B. diacanthus (Tristan da Cunha Island Group), B. variegatus (New Zealand) and B. angustifrons (Tasmania). Chromosomes were analyzed both by conventional karyotyping and by cytogenetic mapping of ribosomal genes using fluorescence in situ hybridization. The three species displayed a strongly conserved karyotype consisting entirely of telocentric chromosomes (diploid number = 48; Fundamental Number = 48), in agreement with our previously published hypothesis that the bovichtid karyotype is the basal state for notothenioid fishes. The chromosomal distribution of ribosomal genes differed from those of most notothenioid species studied to date, with the 45S and 5S genes separated on two different chromosome pairs. Separation of two classes of ribosomal genes is the most widespread condition in teleosts, including the bovichtids. Most notothenioid lineages on the other hand exhibit a derived consolidation of these genes.

The study of globin genes has greatly contributed to the understanding of universal mechanisms of gene evolution and of gene expression regulation. In Notothenioidei, the dominant suborder of Antarctic fish, the investigation of the organization of globin genes is particularly interesting in order to characterize the evolutionary mechanisms which led to the hemoglobin loss of the family Channichthyidae. The Antarctic fish Trematomus newnesi has an oxygen transport system with unique characteristics among notothenioids. The cloning and sequence analysis of the ?- and ?-globin cDNAs of the major hemoglobin of T. newnesi, with the determination of the linkage of the relative genes, is herewith reported. The orientation of T. newnesi Hb 1 ?- and ?-globin genes was head-to-head, corresponding to the classical organization of fish globin genes. Seven of the eight notothenioid families are red-blooded but have reduced erythrocyte number and hemoglobin multiplicity; all 16 species of the eighth family (Channichthyidae) are completely devoid of hemoglobin (white-blooded icefish) and represent a novel and unique phenotype in the evolutionary history of fish and vertebrates. Therefore, studies have aimed at evaluating the pathway of genomic change and the mechanisms that have led to the loss of hemoglobin expression by the white-blooded icefish. Adult and embryonic/juvenile gene complexes from a closely related, red-blooded notothenioid species were isolated and characterized. The results indicate that in channichthyids the adult ?-globin gene is absent and that the inability to express hemoglobin in these fishes might have arisen from a large-scale deletional event.

The suborder Notothenioidei dominates the Antarctic ichthyofauna. The non-Antarctic monotypic family Pseudaphritidae is one of the most primitive families. The characterization of the oxygen-transport system of euryhaline Pseudaphritis urvillii is herewith reported. Similar to most Antarctic notothenioids, this temperate species has a single major hemoglobin (Hb 1, over 95% of the total). Hb 1 has strong Bohr and Root effects. It shows two very uncommon features in oxygen binding: At high pH values, the oxygen affinity is exceptionally high compared to other notothenioids, and subunit cooperativity is modulated by pH in an unusual way, namely the curve of the Hill coefficient is bell-shaped, with values approaching 1 at both extremes of pH. Molecular modeling, electronic absorption and resonance Raman spectra have been used to characterize the heme environment of Hb 1 in an attempt to explain these features, particularly in view of some potentially important nonconservative replacements found in the primary structure. Compared to human HbA, no major changes were found in the structure of the proximal cavity of the alpha-chain of Hb 1, although an altered distal histidyl and heme position was identified in the models of the beta-chain, possibly facilitated by a more open heme pocket due to reduced steric constraints on the vinyl substituent groups. This conformation may lead to the hemichrome form identified by spectroscopy in the Met state, which likely fulfils a potentially important physiological role.

On the basis of histological examination, we present the first description of oogenesis in H. spinosus. Eight stages of oocyte development were identified using morphological and histochemical criteria. The development of the oocytes is synchronous, and two clutches grow simultaneously in the ovary. Spawning takes place during the end of summer and the beginning of autumn. Absolute fecundity is low, ranging from 787 to 1504 oocytes. The low fecundity is probably related to parental behaviour. Our data indicate that the features of the reproductive biology of H. spinosus, in spite of its unusual environment, are similar to those observed in other teleost fish. Fil: Van Der Molen, Silvina. Universitat Autònoma de Barcelona; España. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico; Argentina Fil: Matallanas, Jesús. Universitat Autònoma de Barcelona; España

Species of the suborder Notothenioidei dominate the fish fauna of coastal Antarctic waters. Members of one notothenioid family, Channichthyidae (Antarctic icefishes), are unique among all vertebrates in lacking the circulating oxygen-binding protein hemoglobin. Icefish species also do not uniformly express the intracellular oxygen-binding protein myoglobin (Mb) in their oxidative muscles. Our laboratory previously characterized the pattern of cardiac Mb expression in 13 of the 16 known icefish species. In this paper, we complete the survey of cardiac Mb expression among all 16 known species of icefishes. Using PAGE and immunoblot analyses, we demonstrate that both Channichthys rhinoceratus and Cryodraco atkinsoni express Mb in heart ventricle, while Champsocephalus esox does not express the protein. We report Mb gene sequences from Channichthys rhinoceratus and Champsocephalus esox genomic DNA. The Mb gene of C. esox contains the identical 5-bp duplication/insertion to that observed in congeneric Champsocephalus gunnari, a species that also does not produce Mb. This duplication in exon 2 of the Champsocephalus spp. gene causes a shift in reading frame at a position normally encoding for amino acid 91 and also results in a premature stop codon, thus disrupting translation of the normal protein. Thus, 6 of the 16 known icefish species do not express cardiac Mb. These results confirm earlier conclusions that losses of Mb expression have occurred via at least four independent events during the evolution of the icefish family. Extreme similarity of Mb genes in Champsocephalus congeners further suggests recent speciation despite early divergence of this group from the lineage leading to more derived icefishes.

Analysis of the molecular properties of proteins extracted from organisms living under extreme conditions often highlights peculiar features. We investigated by UV-visible spectroscopy and X-ray crystallography the oxidation process, promoted by air or ferricyanide, of five hemoglobins extracted from Antarctic fishes (Notothenioidei). Spectroscopic analysis revealed that these hemoglobins share a common oxidation pathway, which shows striking differences from the oxidation processes of hemoglobins from other vertebrates. Indeed, simple exposure of these hemoglobins to air leads to the formation of a significant amount of the low-spin hexacoordinated form, denoted hemichrome. This hemichrome form, which is detected under a variety of experimental conditions, can be reversibly transformed to either carbomonoxy or deoxygenated forms with reducing agents. Interestingly, the spectra of the fully oxidized species, obtained by treating the protein with ferricyanide, show the simultaneous presence of peaks corresponding to different hexacoordinated states, the aquomet and the hemichrome. In order to assign the heme region state of the alpha and beta chains, the air-oxidized and ferricyanide-oxidized forms of Trematomus bernacchii hemoglobin were crystallized. Crystallographic analysis revealed that these forms correspond to an alpha(aquomet)-beta(bishistidyl-hemichrome) state. This demonstrates that the alpha and beta chains of Antarctic fish hemoglobins follow very different oxidation pathways. As found for Trematomus newnesi hemoglobin in a partial hemichrome state [Riccio, A., Vitagliano, L., di Prisco, G., Zagari, A. & Mazzarella, L. (2002) Proc. Natl Acad. Sci. USA99, 9801-9806], the quaternary structures of these alpha(aquomet)-beta(bishistidyl-hemichrome) forms are intermediate between the physiological R and T hemoglobin states. Together, these structures provide information on the general features of this intermediate state.

In the subzero shelf waters of Antarctica, fishes of the perciform suborder Notothenioidei dominate the fish fauna and constitute an adaptive radiation and a species flock. The 16 species of dragonfishes of the family Bathydraconidae live from surface waters to nearly 3,000 m and have the greatest overall depth range among noto- thenioid families. We examined the anatomy and histol- ogy of the brain, retina, and cephalic lateral line system of nine bathydraconid species representing 8 of the 11 known genera. We evaluate these data against a cla- dogram identifying three clades in the family. We provide a detailed drawing of the brain and cranial nerves of Gymnodraco acuticeps and Akarotaxis nudiceps. Bathy- draconid brain morphology falls into two categories. Brains of most species are similar to those of generalized perciforms and some basal notothenioids (Class I). How- ever, brains of deep-living bathydraconids (members of the tribe Bathydraconini minus Prionodraco) have a re- duced telencephalon and tectum that renders the neural axis visible — the stalked brain morphology (Class II). All bathydraconids have duplex (rod and cone) retinae but there is considerable interspecific variation in the ratio of cones:rods and in the number of cells in the internal nuclear layer. Retinal histology reflects habitat depth but is not tightly coupled to phylogeny. Although the deep- living species of Bathydraconini have rod-dominated ret- inae, the retinae of some sister species are photopic. An expanded cephalic lateral line system is also characteris- tic of all members of the Bathydraconini as exemplified by Akarotaxis. This morphology includes large lateral line pores, wide membranous canals, hypertrophied canal neu- romasts, and large anterodorsal lateral line nerves, emi- nentia granulares, and crista cerebellares. The saccular otoliths are also enlarged in members of this tribe. Neural diversification among bathydraconids on the Antarctic shelf has not involved the evolution of sensory specialists. Brain and sense organ morphologies do not approach the specialized condition seen in primary deep-sea fishes or even that of some secondary deep-sea fishes including sympatric non-notothenioids such as liparids (snailfishes) and muraenolepidids (eel cods). The brains and sense organs of bathydraconids, including the deep-living spe- cies, reflect their heritage as perciform shorefishes. J. Morphol. 258:130 -150, 2003. © 2003 Wiley-Liss, Inc. Fish of the perciform suborder Notothenioidei dominate the diversity, abundance, and biomass in the subzero waters of the Antarctic continental shelf. Here the 96 currently recognized Antarctic species form an adaptive radiation (Eastman, 1993, 2000; Clarke and Johnston, 1996) and possibly a species flock (Eastman and Clarke, 1998; Eastman and McCune, 2000). A species flock is an assemblage of a disproportionately high number of closely re- lated species which evolved rapidly within a circum- scribed area where most species are endemic (Rib- bink, 1984). With little competition from a sparse non-notothenioid fauna, notothenioids underwent considerable morphological and ecological diversifi- cation. Although they are a benthic group and lack a swim bladder, one hallmark of the notothenioid ra- diation has been the evolutionary alteration of buoy- ancy enabling the utilization of unfilled niches in the water column, especially those involving pelagic or partially pelagic zooplanktivory and piscivory. The Bathydraconidae (dragonfishes), one of the five Antarctic notothenioid families, includes 16 spe- cies and 11 genera, many of these monotypic (East- man and Eakin, 2000). Bathydraconids are phyleti- cally derived within the Notothenioidei and are the sister group of the hemoglobin-less icefishes of the family Channichthyidae. Some morphological (Hast- ings, 1993) and molecular (Bargelloni et al., 2000) datasets yield cladograms suggesting that the Bathydraconidae is paraphyletic, but other molecu- lar data support neither paraphyly nor monophyly

Antarctic fish of the suborder Notothenioidei represent one of the most notable examples of adaptive radiation in the marine environment. The evolutionary relationships between and within the eight families of this suborder have been well established by numerous studies, whereas the microevolutionary processes of notothenioid species remain largely unexplored. In the present paper we investigated the evolutionary relationships between three closely related species of the genus Chionodraco (family Channichthyidae), namely Chionodraco hamatus, Chionodraco rastrospinosus, and Chionodraco myersi by analysing portions of the mitochondrial genome (D-loop and 16S rRNA). The taxonomic status of C. hamatus and C. rastrospinosus as separate species has been questioned because of the limited number of key morphological characters that distinguish these two taxa. Our results, based on the analysis of several specimens belonging to both morphological groups revealed a small genetic differentiation among haplotypes, however, a clear separation between the two nominal species emerged since all individuals of each of the two taxa clustered together in distinct monophyletic groups. C. myersi appeared more distantly related in the phylogenetic analysis. For one species, C. hamatus, sampling was carried out at three different geographic locations in the area of the Ross Sea and Weddell Sea. The results showed that the partition of the genetic variation within this species is not compatible with the hypothesis of panmixia as gene flow between populations was significantly reduced.

8 pages, 5 figures, 5 tables, An ecomorphological study was performed on ten species of the family Artedidraconidae, the most benthic of the suborder Notothenioidei. These species are sympatric on the shelf of the Weddell Sea. The results show that the four genera comprising the family (Artedidraco, Dolloidraco, Histiodraco and Pogonophryne) can be differentiated by just a few morphometric features of their sensory organs and mouth. The genera were also clearly defined by the composition of their diet (benthic and epibenthic), the size of their prey, and their bathymetric distribution. Comparison of the morphological and ecological data shows a very close connection between sensory organs development, the mouth and bathymetric distribution. The morphological and ecological divergence observed can be explained as the result of the rapid adaptive radiation of the artedidraconids. The results confirm that characteristics of the sensory organs and alimentary structures are very suitable for the ecomorphological study of fishes, The authors wish to thank Dr Wolf Arntz for his invitation to participate in the ANT XV/III survey. We express our gratitude to the scientific fishing teams, Drs A. Schröeder and R. Knust, for their help during this cruise, Drs Jordi Lleonart, Beatriz Morales-Nin and Eduardo Balguerias for scientific support, and Drs J. Montgomery and J.T. Eastman for their valuable comments. We also acknowledge the skill and professionalism of the RV Polarstern crew members. This work was undertaken as part of the SCAR EASIZ programme, and was supported by the Alfred Wegener Institute and the Spanish Antarctic Programme

In the high-latitude shelf waters of Antarctica, fishes in the perciform suborder Notothenioidei dominate the fish fauna and constitute an adaptive radiation and a species flock. The 25 species of notothenioid plunderfishes, comprising four genera of the family Artedidraconidae, contribute substantially to fish species diversity on the high Antarctic shelf. A mental barbel is an autapomorphy for the family. Dolloidraco longedorsalis is the most abundant artedidraconid at depths over 400 m in these waters. In this article we present the anatomy and histology of the brain and special sense organs of Dolloidraco and compare it to the brains of other artedidraconids, closely related harpagiferids, and more generally to other notothenioids. We provide a detailed drawing of the brain and cranial nerves. The brain of Dolloidraco is simple, without external hypertrophy of sensory or motor regions, but contains several unusual features associated with the ventricular system and CSF, including well-developed circumventricular organs, subependymal expansions, and subarachnoid cisterns; and a ventricle in the corpus cerebellum. The brain of Dolloidraco also contains a lobed chief sensory nucleus of the trigeminal nerve that is correlated across species with barbel length. The eyes are large and contain a small choroid rete, a structure previously thought to be absent from members of this family. We document the histology of the duplex retina, olfactory apparatus, cutaneous taste buds, and barbel musculature and innervation. We discuss the role of pedomorphy in producing simplified brain morphologies. We consider the possibility that Dolloidraco is a somatosensory specialist—an unusual feature among vertebrates—and decide that this is unlikely. J. Morphol. 255:358–377, 2003. © 2003 Wiley-Liss, Inc.

A total of 1,103 inshore notothenioid fish were caught by means of trammel-nets in 4 sites surrounding Cierva Point (Moss Island 1; Moss Island 2; Sterneck Island; Leopardo Island), Danco Coast, West Antarctic Peninsula, during February and March 2000. The families Nototheniidae, Channichthyidae and Bathydraconidae were represented in the samples, Notothenia coriiceps being the dominant fish of the area. Gobionotothen gibberifrons and Trematomus newnesi followed in importance. In general, the fish sampled agreed in terms of number and mass with those of the South Shetland Islands area, except for a marked higher occurrence of G. gibberifrons in the Danco Coast. This supports the hypothesis that the commercial fishery around the South Shetland Islands at the end of the 1970s was responsible for the decrease in the inshore population of G. gibberifrons in that area during the last 17 years. Information on morphometry, reproduction and diet of the fish species caught is provided., Facultad de Ciencias Médicas

El continente Antártico, la región más meridional de la tierra, despliega una serie de características únicas y distintivas. Las temperaturas de las aguas del Océano Antártico exhiben poca variación estacional. A pesar de la alta estabilidad de la temperatura en la columna de agua, cercana al punto de congelación del agua de mar, existe una marcada estacionalidad en otros factores como la formación de capas de hielo y períodos de luz, y sujeta a estos cambios, una extrema estacionalidad en la producción primaria. Estas condiciones parecen haber favorecido a una serie de adaptaciones singulares en la fauna íctica, por ejemplo: metabolismo bajo, madurez tardía, y huevos con abundante vitelo. En este contexto, los peces antárticos constituyen un claro ejemplo de adaptación al medio. Las aguas de plataforma y talud están dominadas, en términos de biomasa y diversidad, por un solo suborden endémico: Notothenioidei. Como en la mayoría de los vertebrados, la reproducción de los peces es un fenómeno cíclico. En los teleósteos en general, esta periodicidad es anual y se rige por las variaciones estacionales de las condiciones ambientales. Se ha sugerido, como norma general, un acoplamiento espacio-temporal de los ciclos reproductivos con los ciclos de producción primaria. Sin embargo, algunas de las especies antárticas parecen desviarse de este criterio. El desacoplamiento de los ciclos reproductivos con las fluctuaciones temporales de la producción primaria podría representar una ventaja evolutiva para algunas especies, dado que su éxito reproductivo constituiría un suceso independiente de las variaciones anuales.Para una comprensión de la biología de los peces que viven en las peculiares condiciones que impone el Océano Austral, es de gran interés el estudio de sus adaptaciones. En los Notothenioidei, la reproducción parece ser un parámetro clave de estas adaptaciones. Dado que han experimentado una gran radiación adaptativa, es de suma importancia apreciar si las estrategias reproductivas que presentan sufrieron también cambios a lo largo de su historia evolutiva. Es decir, examinar si los parámetros que describen la biología reproductiva de las especies, presentan alguna particularidad relacionada con el resto de las adaptaciones que presenta este grupo.El objetivo general de este trabajo es reconstruir el ciclo biológico de varias especies del suborden Notothenioidei, a partir del estudio histoquímico de la ovogénesis, interpretando luego su papel ecológico y evolutivo. Los parámetros reproductivos vinculados a la duración del período reproductivo, fecundidad, madurez sexual, desove y estacionalidad fueron analizados en ejemplares pertenecientes a las familias Channichthyidae, Harpagiferidae, Artedidraconidae y Bathydraconidae (Pisces: Perciformes).Los resultados obtenidos reflejan que las diferencias en las estrategias reproductivas que presenta este grupo se basan principalmente en el aprovechamiento diferencial de los recursos, ya sean espaciales y temporales o tróficos. Es decir que, a pesar de la alta diversidad fenotípica que muestra este grupo en lo referente a estrategias reproductivas, los Notothenioidei no han desarrollado adaptaciones específicas en este sentido, que los distingan de otros grupos de teleósteos., The Antarctic continent shows the most extreme environmental conditions. The stable temperatures of the Southern Ocean are at or near the freezing point of sea water. Although antarctic waters exhibit low seasonal variability in temperature, light, development of sea-ice and primary production show marked seasonal fluctuations. Thus the antarctic fauna is subject to extreme environmental conditions and exhibit particular adaptations. The major features of the biology of antarctic marine poikiloterms are low metabolism, seasonal growth rates, large yolky eggs and delayed maturity. Many authors also suggested that reproductive cycles appear to be closely linked to the primary production cycleThe antarctic fauna is smaller and much less diverse on a higher taxonomic level (order, family) than might be expected given the considerable age and large area of the ecosystem. The shelf and slope waters are dominated both in therms of species and biomass by a single, endemic perciform suborder, the Notothenioidei. Some of the common characteristics of this suborder are: slow growth, prolonged gametogenesis, low fecundity and large yolky eggs. As in most vertebrates, reproduction in fish is a cyclic phenomenon, generally in Teleost fishes this periodicity is annual and it is influenciated by environmental changes. However, some antarctic species show a different pattern and exhibit uncoupled reproductive cycles. It is possible that there could be selective advantages to be gained by species that are able to be independent of the production cycle and thereby avoid large variations in breeding success from year-to-year changes in the environment. To a better understanding of the biology of fish living in the extreme conditions of marine polar environment it is of great interest the study of the antarctic fauna adaptations. Reproduction is a key parameter in the study of notothenioidei adaptations. Describing oogenesis stages and sexual maturation, spawning characteristics and fecundity, the life cycle of these species can be reconstructed, giving basis knowledge about their biology.The main objective of this project is to describe the characteristics of Notothenioiids reproductive cycle, its relation with environmental parameters and its intraspecific variation. Based in oogenesis histochemical analysis, reproductive parameters such as fecundity, spawning and seasonality were evaluated in 9 species belonging to the families Channichtyidae, Artedidraconidae, Harpagiferidae and Bathydraconidae (Pisces: Perciformes). Results reveal that the oocyte development events are very similar to those described for other notothenioids and for most other Teleost. Reproductive strategies in notothenioids differ mainly in the differential exploitation of temporal or spatial resources. Thus, these observations suggest that specialisation in Antarctic fish are more related to ecological diversification than to any kind of adaptation concerning oogenesis.

The blood of Antarctic icefishes (family Channichthyidae, suborder Notothenioidei) is completely devoid of hemoglobin. Icefishes have developed compensatory adaptations that reduce oxygen demand and enhance oxygen transport. Oxygen delivery to tissues occurs by carrying the gas physically dissolved in the plasma. To evaluate the evolutionary pathway leading to the icefish hemoglobinless phenotype, the adult and embryonic/juvenile gene complexes from a closely related, red-blooded notothenioid species were isolated and characterized. The hybridization pattern of notothenioid adult globin cDNAs showed that the genomes of three icefish species retain transcriptionally inactive alpha1-globin-related DNA sequences, which are identical truncated variants of the alpha1-globin gene of the red-blooded fish, containing part of intron 2, all of exon 3, and the 3'-untranslated region. The icefish genomes have no beta-globin genes. Furthermore, Southern blots of genomic DNA from red- and white-blooded (two species) notothenioids, probed with fragments of the genes flanking the ends of the embryonic/juvenile complex, indicated that icefishes have also lost embryonic/juvenile globin genes. It is proposed that inability to express hemoglobin arose from a single, large-scale deletional event, which removed all icefish globin genes with the exception of the 3' end of alpha1.

The Arctic fish Anarhichas minor, a benthic sedentary species, displays high hemoglobin multiplicity. The three major hemoglobins (Hb 1, Hb 2, and Hb 3) show important functional differences in pH and organophosphate regulation, subunit cooperativity, and response of oxygen binding to temperature. Hb 1 and Hb 2 display a low, effector-enhanced Bohr effect and no Root effect. In contrast, Hb 3 displays pronounced Bohr and Root effects, accompanied by strong organophosphate regulation. Hb 1 has the beta (beta(1)) chain in common with Hb 2; Hb 3 and Hb 2 share the alpha (alpha(2)) chain. The amino acid sequences have been established. Several substitutions in crucial positions were observed, such as Cys in place of C-terminal His in the beta(1) chain of Hb 1 and Hb 2. In Hb 3, Val E11 of the beta(2) chain is replaced by Ile. Homology modeling revealed an unusual structure of the Hb 3 binding site of inositol hexakisphoshate. Phylogenetic analysis indicated that only Hb 2 displays higher overall similarity with the major Antarctic hemoglobins. The oxygen transport system of A. minor differs remarkably from those of Antarctic Notothenioidei, indicating distinct evolutionary pathways in the regulatory mechanisms of the fish respiratory system in the two polar environments.

In the framework of the I.C.E.FISH (International Collaborative Effort on Antarctic Fish Adaptive Evolution) project, during the 1998–1999 season at the Italian shore-based Terra Nova Bay Station, the broad fields of adaptation and evolution of the fish suborder Notothenioidei were tackled through the integration of many disciplines. As a representative contribution of I.C.E.FISH, a cytogenetic study of the bathydraconid fish Gymnodraco acuticeps is reported. The Bathydraconidae is a heterogeneous notothenioid taxon whose intra-family relationships are very uncertain. The conventional karyotype and the location of nuclear ribosomal genes, by means of fluorescence in situ hybridisation, contributed to species-specific characterisation and provided the basis for comparative analyses. The karyotype of G. acuticeps consists of 48 chromosomes (2 metacentric, 2 submetacentric and 44 acrocentric); major ribosomal cistrons correspond to a large region in a single chromosome pair. Mapping of the available karyotypic data on a molecular phylogenetic tree provided information on chromosomal diversification during the cladogenesis of the bathydraconids.

The radiation of notothenioid fishes provides an excellent system to explore issues of evolution and adaptation. Most studies emphasize adaptation to the extreme Antarctic environment; however, recent work provides cogent examples of disaptation or evolutionary loss of function. The nature and extent of regressive change is revealed by subsequent adaptive recovery. Ancestral notothenioids were benthic but some became secondarily pelagic through the retention of larval characters. Paedomorphosis has produced detrimental changes in lateral-line sensory systems that have been made good by compensatory adaptation. In the icefish family, compensatory adaptation has followed the loss of the oxygen-binding pigments haemoglobin and myoglobin.

Six postlarval and an adult bathydraconid were collected in Terra Nova Bay, Ross Sea, during January-February 1988 and February 1998, respectively. The adult was identified as Acanthodraco dewitti Skora, 1995 (type locality South Shetland Islands), a species not recorded in the Ross Sea. The postlarvae may be those of A. dewitti. The morphology and pigmentation patterns of the postlarvae are described and compared with those of other bathydraconids.

Published and unpublished data are used to investigate possible mechanisms of species diversification in two key groups of Antarctic organisms: the Notothenioidei and the Euphausiidae. Species distributions are mapped onto molecular phylogenies, and this evidence is interpreted in light of the various ecological and historical factors which characterize the Southern Ocean. The joint effect of diverse agents (vicariance, “jump” dispersal) appears to be determinant in several cases for species divergence. A review of results from population genetics studies, together with new molecular evidence, confirm the importance of physical barriers (oceanographic fronts) in reducing migration, thereby promoting speciation, at least in notothenioids.

Icefish (Notothenioidei, Channichthyidae) are a group of Antarctic fish displaying unique phenotype characteristics. Besides the reported lack of hemoglobin, icefish have a very low metallothionein (MT) content compared to Antarctic red-blooded fish. Despite the low amount of MT protein in icefish, cDNA encoding two MT isoforms could be produced from total hepatic RNA by RT-PCR. The mRNA 5’ end fragments were amplified by 5’-RACE. Steady-state mRNA levels were assessed in both icefish and red-blooded fish by high-stringency hybridization of the MT probe with total RNA. The results showed the presence of large amounts of MT mRNA in icefish liver. A further increase in MT mRNA was observed in Cd-treated icefish which was accompanied by the appearance of large amounts of MT protein. A comparison of the sequences of the 5’ ends of the mRNAs showed the lack of a short motif in the 5’-UTR of only one of the two MT isoforms. On the basis of these results, we suggest that translation of the mRNA encoding an MT isoform is regulated in icefish by specific motifs in the 5’end.

The hematological properties and the oxygen-transport system of the antarctic fish Pleuragramma antarcticum were investigated. Most blood parameters are at the lower end of the range of values known for red-blooded antarctic fish, suggesting a link with the sluggish mode of life of this species. P. antarcticum is the only species of the family Nototheniidae and of the suborder Notothenioidei having three major hemoglobins, which were isolated and fully characterized. The complete amino acid sequence of the α- and β-globin chains was determined. The three hemoglobins showed strong Bohr and Root effects, and their oxygen-binding properties were differently regulated by temperature. None of the three hemoglobins of P. antarcticum can be considered as evolutionary (or larval) remnants. Therefore, this oxygen-transport system is one of the most specialized ever found in fish. The data suggest a strong relationship between hematological/biochemical adaptation and life style.

The monophyly of the antarctic fish suborder Notothenioidei and the monophyly of its earliest family the Bovichtidae have been investigated with 12S and 16S mitochondrial DNA sequences. New data from Cottoperca , Pseudaphritis , Harpagifer and several outgroups, in addition to available sequences, show that the bovichtids are paraphyletic. Pseudaphritis is the sister group of all the non-bovichtid notothenioids. The same results are found from two independent genetic markers, the nuclear 28S rDNA and the 12S and 16S mitochondrial rDNA. This reliably refutes a previous hypothesis that placed Pseudaphritis as the sister group of all the remaining notothenioids (including Cottoperca and Bovichtus ). Bootstrap analyses show that the Notothenioidei are monophyletic (although members of the suborder Trachinoidei have not been surveyed). Subsequent data from hemoglobin composition confirm the present relationships. After discussions between members of the European Science Foundation (ESF) network during its last two meetings, we point out here some fundamental aspects of comparative biology to improve understanding between the physiologist community and phylogeneticists. The most important points are differences in how the concept of homology is used and differences in the consideration of adaptation. When adaptation is evoked or questioned, endless speculations and untestable scenarios are often developed. We strongly advocate the use of phylogenetic trees for testing hypotheses of adaptation (through multiple character mapping). Such a “research program” in comparative biology has the power to improve knowledge because it can potentially lead to new experiments for testing adaptive hypotheses.

A critical list of the Antarctic fish Digenea recently published [1] concerns 36 species. However, a genuine occurrence of one species, Lecithocladium falklandicum, in the Antarctic is doubtful. A validity of another one, Gonocerca lobata, is questionable. These two species are omitted from the present review. On the other hand, new material collected in the Weddell Sea has recently been reported by the present author and a further ten undescribed species were recognized. Their descriptions have been recently published or are in press [2, 3, 4, 5, 6, 7]. Thus 44 species are taken into account. This is not a final number of digeneans occurring in fish in the Antarctic, as strongly contracted and/or damaged undeterminable (even to generic level) specimens of not less than two further species are in the present author’s collection. The presence of further undescribed species is very possible, as only limited data are available for parasites of Antarctic fishes of such important families as Liparididae, Zoarcidae, Murenolepididae and Macrouridae, as well as some Notothenioidei like Artedidraconidae and Bathydraconidae occurring in deep waters. It should be noted that only one species, Lecithochirium whitei, is known as a parasite specific to a bathypelagic fish, Bathylagus antarcticus. All others normally occur in benthic and bentho-pelagic fishes; only a few species and specimens have been reported also from pelagic fishes.

The suborder Notothenioidei represents a moderate radiation of teleosts into various niches, mostly at less than 500 m depths, and adaptation to subzero temperatures [1,2]. The decrease of temperature of the Antarctic continent 38 million years ago (Ma) provoked important species extinction and the dominance of Notothenoidei.

During recent years cytogenetic studies on Antarctic fish have received increasing attention [1–4]. Representatives from all the families of the Notothenioidei suborder have been studied in High Antarctic, SubAntarctic and cold-PeriAntarctic regions (1).

The family Channichthyidae contains 11 genera and 15 species [1] and is considered monophyletic [2,3, 4, 5]. Interrelationships of channichthyids were first investigated by Iwami [2] based on morphological characters.

The perciform suborder Notothenioidei is a highly diversified group of fish inhabiting the waters around the Antarctic continent. This is a unique environment, characterized, especially in the high Antarctic Zone, by temperatures as low as −2 °C, presence of sea ice, and large seasonal fluctuations of the primary production. At the Oligocene-Miocene boundary, about 25–22 million years ago, with the opening of the Drake Passage and the formation of a circumpolar hydrographic barrier, the Polar Front Zone, the Antarctic marine environment became effectively isolated.

The fish fauna around Antarctica is dominated by a single perciform suborder, the Notothenioidei, whose phylogeny has been well defined by traditional and molecular techniques. They have undergone a remarkable adaptive radiation (into a variety of forms from small, sculpin-like to large, hake-like species) that is possibly unique in the marine environment, and which molecular evidence suggests is a relatively recent event [1]. The last few decades have seen the development of extensive scientific and commercial fishing in the Southern Ocean that has provided much data on the growth and reproduction of a wide range of species. In contrast, financial constraints and logistical difficulties have meant that studies into the physiological mechanisms that allow these fishes to successfully exploit this extremely cold and highly seasonal environment have mainly concentrated on a few species, predominantly from Nototheniidae and Channichthyidae [2,3]. Antarctic pelagic fishes are diverse in their origins, coming from bathypelagic, mesopelagic and, of most interest for our purpose, from coastal fish families that have become secondarily adapted to midwater life. However, most species (about 75% of those so far recorded) inhabit benthic or demersal niches, which is thought to reflect the benthic lifestyle of the parent taxa, and helps explain the lack of a functional swimbladder [2]. Those species that are now able to exploit the water column have had to evolve means of adjusting buoyancy, including reduced skeletal ossification and extensive lipid deposits [3].

The frigid waters of the polar oceans delimit the cold extreme for marine life. This is particularly true in the case of the thermally isolated Antarctic Ocean which is perpetually near or at freezing (−1.9 °C) due to the thermal barrier imposed by the Antarctic Circumpolar Current [1]. The most fundamental survival challenge faced by teleost fishes in these waters is a physical one — the threat of being frozen. The body fluids of marine teleosts including polar species are hyposmotic to sea water, 300–600 mOsM [2,3] versus 1000 mOsM, and thus have a higher colligative freezing point than the latter, −0.56 °C to −1.1 °C versus −1.86 °C. By these simple physical considerations alone, freezing death would be unavoidable especially in the presence of ice. Unlike some reptiles and amphibians, fish cannot survive even partial freezing of their body fluids. A number of polar and subpolar fishes had overcome this environmental challenge with a biological solution — they evolved ice-binding antifreeze proteins which enabled them to successfully colonize icy habitats that were otherwise out of their reach. The impact of the evolution of these unique antifreezing proteins on organismal and ecological success is manifested most strikingly in the case of the Antarctic notothenioid fishes — a single teleost suborder (Notothenioidei) that has come to dominate today’s Antarctic fish fauna in terms of species number (∼50%) and biomass (≥90%) [2,4, 5, 6].

Amino acid sequences of alpha- and beta-chains of human hemoglobin and of hemoglobins of coelacanth and 24 teleost fish species, including 11 antarctic and two temperate Notothenioidei, were analyzed using maximum parsimony. Trees were derived for the alpha- and beta-chains separately and for tandemly arranged sequences, using the human and coelacanth sequences as outgroups in all analyses. The topologies of the trees of the alpha- and beta-chains are highly congruent and indicate a specific pattern of gene duplications and gene expression of teleost hemoglobins which has not yet been investigated into more detail. The Notothenioid fish generally contain a single major hemoglobin and often a second minor component. The alpha- and beta-chains of the major components form a monophyletic group in all investigated trees, with the nonantarctic Pseudaphritis as their sister taxon. The minor chains also are a monophyletic group and form an unresolved cluster with the major chains and the hemoglobins of tuna and red gurnard. The Notothenioid families Nototheniidae and Bathydraconidae appear to be paraphyletic.

The concept of species flocks has been central to previous interpretations of patterns and processes of explosive species radiations within several groups of freshwater fishes. Here, molecular phytogenies of species-rich Sebastes rockfishes from the northeastern Pacific Ocean were used to test predictions of null theoretical models that assume random temporal placements of phylogenetic nodes. Similar appraisals were conducted using molecular data previously published for particular cichlid fishes in Africa that epitomize, by virtue of a rapid and recent radiation of species, the traditional concept of an intralacustrine "species flock." As gauged by the magnitudes of genetic divergence in cytochrome b sequences from mitochondrial DNA, as well as in allozymes, most speciation events in the Sebastes complex were far more ancient than those in the cichlids. However, statistical tests of the nodal placements in the Sebastes phylogeny suggest that speciation events in the rockfishes were temporally nonrandom, with significant clustering of cladogenetic events in time. Similar conclusions also apply to an ancient complex of icefishes (within the Notothenioidei) analyzed in the same fashion. Thus, the rockfishes (and icefishes) may be interpreted as ancient species flocks in the marine realm. The analyses exemplified in this report introduce a conceptual and operational approach for extending the concept of species flocks to additional environmental settings and evolutionary timescales.

Background Among the cold-adapted Antarctic notothenioid fishes, the high-latitude bald notothen Pagothenia borchgrevinki is particularly notable as the sole cryopelagic species, exploiting the coldest and iciest waters of the Southern Ocean. Because P. borchgrevinki is a frequent model for investigating notothenioid cold-adaptation and specialization, it is imperative that “omic” tools be developed for this species. In the absence of a sequenced genome, a well annotated reference transcriptome of the bald notothen will serve as a model of gene expression in the coldest and harshest of all polar marine environments, useful for future comparative studies of cold adaptation and thermal responses in polar teleosts and ectotherms. Results We sequenced and annotated a reference transcriptome for P. borchgrevinki, with added attention to capturing the transcriptional responses to acute and chronic heat exposures. We sequenced by Roche 454 a normalized cDNA library constructed from pooled mRNA encompassing multiple tissues taken from environmental, warm acclimating, and acute heat stressed specimens. The resulting reads were assembled into 42,620 contigs, 17,951 of which could be annotated. We utilized this annotated portion of the reference transcriptome to map short Illumina reads sequenced from the gill and liver of environmental specimens, and also compared the gene expression profiles of these two tissue transcriptomes with those from the temperate model fish Danio rerio. From this, we identified a conserved group of 58 GO terms, in which terms related to transcription and its regulation, ubiquitin-protein ligase activity, protein ubiquitination, and protein binding among others are more prevalent in the bald notothen, suggesting the pertinent genes play essential roles in cold temperature functioning. Conclusion We sequenced multiple tissue transcriptomes from native and heat-exposed experimental specimens of the high Antarctic, cryopelagic notothenioid P. borchgrevinki to construct a reference transcriptome. In a proof of concept, we utilized the annotated reference transcriptome to profile the gene expression patterns of gill and liver, and identified a suite of over and under-represented GO terms when compared to the tropical water zebrafish suggesting these functions may be important for surviving in freezing waters. The transcriptome resource from this study will aid future investigations of cold adaptation and thermal response of polar ectothermic species.

Introduction More than any other habitat on earth, Antarctica is a unique natural laboratory, ideal for studying temperature adaptations. Hence special attention will be given to its paleogeography and to the adaptive mechanisms of Antarctic marine organisms. For example some adaptations (freezing avoidance, efficient enzymatic catalysis and cytoskeletal polymer assembly, decreased blood viscosity through reduction or elimination of erythrocytes and haemoglobin) represent a unique character of Antarctic fish and will be examined in detail. Specialisations in haematology and in the oxygen transport system were also developed by other polar and temperate organisms: Arctic mammals (reindeer, musk ox, whale), birds (penguin), reptiles (turtle), crustaceans (krill), cephalopods (squid). We describe the molecular mechanisms of the oxygen transport system in relation to requirements for function at low temperature. It is pertinent to mention the difficulty in establishing consensus on objective criteria to identify a phenotypic trait as an adaptation. Thus, adaptation remains ‘a slippery concept’. The reader will find extensive discussion on this and other issues in two recent reviews (Reeve & Sherman, 1993; Garland & Carter, 1994). The Antarctic In the late Precambrian, 590 million years ago (Ma), Antarctica was the central part of the supercontinent Gondwana, which remained intact for 400 million years, during the Paleozoic and part of the Mesozoic, through the Jurassic; fragmentation began and continued during the Cretaceous. The continental drift took Antarctica to its present position about 65 Ma, at the beginning of the Cenozoic.

In the absence of fossils, the origin of Notothenioidei, a perciform suborder dominating the fish fauna of the Southern Ocean, remains conjectural; some morphoecological evidence suggests relationships to zoarcoids. To test this point we have compared the karyotype morphology and genome size of two species of zoarcids from the Ross Sea to those of one species each of the notothenioid families. Artedidraconidae, Bathydraconidae, Channichthyidae and Nototheniidae from the same region. A karyotype of 48, mostly acrocentric, chromosomes, localization of nucleolar organizers on a pair of small dibrachial chromosomes, a genome size of about 3 pg of DNA, characterize both zoarcids; similar features can be found in the karyology of the notothenioids (especially the Nototheniidae). However, all shared characters appear as plesiomorphic in teleost karyology, which does not help in producing new data on the problem of notothenioid relationships.

Antifreeze glycopeptides (AFGP) have been isolated from the fully pelagic high-Antarctic silverfish Pleuragramma antarcticum of the suborder Notothenioidei (Perciformes). The fishes were caught during the PRV Polarstern expedition EPOS III (Jan-Mar, 1989) in the eastern and southeastern Weddell Sea. Glycoconjugate and amino acid analysis of antifreeze glycopeptides (AFGP) indicate that the glycopeptide structure is identical to the polymers of H2N[Ala-Ala(beta-galactosyl(1--3)-alpha-N- acetylgalactosamine)Thr]nAla-Ala-COOH of previously studied Antarctic notothenioids. The content of AFGPs in P. antarcticum is lower than in other notothenioid fish from the same region. Antifreeze activity shows a maximal hysteresis of 1.19 degrees C at a concentration of 20 mg/ml AFGP. A linear increase in activity of the antifreeze glycopeptides could be demonstrated concomittant with a decreasing ice content. The freezing point of blood serum is -1.9 degrees C.

The Notothenioidei, a perciform suborder of 120 species, dominates the ichthyofauna of the Southern Ocean around Antarctica. Unlike most teleost groups, notothenioids have undergone a corresponding ecological and phyletic diversification and therefore provide an excellent opportunity to study the divergence of the nervous system in an unusual environment. Our goal is to evaluate notothenioid brain variation in light of this diversification. To provide a baseline morphology, we examine the gross morphology and histology of the brain of Trematomus bernacchii, a generalized member of the family Nototheniidae. We then examine the variation in brain gross anatomy (32 species) and histology (10 species) of other notothenioids. Our sample represents about 27% of the species in this group and includes species from each of the six families, as well as species representing diverse ecologies. For comparison we reference the well-studied brains of two species of temperate perciformes (Perca flavescens and Lepomis humilis). Our results show that, in general, notothenioid brains are more similar to the brains of temperate perciforms than to the unusual brains of cave-dwelling and deep-sea fishes. Interspecific variation in gross brain morphology is comparable to that in Old World cyprinids and is illustrated for 17 species. Variation is especially noteworthy in the ecologically and geographically diverse family Nototheniidae. Measurements indicate that sensory regions (olfactory bulbs, eminentia granularis, and crista cerebellaris) exhibit the most pronounced variation in relative surface area. Association areas, including the corpus cerebelli and the telencephalon, exhibit moderate variation in size, shape, and lobation patterns. Regulatory areas of the brain, including the saccus vasculosus and the subependyma of the third ventricle, are also variable. These regions are best developed in species living in the subfreezing water close to the continent. In some species the expanded ependymal lining forms ventricular sacs, not previously described in any other vertebrate. Three species, including two nototheniids (Eleginops maclovinus and Pleuragramma antarcticum) and the only artedidraconid in our sample, have distinctive brains. The unique brain morphology of Pleuragramma is probably related to a sensory (lateral line) specialization for feeding. Within the Nototheniidae, a phyletic effect on cerebellar morphology is evident in the Coriiceps group and in the Pleuragramminae. Neither phyletic position nor ecological factors (water temperature, position in the water column, dietary habits) alone fully explain the pattern of notothenioid brain diversification.

Fish of the suborder Notothenioidei have successfully radiated into the Southern Ocean and today comprise the dominant fish sub-order in Antarctic waters in terms of biomass and species abundance. During evolution in the cold and stable Antarctic climate, the Antarctic lineage of notothenioids developed several unique physiological adaptations, which make them extremely vulnerable to the rapid warming of Antarctic waters currently observed. Only recently, a further phenomenon exclusive to notothenioid fish was reported: the translocation of the mitochondrial gene encoding the NADH Dehydrogenase subunit 6 (ND6), an indispensable part of complex I in the mitochondrial electron transport system. This study investigated the potential physiological consequences of ND6 translocation for the function and thermal sensitivity of the electron transport system in isolated liver mitochondria of the two nototheniid species Notothenia coriiceps and Notothenia rossii, with special attention to the contributions of complex I (NADH DH) and complex II (Succinate DH) to oxidative phosphorylation. Furthermore, enzymatic activities of NADH∶Cytochrome c Oxidoreductase and Cytochrome C Oxidase were measured in membrane-enriched tissue extracts. During acute thermal challenge (0–15°C), capacities of mitochondrial respiration and enzymatic function in the liver could only be increased until 9°C. Mitochondrial complex I (NADH Dehydrogenase) was fully functional but displayed a higher thermal sensitivity than the other complexes of the electron transport system, which may specifically result from its unique amino acid composition, revealing a lower degree of stability in notothenioids in general. We interpret the translocation of ND6 as functionally neutral but the change in amino acid sequence as adaptive and supportive of cold stenothermy in Antarctic nototheniids. From these findings, an enhanced sensitivity to ocean warming can be deduced for Antarctic notothenioid fish.

The major haemoglobin of the sub-Antarctic fish Eleginops maclovinus was structurally and functionally characterised with the aim to compare molecular environmental adaptations in the O(2)-transport system of sub-Antarctic fishes of the suborder Notothenioidei with those of their high-latitude relatives. Ligand-binding kinetics of the major haemoglobin of E. maclovinus indicated strong stabilisation of the liganded quaternary T state, enhanced in the presence of the physiological allosteric effector ATP, compared to that of high-Antarctic Trematomus bernacchii. The activation enthalpy for O(2) dissociation was dramatically lower than that in T. bernacchii haemoglobin, suggesting remarkable differences in temperature sensitivity and structural changes associated with O(2) release and exit from the protein. The haemoglobin functional properties, together with the X-ray structure of the CO form at 1.49 Å resolution, the first of a temperate notothenioid, strongly support the hypothesis that in E. maclovinus, whose life-style varies according to changes in habitat, the mechanisms that regulate O(2) affinity and the ATP-induced Root effect differ from those of high-Antarctic Notothenioids.

The paper demonstrates results of the stock monitoring of blackfin icefish in the region of South Georgia in the period of 1988/89-1991/92 as well as summarises seventeen-year period ofobservation (1976/77 -1991/92). The influence of fishing on stock strucrture and supposed reasons of fluctuation ofyear-classes number are discussed.

Most fishes of the perciform suborder Notothenioidei are endemic to the subzero marine waters of Antarctica. A number of remarkable physiological attributes allow them to inhabit this extreme environment; for example, the blood of almost all notothenioid species contains antifreeze glycopeptides. To establish a molecular phylogenetic hypothesis for these fishes, DNA sequences from two mitochondrial genes, portions of the 12s and 16s ribosomal genes (928 base pairs [ bp] ), were determined for 18 species. These belong to 15 genera in five families of the suborder. The DNA data suggest that two of these families are unnatural groups and consequently that the classification and phylogeny of this suborder is in need of revision. In terms of DNA variation, the Bovichtidae are a distantly related sister group to the other families of the suborder that includes the icefishes, the only vertebrates without hemoglobin. The fishes of the suborder (except the Bovichtidae) seem to have speciated rapidly, forming an adaptive radiation in the Antarctic waters. A phylogenetic analysis of published hemoglobin amino acid sequences for other notothenioid fishes supports these results from mtDNA. On the basis of molecular phylogeny, the evolution of antifreeze glycopeptides was studied. The age of the radiation of notothenioid fishes had been estimated to be at least 38 Mya. However, the level of mtDNA variation detected in notothenioid fishes appears to be too low to agree with this date of origin and might instead suggest a younger age ( lo- 15 Mya). Alternatively, the low level of detected mtDNA variation would agree with the traditional old-age estimate if an extremely slow rate of mtDNA evolution is postulated for this group. This slow-rate hypothesis, if true, could be explained by decreased metabolic rates slowing down the tempo of molecular evolution.

The present paper evaluates the relevance of otoliths, vertebrae and other osseous elements to the age determinations; the age composition of stock, growth rate, length-weight relationship, condition, stage of gonads maturity as well as the age at first maturity were investigated, too.

The Antarctic notothenioid, Pagothenia bernacchii, were found to have plasma total and free calcium levels, plasma inorganic phosphate and whole body calcium efflux rates which were similar to those seen in other teleosts. But total bone calcium was lower than reported for other teleosts. A single injection of vitamin D3 (5 ng g(-1) fish) increased plasma total and plasma free calcium and these increases were associated with an increase in whole body calcium efflux and bone calcification. Conversely, the same treatment with 1,25-(OH)2-D3 reduced plasma free calcium. This seco-steroid also increased the specific activity of (45)Ca in bone at 40h post-injection but did not significantly effect total bone calcium, plasma total calcium or whole body calcium efflux. 25-OH-D3 at the same dose had no effect on any of the parameters tested and none of the seco-steroids tested had any effect on plasma total inorganic phosphate. These data show that both D3 and 1,25-(OH)3-D3 can have calcitropic effects in this marine teleost and that these two forms of vitamin D can exert different effects within the same species.