Amoebic gill disease increases energy requirements and decreases hypoxia tolerance in Atlantic salmon (Salmo salar) smolts.

Globally, Atlantic salmon (Salmo salar Linnaeus) aquaculture is now routinely affected by amoebic gill disease (AGD; Neoparamoeba perurans). The disease proliferates throughout the summer and is implicated in decreasing tolerance of salmon to environmental perturbations, yet little empirical evidence exists to support these observations. Using salmon acclimated to 15 or 19 °C, our aim was to determine the effects of clinically light-moderate (industry-relevant) AGD on metabolism (MO2rest and MO2max), aerobic scope (MO2max - MO2rest), excess post-exercise oxygen consumption (EPOC), and hypoxia tolerance. An increase in MO2rest (~8% and ~ 13% increase within the 15 and 19 °C acclimation groups, respectively) with increasing disease signs demonstrated an increase in baseline energy requirements as the disease progressed. Conversely, MO2max remained stable at both temperatures (~364 mg O2 kg-1 h-1), resulting in a decline in aerobic scope by 13 and 19% in the 15 and 19 °C groups, respectively. There was evidence of a decrease in hypoxia tolerance as the dissolved oxygen concentrations at loss of equilibrium increased by ~8% with more severe lesion coverage of the gills. These results suggest an increase in basal energy requirements and reduction in hypoxia tolerance as AGD proliferates, lending support to the idea that AGD reduces environmental tolerance. However, the lack of an effect of acclimation temperature indicates that the temperature-disease interaction may be more complicated than currently thought.

Aerobic and anaerobic movement energetics of hybrid and pure parental abalone.

The underlying mechanisms controlling growth heterosis in marine invertebrates remain poorly understood. We used pure blacklip (Haliotis rubra) and greenlip (Haliotis laevigata) abalone, as well as their hybrid, to test whether differences in movement and/or aerobic versus anaerobic energy use are linked to a purported increased growth rate in hybrids. Abalone were acclimated to control (16 °C) and typical summer temperatures (23 °C), each with oxygen treatments of 100% air saturation (O2sat) or 70% O2sat. The experiment then consisted of two phases. During the first phase (chronic exposure), movement and oxygen consumption rates (MO2) of abalone were measured during a 2 day observation period at stable acclimation conditions. Additionaly, lactate dehydrogenase (LDH) and tauropine dehydrogenase (TDH) activities were measured. During phase two (acute exposure), O2sat was raised to 100% for abalone acclimated to 70% O2sat followed by an acute decrease in oxygen to anoxia for all acclimation groups during which movement and MO2 were determined again. During the chronic exposure, hybrids and H. laevigata moved shorter distances than H. rubra. Resting MO2, LDH and TDH activities, however, were similar between abalone types but were increased at 23 °C compared to 16 °C. During the acute exposure, the initial increase to 100% O2sat for individuals acclimated to 70% O2sat resulted in increased movement compared to individuals acclimated to 100% O2sat for hybrids and H. rubra when compared within type of abalone. Similarly, MO2 during spontaneous activity of all three types of abalone previously subjected to 70% O2sat increased above those at 100% O2sat. When oxygen levels had dropped below the critical oxygen level (Pcrit), movement in hybrids and H. laevigata increased up to 6.5-fold compared to movement above Pcrit. Differences in movement and energy use between hybrids and pure species were not marked enough to support the hypothesis that the purportedly higher growth in hybrids is due to an energetic advantage over pure species.

Evidence for multiple sex-determining loci in Tasmanian Atlantic salmon (Salmo salar).

Phenotypic sex in salmonids is determined primarily by a genetic male heterogametic system; yet, sex reversal can be accomplished via hormonal treatment. In Tasmanian Atlantic salmon aquaculture, to overcome problems associated with early sexual maturation in males, sex-reversed females are crossed with normal females to produce all female stock. However, phenotypic distinction of sex-reversed females (neo-males) from true males is problematic. We set out to identify genetic markers that could make this distinction. Microsatellite markers from chromosome 2 (Ssa02), to which the sex-determining locus (SEX) has been mapped in two Scottish Atlantic salmon families, did not predict sex in a pilot study of seven families. A TaqMan 64 SNP genome-wide scan suggested SEX was on Ssa06 in these families, and this was confirmed by microsatellite markers. A survey of 58 families in total representing 38 male lineages in the SALTAS breeding program found that 34 of the families had SEX on Ssa02, in 22 of the families SEX was on Ssa06, and two of the families had a third SEX locus, on Ssa03. A PCR test using primers designed from the recently published sdY gene is consistent with Tasmanian Atlantic salmon having a single sex-determining gene that may be located on at least three linkage groups.

Development of genetic markers in abalone through construction of a SNP database.

In the absence of a reference genome, single-nucleotide polymorphisms (SNP) discovery in a group of abalone species was undertaken by random sequence assembly. A web-based interface was constructed, and 11 932 DNA sequences from the genus Haliotis were assembled, with 1321 contigs built. Of these, 118 contigs that consisted of at least ten annotation groups were selected. The 1577 putative SNPs were identified from the 118 contigs, with SNPs in several HSP70 gene contigs confirmed by PCR amplification of an 809-bp DNA fragment. SNPs in the HSP70 gene were compared across eight abalone species. A total of 129 polymorphic sites, including heterozygote sites within and among species, were observed. Phylogenetic analysis of the partial HSP70 gene region showed separation of the tested abalone into two groups, one reflecting the southern hemisphere species and the other the northern hemisphere species. Interestingly, Haliotis iris from New Zealand showed a closer relationship to species distributed in the northern Pacific region. Although HSP genes are known to be highly conserved among taxa, the validation of polymorphic SNPs from HSP70 in this mollusc demonstrates the applicability of cross-species SNP markers in abalone and the first step towards universal nuclear markers in Haliotis.

A DNA-based method for identification of krill species and its application to analysing the diet of marine vertebrate predators.

Accurate identification of species that are consumed by vertebrate predators is necessary for understanding marine food webs. Morphological methods for identifying prey components after consumption often fail to make accurate identifications of invertebrates because prey morphology becomes damaged during capture, ingestion and digestion. Another disadvantage of morphological methods for prey identification is that they often involve sampling procedures that are disruptive for the predator, such as stomach flushing or lethal collection. We have developed a DNA-based method for identifying species of krill (Crustacea: Malacostraca), an enormously abundant group of invertebrates that are directly consumed by many groups of marine vertebrates. The DNA-based approach allows identification of krill species present in samples of vertebrate stomach contents, vomit, and, more importantly, faeces. Utilizing samples of faeces from vertebrate predators minimizes the impact of dietary studies on the subject animals. We demonstrate our method first on samples of Adelie penguin (Pygoscelis adeliae) stomach contents, where DNA-based species identification can be confirmed by prey morphology. We then apply the method to faeces of Adelie penguins and to faeces of the endangered pygmy blue whale (Balaenoptera musculus brevicauda). In each of these cases, krill species consumed by the predators could be identified from their DNA present in faeces or stomach contents.

Genetic differentiation in the Antarctic coastal krill Euphausia crystallorophias.

The population genetics of the Antarctic neritic krill species Euphausia crystallorophias was examined by nucleotide sequence variation in its mitochondrial DNA. A 616 base pair region of the cytochrome c oxidase subunit I (COI) gene was screened for mutations by single-strand conformational polymorphism (SSCP) combined with restriction digestion. E. crystallorophias caught in three different regions of the Antarctic coastline were used--two samples from the Mertz Glacier Polynya and one sample each from the western side of the Antarctic Peninsula and from the Davis Sea. Significant genetic differences between krill samples were identified. However, the extent of these differences did not correlate with the degree of geographic separation between the sampling sites. This suggests that the genetic structuring may be the result of small-scale differentiation rather than differentiation between resident populations in separate parts of the Southern Ocean. The possibility that genetic differences between samples within a region are as important as differences between regions has implications for other studies of krill population genetics.

Unusually high levels of non-saponifiable lipids in the fishes escolar and rudderfish identification by gas and thin-layer chromatography.

Analysis of the non-saponifiable lipids of the fishes Lepidocybium flavobrunneum and Ruvettus pretiosus (escolar), and Centrolophus niger and Tubbia spp. (rudderfish) was performed. The analyses were used to clarify the cause of recent reports of illness (diarrhoea) in Australia from consumption of purported rudderfish. Both escolar and rudderfish contained very high levels of oil (generally between 14 to 25%, as % wet mass) in the fillet and the oil compositions were different to most seafood. Escolar oil contained mainly wax ester (>90% of oil). The oil from five specimens of rudderfish contained mainly diacylglyceryl ether (DAGE, >80% of oil) or hydrocarbon (>80% of oil, predominately squalene). One rudderfish specimen contained mainly polar lipid. Major differences in oil content and composition, including fatty alcohol and glyceryl ether diols (derived from DAGE), were observed between purported individuals of the same species or related species of rudderfish, raising the possibility of geographic or seasonal differences affecting the oil composition. The oil composition of fish fillet samples associated with the health issues were consistent with the profiles for escolar, rather than rudderfish species. These findings, in particular the lipid class and fatty alcohol profiles, were supported by general protein fingerprinting results and were consistent with the samples originating from individuals of the escolar species L. flavobrunneum. The high wax ester content of the escolar group clarifies the reported diarrhoeal effects to consumers. Purgative properties of high wax ester containing fish oils have been reported for escolar and other species. The results highlight the potential for non-saponifiable lipid profiles to be used for identification of fish fillets and oils to at least group level.

28S rDNA evolution in the Eumalacostraca and the phylogenetic position of krill.

The Malacostraca are an ancient and morphologically diverse class of Crustacea. The phylogenetic position of one order within this class, the Euphausiacea ("krill," subclass Eumalacostraca) was investigated using 28S rDNA sequences from representatives of several malacostracan orders. Phylogenies for these sequences were estimated by maximum-likelihood and maximum-parsimony analysis. The results of these analyses produced a new scheme for evolution within the Eumalacostraca. The new phylogenies suggested that Euphausiacea are most closely related to the Mysida and not the Decapoda, as is generally thought. Furthermore, the Mysida were found not to be closely related to the Lophogastrida, which are often considered their sister taxon. These hypotheses were tested against the hypotheses of monophyly for the Eucarida, Mysidacea, and Peracarida and found to be significantly better on the basis of the 28S rDNA data.