Characterisation of the Mitochondrial Genome and the Population Genetics of Polyprion Oxygeneios (Hapuku) from Around New Zealand

<p><b>Polyprion oxygeneios (hapuku) is an important commercial and recreational fishery species within New Zealand. Moreover, P. oxygeneios are currently being developed as a high-value New Zealand aquaculture species. There have been no previous studies on New Zealand’s P. oxygeneios that have been able to detect genetic differences among samples, which may be of use to either broodstock or fisheries managers. An understanding of the genetic structure of commercially harvested species maximises the potential for sustainable harvesting through effective management schemes. The primary goal of this thesis was to investigate the population genetic structure of P. oxygeneios using molecular markers to analyse samples collected from sites within New Zealand’s Exclusive Economic Zone (EEZ).</b></p> <p>The DNA sequence of the whole mitochondrial genome of P. oxygeneios was determined and it showed a similar structure and gene organisation to that of other species across a wide range of taxa. A set of species-specific control region primers was developed for P. oxygeneios and Polyprion americanus, and additional primers were designed for the 16S and ND6 genes of P. oxygeneios. A ~488 bp portion of the mitochondrial DNA (mtDNA) control region sequence from 274 individuals, and genotypes from 259 individuals using nine polymorphic microsatellite loci, were used to investigate the phylogeography and population genetic structure of P. oxygeneios. The mitochondrial DNA data failed to detect any significant differentiation between sample sites. However, the microsatellite DNA analyses showed that individuals sampled from the west coast of the South Island (Hokitika) were genetically distinct from individuals sampled at all other New Zealand sites. These two groups might be representative of two discrete populations of P. oxygeneios within New Zealand’s EEZ. These results suggest that the west coast South Island P. oxygeneios fishery should continue to be managed as a separate stock, with some possible revision of the Cook Strait fishery required. Analyses of the mtDNA and microsatellite DNA data of P. oxygeneios broodstock held at NIWA’s Bream Bay Aquaculture Park showed that they were not significantly differentiated from the wild populations (excluding Hokitika). Simulations also described the appropriate sampling efforts required to capture an appropriate level of genetic diversity when either establishing a new broodstock or supplementing an existing broodstock with new individuals. Continued management of the broodstock will be required to maintain the high levels of genetic diversity that have been captured in the founding broodstock in future generations.</p>

Characterisation of the Mitochondrial Genome and the Population Genetics of Polyprion Oxygeneios (Hapuku) from Around New Zealand

<p><b>Polyprion oxygeneios (hapuku) is an important commercial and recreational fishery species within New Zealand. Moreover, P. oxygeneios are currently being developed as a high-value New Zealand aquaculture species. There have been no previous studies on New Zealand’s P. oxygeneios that have been able to detect genetic differences among samples, which may be of use to either broodstock or fisheries managers. An understanding of the genetic structure of commercially harvested species maximises the potential for sustainable harvesting through effective management schemes. The primary goal of this thesis was to investigate the population genetic structure of P. oxygeneios using molecular markers to analyse samples collected from sites within New Zealand’s Exclusive Economic Zone (EEZ).</b></p> <p>The DNA sequence of the whole mitochondrial genome of P. oxygeneios was determined and it showed a similar structure and gene organisation to that of other species across a wide range of taxa. A set of species-specific control region primers was developed for P. oxygeneios and Polyprion americanus, and additional primers were designed for the 16S and ND6 genes of P. oxygeneios. A ~488 bp portion of the mitochondrial DNA (mtDNA) control region sequence from 274 individuals, and genotypes from 259 individuals using nine polymorphic microsatellite loci, were used to investigate the phylogeography and population genetic structure of P. oxygeneios. The mitochondrial DNA data failed to detect any significant differentiation between sample sites. However, the microsatellite DNA analyses showed that individuals sampled from the west coast of the South Island (Hokitika) were genetically distinct from individuals sampled at all other New Zealand sites. These two groups might be representative of two discrete populations of P. oxygeneios within New Zealand’s EEZ. These results suggest that the west coast South Island P. oxygeneios fishery should continue to be managed as a separate stock, with some possible revision of the Cook Strait fishery required. Analyses of the mtDNA and microsatellite DNA data of P. oxygeneios broodstock held at NIWA’s Bream Bay Aquaculture Park showed that they were not significantly differentiated from the wild populations (excluding Hokitika). Simulations also described the appropriate sampling efforts required to capture an appropriate level of genetic diversity when either establishing a new broodstock or supplementing an existing broodstock with new individuals. Continued management of the broodstock will be required to maintain the high levels of genetic diversity that have been captured in the founding broodstock in future generations.</p>

Ontogeny of the Hapuka (Polyprion oxygeneios) Immune System

<p>In this study the ontogeny of the hapuka (Polyprion oxygeneios) immune system was studied during larval development. In teleost fish, the head kidney, thymus, and spleen are generally regarded as important immune organs. The head kidney was observed at 4 days post hatch (dph), the spleen at 16 dph and lastly the thymus at 20 dph and all 3 lymphoid organs were relatively well developed by 45 dph. The immune genes CSF1R, C3, MHCIIα, TCRα, TCRβ, RAG1, IgM and IgZ were examined by RT-PCR to investigate the leucocyte development. Macrophages appear to be present from hatch with both CSF1R and MHCIIα expression from 1 dph, while IgM is expressed at 9 dph. T-cells appear later in hapuka with TCRβ expression first detected at 32 dph whereas TCRα was not expressed until after 63 dph. Immunostaining using a monoclonal antibody against fish IgM detected IgM in the head kidney at 12 dph, the spleen at 32 dph, the intestinal tract at 45 dph and lastly the thymus at 50 dph. Comparison of the leucocyte populations in juveniles and adults indicated that innate cell populations are late to develop, while the adaptive cells mature earlier in hapuka than expected. Finally, the maternal transfer of immunity was examined and while lysozyme and IgM appear to be transferred, complement does not. Overall this study provides insight into the developmental sequence of immune organs and cells and will be useful in understanding the timing of immune competence in juveniles and adult hapuka.</p>

Ontogeny of the Hapuka (Polyprion oxygeneios) Immune System

<p>In this study the ontogeny of the hapuka (Polyprion oxygeneios) immune system was studied during larval development. In teleost fish, the head kidney, thymus, and spleen are generally regarded as important immune organs. The head kidney was observed at 4 days post hatch (dph), the spleen at 16 dph and lastly the thymus at 20 dph and all 3 lymphoid organs were relatively well developed by 45 dph. The immune genes CSF1R, C3, MHCIIα, TCRα, TCRβ, RAG1, IgM and IgZ were examined by RT-PCR to investigate the leucocyte development. Macrophages appear to be present from hatch with both CSF1R and MHCIIα expression from 1 dph, while IgM is expressed at 9 dph. T-cells appear later in hapuka with TCRβ expression first detected at 32 dph whereas TCRα was not expressed until after 63 dph. Immunostaining using a monoclonal antibody against fish IgM detected IgM in the head kidney at 12 dph, the spleen at 32 dph, the intestinal tract at 45 dph and lastly the thymus at 50 dph. Comparison of the leucocyte populations in juveniles and adults indicated that innate cell populations are late to develop, while the adaptive cells mature earlier in hapuka than expected. Finally, the maternal transfer of immunity was examined and while lysozyme and IgM appear to be transferred, complement does not. Overall this study provides insight into the developmental sequence of immune organs and cells and will be useful in understanding the timing of immune competence in juveniles and adult hapuka.</p>

First report of the hapuku wreckfish Polyprion oxygeneios (Polyprionidae) in Argentinian waters

The hapuku wreckfish Polyprion oxygeneios is recorded for the first time in Argentinian waters. Four specimens were caught off Mar del Plata (38°S, Argentina) in depths between 60 and 260 m during austral summer (three by sport fishers and one in a research cruise). Up to the present, this species was consistently misidentified with its congener, the common wreckfish P. americanus, off Argentina. These records represent a connection between previous records from Brazil and the more austral circumglobal distribution of the species.

Induced Spawning of F1 Wreckfish (Hāpuku) Polyprion oxygeneios Using a Synthetic Agonist of Gonadotropin-Releasing Hormone

Wild-caught hāpuku (Polyprion oxygeneios) spawn readily in captivity, but although first filial (F1) hāpuku complete vitellogenesis, females fail to undergo oocyte maturation and spawn or produce poor quality eggs. This study investigated whether administration of a synthetic agonist of gonadotropin-releasing hormone (GnRHa) could improve F1 hāpuku spawning and complete the life-cycle in captivity. Spawning trials were conducted over 2 years in 2013 and 2014, when F1 were aged five and six years. In 2013, females previously conditioned under a variable or constant temperature regime were implanted with GnRHa (100 μg/kg−1) or blank implants constructed of powdered cellulose and cholesterol. Spawning was erratic and egg quality very poor in all tanks. No F2 offspring were produced by communal spawning. In contrast, viable F2 larvae were produced by strip-spawning and in vitro fertilization after a series of GnRHa injections. In 2014, two additional trials were conducted: females received ethylene-vinyl acetate copolymer (EVAc) matrix implants containing GnRHa (100 μg/kg−1) or blank implants and in the second trial, two GnRHa doses (100 μg/kg−1 and 50 μg/kg−1) were tested. Eggs were first detected in all tanks 12–17 days post-implantation when females received 100 µg/kg−1 GnRHa implants, but not in the lower dose or control tanks. In summary, this study achieved induction of female spawning with GnRHa implants (target dose 100 μg/kg−1) and the successful production of F2 hāpuku in captivity by strip-spawning.

Transcriptomic Changes during Previtellogenic and Vitellogenic Stages of Ovarian Development in Wreckfish (Hāpuku), Polyprion oxygeneios (Perciformes)

‘Wreckfish’ a collective of species belonging to the family Polyprionidae, are an important commercial fishery and have significant aquaculture potential. Until now, genomic or transcriptomic information for any species within the genus Polyprion has either remained unpublished or is non-existent. Using Illumina HiSeq, we compared the transcriptomes of hāpuku (Polyprion oxygeneios) ovaries to explore developmental stage-specific variations underlying their reproductive physiology. We sought to identify differentially expressed genes and the associated shifts in biological pathways between previtellogenic and early vitellogenic ovaries. Ovarian tissue was repeatedly biopsied by gonopore cannulation from the same females (n = 3) throughout oogenesis. Reproductive status of initial biopsies was confirmed as being previtellogenic and that in biopsies collected eight weeks later as early vitellogenic. A de novo hāpuku transcriptome was assembled (146,189 transcripts) from RNA-Seq data without a reference genome. On average, each tissue sample contained 17.5 million trimmed reads. Gene annotation was 80% when using BLASTX against Genbank Non Redundant database. Fifty-three transcripts were differentially expressed within the FDR of 0.05 when previtellogenic and early vitellogenic ovaries were compared; this reduced to 35 differentially expressed genes when transcript duplications were pooled. Among these were genes tentatively associated with the electron transport chain, lipid metabolism, steroidogenesis and mineral/solute transportation. These data provide a snap-shot into stage-specific physiological events during oogenesis in the ovary of a teleost and an extensive molecular resource for research on species in the Genus Polyprion.