title: Molecular Characterisation of Immune Genes in Yellowtail Kingfish (Seriola lalandi)

authors: Broomfield, Grant Robert

subjects: seriola

abstract: The effective and efficient introduction of a new candidate species for aquaculture depends greatly on the understanding of the physiology of the organism and the ability to monitor a candidates’ response to changes in their environment. For any farmed fish in an aquaculture setting, there will be a number of bottlenecks that restrict optimal growth and production. Being able to monitor and maintain health is one of these, with stress and pathogenic invasion from microbial organisms leading to a reduction in farming efficiency. Due to the very nature of aquaculture, stress and transmission of pathogens are very common, as fish are contained in close quarters. Thus, characterising and understanding the immune system of these fish will allow for better monitoring of fish health. 
Yellowtail Kingfish (Seriola lalandi) is a strong candidate for introduction into the New Zealand aquaculture industry. The selection of this fish as a candidate for aquaculture is based on its economically beneficial traits that have potential in both domestic and export markets, particularly in the sashimi market in Japan. Because of this, it is important to begin investigations into the immune response of this species, so aspects of its health can be monitored during farming. The focus of this investigation was to identify immune genes in this species and to begin profiling their expression during the development of the fish. 
Using bioinformatics approaches, the available gene databases and the implementation of a transcriptomic library prepared from the spleen tissue of S. lalandi, a number of immune genes were identified, which included, Immunoglobulin D (IgD), Recombination Activation Gene 1 (RAG1) and Interleukin-1β (IL-1b). Primers were designed for each gene, to enable confirmation of the gene sequences, by PCR and RACE-PCR. Using this approach, no confirmed sequence was obtained for IL-1b, however sequence was obtained for part of the RAG1 gene and a large part of the IgD gene, including the 3’end. 
	The availability of these immune gene sequences, allowed methods to be developed for looking at gene expression. Primers were designed using the confirmed sequences for RAG1 and IgD, as well as the unconfirmed sequence for IL-1b. These were used in qPCR to examine the expression of each gene during development, within larvae at hatch, 3 dph, 12 dph and 18 dph. Expression of each gene was found to increase by 12 dph, but then decrease at 18dph, with IL-1b showing the highest relative expression. However, no significant difference in expression was seen statistically between any of the time points.
Lastly, a protocol was developed for the histological sectioning of larval fish at different stages of development. A number of attempts were made to optimise an approach, with fish from 3 dph to 60 dph eventually being paraffin embedded, sectioned and stained with either toluidine blue or haematoxylin and eosin (H&E). Developing structures within the larvae, including immune tissues, were identified. Future work will use prepared sections to stain for the immune genes, to gain a better understanding of their role. 
Discovery of immune genes from kingfish, such as the ones characterised in this investigation, will allow the development of invaluable tools. These tools can be used to monitor the health of fish during both the developmental and adult stages. This paves the way for informative studies that will benefit the future aquaculture of this species.

text: Molecular Characterisation of Immune Genes in Yellowtail Kingfish (Seriola lalandi) The effective and efficient introduction of a new candidate species for aquaculture depends greatly on the understanding of the physiology of the organism and the ability to monitor a candidates’ response to changes in their environment. For any farmed fish in an aquaculture setting, there will be a number of bottlenecks that restrict optimal growth and production. Being able to monitor and maintain health is one of these, with stress and pathogenic invasion from microbial organisms leading to a reduction in farming efficiency. Due to the very nature of aquaculture, stress and transmission of pathogens are very common, as fish are contained in close quarters. Thus, characterising and understanding the immune system of these fish will allow for better monitoring of fish health. 
Yellowtail Kingfish (Seriola lalandi) is a strong candidate for introduction into the New Zealand aquaculture industry. The selection of this fish as a candidate for aquaculture is based on its economically beneficial traits that have potential in both domestic and export markets, particularly in the sashimi market in Japan. Because of this, it is important to begin investigations into the immune response of this species, so aspects of its health can be monitored during farming. The focus of this investigation was to identify immune genes in this species and to begin profiling their expression during the development of the fish. 
Using bioinformatics approaches, the available gene databases and the implementation of a transcriptomic library prepared from the spleen tissue of S. lalandi, a number of immune genes were identified, which included, Immunoglobulin D (IgD), Recombination Activation Gene 1 (RAG1) and Interleukin-1β (IL-1b). Primers were designed for each gene, to enable confirmation of the gene sequences, by PCR and RACE-PCR. Using this approach, no confirmed sequence was obtained for IL-1b, however sequence was obtained for part of the RAG1 gene and a large part of the IgD gene, including the 3’end. 
	The availability of these immune gene sequences, allowed methods to be developed for looking at gene expression. Primers were designed using the confirmed sequences for RAG1 and IgD, as well as the unconfirmed sequence for IL-1b. These were used in qPCR to examine the expression of each gene during development, within larvae at hatch, 3 dph, 12 dph and 18 dph. Expression of each gene was found to increase by 12 dph, but then decrease at 18dph, with IL-1b showing the highest relative expression. However, no significant difference in expression was seen statistically between any of the time points.
Lastly, a protocol was developed for the histological sectioning of larval fish at different stages of development. A number of attempts were made to optimise an approach, with fish from 3 dph to 60 dph eventually being paraffin embedded, sectioned and stained with either toluidine blue or haematoxylin and eosin (H&E). Developing structures within the larvae, including immune tissues, were identified. Future work will use prepared sections to stain for the immune genes, to gain a better understanding of their role. 
Discovery of immune genes from kingfish, such as the ones characterised in this investigation, will allow the development of invaluable tools. These tools can be used to monitor the health of fish during both the developmental and adult stages. This paves the way for informative studies that will benefit the future aquaculture of this species.

year: 2015

title: A standardised sampling protocol for robust assessment of reach-scale fish community diversity in wadeable New Zealand streams

authors: David, Bruno O.

subjects: standard methods

abstract: The New Zealand fish fauna contains species that are affected not only by river system connectivity, but also by catchment and local-scale changes in landcover, water quality and habitat quality. Consequently, native fish have potential as multi-scale bioindicators of human pressure on stream ecosystems, yet no standardised, repeatable and scientifically defensible methods currently exist for effectively quantifying their abundance or diversity in New Zealand stream reaches. Here we report on the testing of a back-pack electrofishing method, modified from that used by the United States Environmental Protection Agency, on a wide variety of wadeable stream reaches throughout New Zealand. Seventy-three first- to third-order stream reaches were fished with a single pass over 150-345 m length. Time taken to sample a reach using single-pass electrofishing ranged from 1-8 h. Species accumulation curves indicated that, irrespective of location, continuous sampling of 150 stream metres is required to accurately describe reach-scale fish species richness using this approach. Additional species detection beyond 150 m was rare (<10%) with a single additional species detected at only two out of the 17 reaches sampled beyond this distance. A positive relationship was also evident between species detection and area fished, although stream length rather than area appeared to be the better predictor. The method tested provides a standardised and repeatable approach for regional and/or national reporting on the state of New Zealand's freshwater fish communities and trends in richness and abundance over time.

text: A standardised sampling protocol for robust assessment of reach-scale fish community diversity in wadeable New Zealand streams The New Zealand fish fauna contains species that are affected not only by river system connectivity, but also by catchment and local-scale changes in landcover, water quality and habitat quality. Consequently, native fish have potential as multi-scale bioindicators of human pressure on stream ecosystems, yet no standardised, repeatable and scientifically defensible methods currently exist for effectively quantifying their abundance or diversity in New Zealand stream reaches. Here we report on the testing of a back-pack electrofishing method, modified from that used by the United States Environmental Protection Agency, on a wide variety of wadeable stream reaches throughout New Zealand. Seventy-three first- to third-order stream reaches were fished with a single pass over 150-345 m length. Time taken to sample a reach using single-pass electrofishing ranged from 1-8 h. Species accumulation curves indicated that, irrespective of location, continuous sampling of 150 stream metres is required to accurately describe reach-scale fish species richness using this approach. Additional species detection beyond 150 m was rare (<10%) with a single additional species detected at only two out of the 17 reaches sampled beyond this distance. A positive relationship was also evident between species detection and area fished, although stream length rather than area appeared to be the better predictor. The method tested provides a standardised and repeatable approach for regional and/or national reporting on the state of New Zealand's freshwater fish communities and trends in richness and abundance over time.

year: 2010

title: The Portrayal of Maori offenders by the Media

authors: Farr, Heather Marama

abstract: In this study I examine the dominant discourses represented in the print and internet media reporting of offending by Maori.  The purpose of which is to analyse the dominant discourse and subject positions associated with offending by Maori.  Foucault Discourse Analysis was utilised whereby the data was collected through articles printed in the New Zealand Herald and the Dominion Post.  This study suggests that there are multiple and complex factors relating to Maori offending, whereby reporting about Maori offending is presented from a mainstream worldview.

text: The Portrayal of Maori offenders by the Media In this study I examine the dominant discourses represented in the print and internet media reporting of offending by Maori.  The purpose of which is to analyse the dominant discourse and subject positions associated with offending by Maori.  Foucault Discourse Analysis was utilised whereby the data was collected through articles printed in the New Zealand Herald and the Dominion Post.  This study suggests that there are multiple and complex factors relating to Maori offending, whereby reporting about Maori offending is presented from a mainstream worldview.

year: 2019