Genetic diversity of Antarctic fish

200 Pages·2015·6.2 MB·English

by  Elaine Fitzcharles

GENETIC DIVERSITY OF ANTARCTIC FISH
Elaine M. Fitzcharles
A Thesis Submitted for the Degree of PhD
at the
University of St Andrews
2014
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Genetic Diversity of Antarctic Fish

Elaine M. Fitzcharles

This thesis is submitted in partial fulfilment for the degree of PhD
at the
University of St Andrews

May 2014
Supervisor of studies
Prof. Alex Rogers (University of Oxford)
Dr Melody Clark (British Antarctic Survey)
Prof. Andrew Brierley (University of St Andrews)

Sponsoring establishment
British Antarctic Survey
Natural Environment Research Council
High Cross
Madingley Road
Cambridge
CB3 0ET
United Kingdom
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DECLARATIONS
1. Candidate’s declarations:
I, Elaine Fitzcharles, hereby certify that this thesis, which is approximately 35000 words in length, has
been written by me, and that it is the record of work carried out by me, or principally by myself in
collaboration with others as acknowledged, and that it has not been submitted in any previous application
for a higher degree.
I was admitted as a research student in September 2005 and as a candidate for the degree of Doctor of
Philosophy in April 2007; the higher study for which this is a record was carried out in the University of St
Andrews between 2005 and 2014.
Date 23/5/11 Signature of candidate ……………………..............

2. Supervisor’s declaration:
I hereby certify that the candidate has fulfilled the conditions of the Resolution and Regulations appropriate
for the degree of Doctor of Philosophy in the University of St Andrews and that the candidate is qualified to
submit this thesis in application for that degree.
Date 23/5/11 Signature of supervisor ……………………..............

3. Permission for publication: (to be signed by both candidate and supervisor)
In submitting this thesis to the University of St Andrews I understand that I am giving permission for it to be
made available for use in accordance with the regulations of the University Library for the time being in
force, subject to any copyright vested in the work not being affected thereby. I also understand that the
title and the abstract will be published, and that a copy of the work may be made and supplied to any bona
fide library or research worker, that my thesis will be electronically accessible for personal or research use
unless exempt by award of an embargo as requested below, and that the library has the right to migrate
my thesis into new electronic forms as required to ensure continued access to the thesis. I have obtained
any third-party copyright permissions that may be required in order to allow such access and migration, or
have requested the appropriate embargo below.
The following is an agreed request by candidate and supervisor regarding the publication of this thesis:
PRINTED COPY
a) No embargo on print copy
ELECTRONIC COPY
a) No embargo on electronic copy
Date 23/5/11
Signature of candidate ……… Signature of supervisor ……………………..............

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Acknowledgements
This part-time PhD was funded by the British Antarctic Survey. First and foremost I
have to thank my supervisors, Alex Rogers and Melody Clark, for having enough faith
in me as a scientist to let me begin this journey, and Andy Brierley for having enough
faith in Alex to agree to take me on. It has been a longer journey than any of us intended
so thank you, especially to Melody, for your continued support when life, work and
unplanned distractions got in the way.
Special thanks go to Martin Collins and Mark Belchier for the fish samples, advice and
support over the years. Your interest in the results has helped to keep me going. Also, to
the number of fisheries scientists who have kindly collected me samples over the years;
Sue Gregory, Judith Brown and Katie Brigden to name just a few. Thanks to Peter
Smith, Steve Parker and Peter McMillan for the samples from the Far Side (Ross Sea),
and Sarah Helyar for the ones from the North and the microsatellite collaboration.
I have to thank Will Goodall-Copestake for the technical help and training in
phylogenetic analysis, as well as general support and encouragement, Huw Griffiths for
the assistance with the GIS maps and Michelle Taylor for her last minute advice and
reassurance. My BAS management has changed over the year but particular thanks go
to Tim Moffat for recognising the importance of this work and giving me the time to
finish it. I also need to thank my colleagues in the biology lab management team, Guy
Hillyard, Paul Geissler and Michelle King, for continued support over the years but
particularly these last few months.
Taking on a part-time PhD and a full-time job is not an easy task and would not have
been possible without the support of my work colleagues in the Angel lab and fellow
PhD students (too many to mention). Particular thanks go to Katrin Linse and Sandra
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McInnes for their advice, support and friendship, and to Natalie Ensor and Mags
Wallace for being there and understanding. On a personal note I have to thank my
brother, Colin Mitchell, for helping to keep me sane, Ria (Bob) Kishen for the shared
frustrations over molecular biology (we never had these problems with McGregor’s
cakes) and Leanne Hulse for giving me a home from home on my visits to St Andrews.
Most of all I have to thank my husband, Scott Fitzcharles, for agreeing to let me do this
and being there for me throughout. There have been some tough times during the course
of this PhD but also the happiest times. Thank you for Frewin and Caelan. Having two
boys while trying to juggle the job and PhD probably wasn’t the most sensible idea but
it was the best one. They may add to the frustrations, exhaustion and general chaos but
they make everything worthwhile.
Finally, thank you to my mother for the continued support in everything I decide to do
in life (even the bad ideas) and always being there when I need her. To my father, I
know how proud you would have been.

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Abstract
Correct species identification is fundamental to all areas of biology, but particularly the
policy related areas of conservation and fisheries management. To enable guidelines to
be developed for environmental management and conservation, such identifications
need links to studies of the evolutionary history, biological factors and environmental
influences driving species divergence and population dynamics for the target species.
This study concerns two genera of gadiform fish, Muraenolepis and Macrourus, found
in southern temperate and Antarctic waters, with a single species, Macrourus berglax,
present in the North Atlantic. With similar distribution patterns to toothfish species,
Dissostichus eleginoides and D. mawsoni, they are a major food source and by-catch of
the toothfish fishery. Both are slow growing and long lived, with different evolutionary
histories, life expectancies and strategies for reproduction. For both genera, the accuracy
of morphological keys, number of described species and their distribution is under
debate.
This study has identified specimens to species level using both morphological and
genetic techniques, redefining the range for morphological features and taxonomic keys.
For Muraenolepis, this has clarified confusion over Mu. marmoratus and Mu. microps
being a single species, confirmed some mis-identification from sexual dimorphism and
provided genetic evidence for the recently described species Mu. evseenkoi. For
Macrourus, this work has identified a new species, now named Ma. caml, and found
that Ma. holotrachys and Ma. berglax are genetically identical, raising the question of
bipolar distribution or recent divergence.
The low level of genetic variation within both species suggests a recent evolution and
expansion into Antarctic waters. Similar geographic species limits imply common
processes influencing divergence, with the oceanographic fronts as potential barriers.
Further investigation of niche overlap and fine scale population structure are required to
fully understand the processes driving speciation and provide the underlying data
required for fisheries management.
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Contents
CHAPTER 1 .................................................................................................................................. 1
General introduction ...................................................................................................................... 1
1.1 Introduction ............................................................................................................................. 1
1.2 Antarctica and the Southern Ocean ......................................................................................... 1
1.3 Evolution of Antarctic Fish ..................................................................................................... 5
1.4 Study Fish Species .................................................................................................................. 6
1.5 Antarctic Fisheries................................................................................................................... 8
1.5 The role of genetics in conservation...................................................................................... 10
1.6 Aims and objectives .............................................................................................................. 12
1.7 This thesis .............................................................................................................................. 13
CHAPTER 2 ................................................................................................................................ 15
Materials and Methods ................................................................................................................ 15
2.1 Sample collection and Morphological Identification ............................................................ 15
2.2 Molecular Methods ............................................................................................................... 21
2.3 Species’ Specific COI Primers for Macrourus Species ........................................................ 23
2.5 Molecular Analysis of Mitochondrial Sequences ................................................................. 25
2.5 High Resolution Melt Analysis (HRMA) for Macrourus Species ........................................ 34
2.6 Microsatellite Markers .......................................................................................................... 40
CHAPTER 3 ................................................................................................................................ 42
The phylogenetics and phylogeography of the genus Muraenolepis .......................................... 42
3.1 Introduction ........................................................................................................................... 42
3.2 Morphological Identification ................................................................................................. 52
3.3 Molecular Identification and Phylogenetics .......................................................................... 58
3.4 Morphological Data............................................................................................................... 66
3.5 Geographic Distribution ........................................................................................................ 69
3.6 Population Genetics............................................................................................................... 72
3.7 Discussion ............................................................................................................................. 76
3.8 Summary of Results .............................................................................................................. 85
CHAPTER 4 ................................................................................................................................ 87
The phylogenetics and phylogeography of the genus, Macrourus.............................................. 87
4.1 Introduction ........................................................................................................................... 87
4.2 Morphological Identification ................................................................................................. 94
4.3 Molecular Identification and Phylogenetics .......................................................................... 98
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4.4 Morphological Data............................................................................................................. 104
4.5 Geographic Distribution ...................................................................................................... 110
4.6 Population Genetics............................................................................................................. 113
4.7 Discussion ........................................................................................................................... 117
4.8 Summary of Results ............................................................................................................ 127
CHAPTER 5 .............................................................................................................................. 129
Phylogenetic Inference for the Order Gadifomes ...................................................................... 129
5.1 Introduction ......................................................................................................................... 129
5.1 Methods ............................................................................................................................... 130
5.2 Results ................................................................................................................................. 132
5.4 Discussion ........................................................................................................................... 137
CHAPTER 6 .............................................................................................................................. 141
Macrourus microsatellite markers ............................................................................................ 141
6.1 Introduction ......................................................................................................................... 141
6.2 Methods ............................................................................................................................... 143
6.3 Results ................................................................................................................................. 146
6.4 Discussion ........................................................................................................................... 149
CHAPTER 7 .............................................................................................................................. 153
Discussion and conclusions ....................................................................................................... 153
7.3 Conclusions ......................................................................................................................... 160
CHAPTER 8 .............................................................................................................................. 163
Cited literature ........................................................................................................................... 163
APPENDIX I ............................................................................................................................. 174
In-house Microsatellite Isolation for Macrourus and Muraenolepis Species ........................... 174
APPENDIX II ........................................................................................................................... 185
Accession numbers for sequences from Genbank with outgroups highlighted in bold. ........... 185

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Table of Figures
Figure 1-1 Bottom topography of the Southern Ocean.....................................................2
Figure 1-2 Block diagram of the circulation of the Southern Ocean.................................3
Figure 1-3 Schematic representation of the convergences and divergences of the
Southern Ocean..................................................................................................................4
Figure 1-4 Typical morphology of the families Muraneolepididae and
Macrouridae.......................................................................................................................7
Figure 1-5 Map of CCAMLR zones for the Antarctic and subAntarctic islands showing
the division into fishing areas............................................................................................9
Figure 2-1 Diagram showing the measurements for species identification.................... 16
Figure 2-2 Key to identification of Muraenolepis species..............................................17
Figure 2-3 Drawing showing the difference in scaling to the underside of the head as
used to distinguish between Macrourus species..............................................................18
Figure 2-4 Key to identify Macrourus species................................................................19
Figure 2-5 COI sequence alignment for the four species showing polymorphic sites....35
Figure 2-6 Graph of Ct values and Difference Plot for Melt Curve from initial test......37
Figure 2-7 Graph of Ct values and Difference Plot for Melt Curve generated using the
redesigned primers...........................................................................................................38
Figure 2-8 HRM difference profiles for Ma. carinatus, Ma. holotrachys, Ma. whitsoni
and Ma. whitsoni..............................................................................................................39
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