Population Genetics and Demographic Resilience in Three Aquatic Invertebrates

567 Pages·2017·20.31 MB·English

by  Greg Macdonald

Population Genetics and Demographic
Resilience in Three Aquatic
Invertebrates

Thesis submitted for the degree of Doctor of Philosophy
by
Hannah C Macdonald

School of Biosciences
Cardiff University

July 2016

Supervisors: Prof. Mike Bruford, Dr Hefin Jones and Prof. Steve Ormerod
DECLARATION
This work has not been submitted in substance for any other degree or award at this or any
other university or place of learning, nor is being submitted concurrently in candidature for
any degree or other award.
Signed ………………………………………… (candidate) Date …08/07/16……

STATEMENT 1
This thesis is being submitted in partial fulfillment of the requirements for the degree of PhD
Signed ………………………………………… (candidate) Date …08/07/16……

STATEMENT 2
This thesis is the result of my own independent work/investigation, except where otherwise
stated. Other sources are acknowledged by explicit references. The views expressed are my
own.
Signed ………………………………………… (candidate) Date …08/07/16……

STATEMENT 3
I hereby give consent for my thesis, if accepted, to be available for photocopying and for
inter-library loan, and for the title and summary to be made available to outside
organisations.
Signed ………………………………………… (candidate) Date …08/07/16……

STATEMENT 4: PREVIOUSLY APPROVED BAR ON ACCESS
I hereby give consent for my thesis, if accepted, to be available for photocopying and for
inter-library loans after expiry of a bar on access previously approved by the Academic
Standards & Quality Committee.
Signed ………………………………………… (candidate) Date …08/07/16……
“In many of the more relaxed civilizations on the Outer Eastern Rim of the Galaxy, the
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much that is apocryphal, or at least wildly inaccurate, it scores over the older, more
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First, it is slightly cheaper; and secondly it has the words
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Acknowledgements

There is one person, without which I can say for certain I’d never have completed this thesis,
my husband and best friend, Greg Macdonald. From deciding whether to start the PhD in
the beginning, which involved moving to the other side of the country and having a long
distance relationship for more than three and a half years, to the countless trials during, he
gave me his full support and encouragement. He also lent me his computer programming
skills and his OCD spelling and formatting checking skills, without which this document
would be filled with my dyslexic spelling. When my funding came to an end he has
financially supported me, and during my write up whilst being pregnant and all of the things
that come with that, he has cooked, cleaned, walked the dog and generally cared for me so
that I could concentrate on trying to hand this in. He’s done all of this with endless patience,
kindness and love. I don’t deserve you. I promise I’ll be a better wife from now on (well, I’ll
try) - I love you so much!
“Love doesn't make the world go round. Love is what makes the ride worthwhile.”
Franklin P. Jones

To all of my lab family, thank you so much for all your help, support, advice and friendship:
Isa-Rita Russo, Mario Barbato, Dave Stanton, Mafalda Costa, Josie Jackson, Jez Smith,
Rosemary Moorhouse-Gann, Luis Cunha, Caitlin Pearson, Marian Pye, Beky Weiser, Pablo
Orozco-terWengel, Jen Stockdale, Tano Cánovas, Frank Hailer, Niall McCann, Hugh Feeley,
Pierfrancesco Sechi, Maria da Silva (Joana); Nia Thomas, Matt Dray and Steve Thomas. You
are all so amazing, you are some of the kindest people I know, so giving of your time and
(considerable) expertise, and I love you all! Particularly to Isa, Mario, Dave, Maf and Pablo –
you were always there to help me and you always knew what to do, from day 1 till the end -
thank you!
“I’ll get by with a little help from my friends.” The Beatles
Thank you to my supervisors, Mike Bruford, Hefin Jones and Steve Ormerod, firstly for giving
me this opportunity in the first place, and for your continued help and guidance along the
way. It was also a pleasure to work within the DURESS team lead by Isabelle Durance.
To my parents and whole family (Burtons and Macdonalds), thank you for your love and
constant support; telling me that I could do it and understanding that I couldn’t visit as
much as I wanted to.
“In every conceivable manner, the family is link to our past, bridge to our future.”
Alex Haley

Small shout out to J.K Rowling and Stephen Fry for the Harry Potter audiobooks which kept
me sane through all the late nights and weekends alone in the lab completing the 12 000+
PCRs.
My dog, Merry (full name Meriadoc Brandybuck Macdonald), who got me out of the house
for a bit of perspective and was always there for a cuddle, he also made working from home
a lot less lonely.
“Happiness is a warm puppy.” Charles M. Schulz

And lastly to my unborn son, you gave me the strictest deadline that any PhD student could
have, and I’m so grateful, without you I’d have been dragging my feet. For you, I tried to
stay (relatively) calm, because I didn’t want you to have physiological issues because your
mum was stressed during pregnancy; you made me look after myself (without you I’d have
survived on cereal, illuminous coloured tinned soup, and caffeine); and I was kept sane
during writing up because I could think of the amazing new adventure you, me, your dad
and Merry will be starting when you arrive! I’m so looking forward to meeting you! Not long
now! I love you. Mum x
“Babies are such a nice way to start people.” Don Herold

Myself and this project was funding by The President Research Scholarship from Cardiff
University and NERC funded DURESS project.
Table of Contents

List of Figures .................................................................................................................. ix
Chapter 1 ........................................................................................................................... ix
Chapter 2 ........................................................................................................................... ix
Chapter 3 ........................................................................................................................... xi
Chapter 4 .......................................................................................................................... xii
Chapter 5 .......................................................................................................................... xii
List of Tables ..................................................................................................................xiv
Chapter 1 ......................................................................................................................... xiv
Chapter 2 ......................................................................................................................... xiv
Chapter 3 ......................................................................................................................... xvi
Chapter 4 ........................................................................................................................ xvii
Chapter 5 ........................................................................................................................ xvii
Thesis Summary ............................................................................................................. xix
Chapter 1 - Introduction ................................................................................................... 1
Chapter 1 - Part 1 ............................................................................................................. 2
1.1 General introduction ........................................................................................................ 2
1.1.1 Why do we care about biodiversity? ......................................................................... 2
1.1.2 Importance of freshwater ecosystems ...................................................................... 3
1.1.3 Freshwater invertebrates .......................................................................................... 4
1.1.4 Importance of genetic studies ................................................................................... 5
1.2 Main aims of study ........................................................................................................... 6
1.3 Study species .................................................................................................................... 7
i
1.3.1 Amphinemura sulcicollis ........................................................................................ 9
1.3.2 Isoperla grammatica ............................................................................................ 10
1.3.3. Baetis rhodani ..................................................................................................... 12
1.4 Why microsatellites were chosen as the genetic marker .............................................. 13
1.5 Invertebrate genetics in freshwater environment ......................................................... 15
Chapter 1 - Part 2 ........................................................................................................... 25
Species-Genetic Diversity Correlation Conceptual Framework ......................................... 25
1.6.0 Abstract ....................................................................................................................... 25
1.6.1 Introduction................................................................................................................. 26
1.6.1.1 SGDC Theory ......................................................................................................... 26
Case one: How can heterogeneous environments cause parallel responses? ............ 27
Case two: How could genetic diversity affect species diversity? ................................. 28
Case three: How could species diversity affect genetic diversity? ............................... 31
1.6.2 Conclusions .............................................................................................................. 32
1.7 Empirical studies ............................................................................................................ 33
1.7.1 Studies showing a positive SGDC ......................................................................... 34
1.7.2 Studies showing a negative SGDC ........................................................................ 38
1.7.3 Studies showing no SGDC .................................................................................... 39
1.8 Limitations/ knowledge gaps ...................................................................................... 42
1.8.1 Species ................................................................................................................. 42
1.8.2 Genetic analysis ................................................................................................... 43
1.9 Conclusion ...................................................................................................................... 45
Chapter 2 - Methods ....................................................................................................... 52
Highlights .............................................................................................................................. 52
2.0 Abstract .......................................................................................................................... 53
2.1 Site Selection .................................................................................................................. 54
2.1.1 All Sites Visited ........................................................................................................ 54
ii
2.1.2 Choosing the final sites used for genetic analysis ................................................... 55
2.1.2.1 Amphinemura sulcicollis ................................................................................... 56
2.1.2.2 Isoperla grammatica ......................................................................................... 56
2.1.2.3 Baetis rhodani ............................................................................................... 56
2.2 Genetic Methods ............................................................................................................ 62
2.2.1. Sample collection.................................................................................................... 62
2.2.2 Extraction methods ................................................................................................. 62
2.2.3 DNA Barcoding ......................................................................................................... 64
2.2.3.1 Sequencing and analysis methods .................................................................... 65
2.2.3.2 Results and discussion ...................................................................................... 67
2.2.3.2.1 Amphinemura sulcicollis ............................................................................ 67
2.2.3.2.2 Isoperla grammatica .................................................................................. 67
2.2.3.2.3 Baetis rhodani ............................................................................................ 68
2.2.3.3 Cryptic species discrimination tool for B. rhodani ............................................ 69
2.3 Next Generation Sequencing ......................................................................................... 74
2.3.1 PrimerPipeline development ................................................................................... 75
2.4 Developing novel microsatellite markers ...................................................................... 75
2.4.1 Method .................................................................................................................... 76
2.4.2 Data analysis ............................................................................................................ 77
2.4.3 Results and discussion ............................................................................................. 77
2.5 Final Dataset ................................................................................................................... 78
2.5.1 Choosing Final Microsatellites ................................................................................. 78
2.5.2 Data quality .............................................................................................................. 79
2.5.3 Binning alleles .......................................................................................................... 80
2.6 Null Alleles ...................................................................................................................... 84
2.6.1. Introduction ............................................................................................................ 84
2.6.2 Data analysis ............................................................................................................ 85
iii
2.6.3 Results ...................................................................................................................... 85
2.6.4 Further investigation into effect on genetic structure ............................................ 91
2.6.4.1 Data analysis ..................................................................................................... 91
2.6.4.1 Results ............................................................................................................... 92
2.6.5 Discussion ................................................................................................................ 96
Chapter 3 - Genetic Structure and Diversity of Three Aquatic Invertebrates: Response to
Environmental Stressors ................................................................................................. 99
Contributions ........................................................................................................................ 99
Highlights .............................................................................................................................. 99
3.0 Abstract ........................................................................................................................ 100
3.1 Introduction .................................................................................................................. 101
3.2 Methods ....................................................................................................................... 105
3.2.1 Sampling and genotyping ...................................................................................... 105
3.2.2 Stressors ................................................................................................................ 105
3.3 Statistical analysis ......................................................................................................... 106
3.3.1 HWE and descriptive statistics .............................................................................. 107
3.3.2 Genetic structure ................................................................................................... 108
3.3.3 Genetic diversity correlations................................................................................ 108
3.3.3.1 Genetic diversity ............................................................................................. 108
3.3.3.2 Stressors .......................................................................................................... 109
3. 4 Results ......................................................................................................................... 110
3.4.1 HWE and descriptive statistics .............................................................................. 110
3.4.2 Genetic structure ................................................................................................... 114
3.4.3 Genetic diversity correlations................................................................................ 121
3.4.3.1 Genetic diversity ............................................................................................. 121
3.4.3.2 Stressors .......................................................................................................... 122
3.5. Discussion .................................................................................................................... 131
iv
3.5.1 HWE and descriptive statistics .............................................................................. 131
3.5.2 Population genetic structuring .............................................................................. 132
3.5.3 Genetic diversity and correlations with stressors ................................................. 134
Chapter 4 - The Relationship between Species and Genetic Diversity in Freshwater
Invertebrates ............................................................................................................... 138
Contributions ...................................................................................................................... 138
Highlights ............................................................................................................................ 138
4.0 Abstract ........................................................................................................................ 139
4.1 Introduction .................................................................................................................. 140
4.2 Methods ....................................................................................................................... 143
4.2.1 Invertebrate sample collection.............................................................................. 143
4.2.2 Species diversity indices ........................................................................................ 144
4.3 Statistical analysis ......................................................................................................... 145
4.3.1 Species genetic diversity correlation ..................................................................... 145
4.3.2 Species diversity vs environmental stressor.......................................................... 146
4.4 Results .......................................................................................................................... 146
4.4.1 Species genetic diversity correlation ................................................................. 146
4.5 Discussion ..................................................................................................................... 154
Chapter 5 - A preliminary investigation of demographic resilience in three invertebrate
species ......................................................................................................................... 158
Highlights ............................................................................................................................ 158
5.0 Abstract ........................................................................................................................ 159
5.1 Introduction .................................................................................................................. 160
5.2 Methods ....................................................................................................................... 161
5.3 Statistical analysis ......................................................................................................... 162
5.3.1 BOTTLENECK .......................................................................................................... 162
5.3.2 MSVAR ................................................................................................................... 163
5.3.3 MPVal ..................................................................................................................... 165
v