defence of honey bee (apis mellifera)

147 Pages·2015·4.34 MB·English

DEFENCE OF HONEY BEE
(APIS MELLIFERA) DRONE
AGAINST SEXUALLY
TRANSMISSIBLE PARASITE
NOSEMA APIS

YAN PENG
BSc (Hon) Murdoch University

This thesis is presented for the degree of Doctor of Philosophy of the University of
Western Australia
School of Animal Biology

December 2014
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DECLARATION

I declare that this thesis is my own account of my research and contains as its main
content work which has not previously been submitted for a degree at any tertiary
education institution

______________________
Yan Peng

Project supervisor:
Professor Boris Baer
Winthrop professor Harvey Millar
Professor Leigh Simmons
Dr. Robert Manning (Department of Agriculture and Food, Western Australia)

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This has been made possible by the sacrifices of many female worker bees, queen
bees and male bees (drones).

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ACKNOWLEDGEMENTS
This thesis and brilliant experience would not have been made possible without the
collective efforts of my supervisors and colleagues from the various Schools and
Centres from The University of Western Australia (UWA), and my supportive family.
I consider myself very lucky as I have four very supportive, intelligent and
kind supervisors, Prof. Boris Baer (coordinating supervisor), Winthrop Prof. Harvey
A. Millar, Prof. Leigh Simmons, and Dr. Rob Manning (external supervisor from the
Department of Agriculture and Food of Western Australia). They have guided me in
areas of evolutionary biology, biochemistry, proteomics, statistics, experimental
design and much more. It’s extremely nice to have Prof. Baer as my supervisor as he
respected my ideas and gave me a lot of freedom to do the experiments I wanted and
trusted me enough to give me that freedom, because of that, I think I have learnt a lot
more than I otherwise would if a project was just handed to me. I admire Winthrop
Prof. Millar very much as he can think of simple ways to solve complex problems. Dr.
Manning has helped with the experimental design for a number of my experiments
and is always a good source of knowledge and wisdom about honey bees.
The Centre of Integrative Research (CIBER) is made up of a family of
researchers who is passionate about the work they do and experts in their own right.
Everyone is talented (and humble about it), kind, and friendly and I am very fortunate
to be part of such a family. I need to dedicate special thanks to Tiffane Bates for her
incredible work as the bee yard manager, who has always fulfilled my request for
drones and worker bees and does this with the bee’s welfare in mind. Often, my
experiments involve painting and feeding thousands of bees and the team has always
volunteered to help when they can, those include Dr. Barbara Baer-Imhoof, Dr. Julia
Grassl, Ellen Paynter, Dr. Lori Lach, Dr. Ryan Dosselli, Dr. Rodolfo Jaffe, Dr. Mat
Welch, Dr. Susanne den Boer, Dr Tamara Hartke and other CIBER members.
During my last 1.5 years of PhD, I have again been blessed to receive guidance
from Dr. Grassl as her knowledge and persistence has help push me to explore more
ideas and this resulted in significant findings and understanding of my own data and
also helped me carry out more elegant experiments to answer hard questions. Although,
not official, I consider her as my fifth and very important supervisor.
It’s been a real privilege to be part of the Centre of Excellence for Plant Energy
Biology (PEB) under Winthrop Prof. Harvey’s team. I would like to thank Dr. Ralitza
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Alexova, Dr. Ricarda Fenske, Dr. Holger Eubel, and Dr. Clark Nelson for their help
with proteomics, Dr. John Bussell for his help with microscopy, Ian Castleden and
Hayden Walker for all things computational, Dr. Elke Stroeher for teaching me DNA
extraction and PCR, Dr. Shaobai Huang, Dr. Richard Jacoby, Dr. Tiago Tomaz, and
Dr. Nicolas Taylor for other skills. I also like to thank my office mates for the good
times I had in the office, those are, Dr Yunshin Sew, Dr Rachel Shingaki-Wells, Dr.
Richard Jacoby, Dr Ellen Paynter, and to-be doctors, Sandra Kerbler and Leila
Heidarvand.
Apart from being taught invaluable skills by all the researchers and students of
past and the present, the laboratory has access to State-of-the-Art equipment and tools
- something I have often taken for granted until I go to other laboratories and realize
with horror (or as diplomatic researchers would say “great surprise”) at how lucky I
was. The administrative staffs, Judith Moyle (now retired), Jennifer Gillett, Deborah
Yeoman, and Geetha Shute have made my PhD a breeze because whenever I need any
reagent, equipment or travel and accommodation arrangement for conferences, I just
need to tell them what I need and they organize everything. I think all research Centres
should thrive to have such efficient and dedicated administrators. Other unsung heroes
I like to thank is Rosemarie Farting and Lyn Loh for their work doing all the
autoclaving (and other work) which help make the Centre go round!
Also from PEB, I like to thank people of the Prof. Ian Small’s laboratory for
their help with PCR and other DNA related skills, such as Dr. Catherine Colas Des
Francs-Small, Dr. Kate Howell and Dr. Sandra Tanz. Although I was part of Winthrop
Prof. Harvey’s team, I am physically located in the Prof. Small’s laboratory, so I thank
Prof. Small for giving me the bench space and all the members for being so kind to
me for the time I was there.
From the Center for Microscopy, Characterisation and Analysis, I would like
to thank Assoc. Prof. Matthew Linden, Tracey Lee-Pullen, Prof. Kathy Heels, and Dr.
Paul Rigby for their help with the use of the flow cytometer and various microscopes.
From the School of Animal Biology, I would also like to thank Thomas Stewart and
Michael Archer for patiently teaching me and helping me with various aspects of
histology.
Throughout the different projects, I had many statistical queries and it is Assoc.
Prof Kevin Murray, Mrs. Laura Firth and Mr. Martin Firth from the Centre of Applied
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Statistics (UWA) and Mr. Mario D'Antuono from the DAFWA whom has given their
time and effort in helping me and I am grateful for that.
Lastly, but not least, I need to thank my loving husband, Liang Huang for his
unconditional love and support during my studies. I was fortunate enough to become
a mother during my PhD studies and Liang’s parents, Shuping Huang and Cunhe
Huang has sacrificed their valuable time to live in Australia to give the very best care
and love to our daughters, Joy Qingwan Huang and Jasmine Qingyuan Huang.
Without their support, I would not have had such a carefree time to pursue my studies,
therefore their support has been truly invaluable and irreplaceable.

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DECLARATION AND STATEMENT OF
CONTRIBUTION
This thesis contains published work and/ or work prepared for publication, some of
which has been co-authored. The bibliographical details of the work and where it
appears in the thesis are outlined below.

Published
PENG, Y., LEE-PULLEN, T. F., HEEL, K., MILLAR, A. H. & BAER, B. 2014.
Quantifying spore viability of the honey bee pathogen Nosema apis using flow
cytometry. Cytometry Part A, 85, 454-462.
This peer reviewed manuscript is presented as Chapter Three of this thesis.
I undertook all the experimental work in this paper, receiving some
technical training from Kathy Heels and Tracey Lee-Pullen. I conducted
all the data analysis with some guidance from Boris Baer. I wrote the
manuscript, which was subsequently edited by all the co-authors within
this manuscript.

PENG, Y., BAER-IMHOOF, B., MILLAR, A. H. & BAER, B. 2015. Consequences
of Nosema apis infection for male honey bee and their fertility. Scientific Reports, 5,
doi:10.1038/srep10565.
This manuscript has been submitted and is presented as submitted to
Scientific Reports to form Chapter Two. I wrote 80 % of this manuscript
and performed 80 % of experimental work and statistical analysis, while
co-author Barbara Baer-Imhoof performed another 20 % of the
experimental work, manuscript writing and statistical analysis for a small
section of this work. The manuscript was subsequently edited by all co-
authors.

Submitted
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PENG, Y., GRASSL, J., MILLAR, A. H. & BAER, B. Seminal fluid of honeybees
contain multiple mechanism to combat infections of the sexually transmitted
pathogen Nosema apis. Proceedings B.
This manuscript is currently in preparation, and 90 % of the content within
this paper will form the whole of Chapter Four. I conducted 90 % of the
experimental work and wrote the 90 % of the manuscript and carried out
the statistical analysis with some assistance from Boris Baer and Julia
Grassl. This manuscript was subsequently edited by all co-authors.

In preparation
PENG, Y., GRASSL, J., MILLAR, A. H. & BAER, B. 2014. Nosema apis infection
compromises antimicrobial effect of honey bee seminal fluid. In preparation.
This manuscript is currently in preparation, and 80 % of the content within
this paper will form the whole of Chapter Five. I conducted 80 % of the
experimental work and wrote the 90 % of the manuscript. This manuscript
was subsequently edited by all co-authors.

Student Signature

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Coordinating Supervisor Signature

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