On the red algal genus grateloupia in the Gulf of Mexico, featuring the organellar genomes of

80 Pages·2015·6.05 MB·English

ON THE RED ALGAL GENUS GRATELOUPIA IN THE GULF OF MEXICO,

FEATURING THE ORGANELLAR GENOMES

OF GRATELOUPIA TAIWANENSIS

by

MICHAEL SCOTT DEPRIEST, JR.

JUAN M. LÓPEZ-BAUTISTA, COMMITTEE CHAIR
DEBASHISH BHATTACHARYA
PHILLIP M. HARRIS
MARTHA J. POWELL
AMELIA K. WARD

A DISSERTATION

Submitted in partial fulfillment of the requirements
for the degree of Doctor of Philosophy
in the Department of Biological Sciences
in the Graduate School of
The University of Alabama

TUSCALOOSA, ALABAMA

2015
Copyright Michael Scott DePriest, Jr., 2015
ALL RIGHTS RESERVED
ABSTRACT

Red algae (Rhodophyta) are economically useful for their gelling compounds,
ecologically critical to marine benthic systems, and evolutionarily poised at the intersection of
primary and secondary endosymbiotic lineages. Molecular sequencing has transformed our
understanding of red algae, revealing genetic and genomic characteristics that had once been
completely unknown. In Grateloupia, a red algal genus that is morphologically simple and
notoriously difficult-to-identify, sequencing has greatly assisted in identification of species and
phylogenetic placement of troublesome taxonomic groups. However, analysis of DNA has also
proven useful for genomic comparisons on a larger scale, in order to resolve deep evolutionary
questions in terms of overall genome architecture and gene content. Grateloupia is a prime
candidate for genomic research, representing an order that had previously not been explored. In
this study, sequencing-based analyses were applied at both levels, examining species of
Grateloupia both within the genus and from a greater phylogenetic perspective. Phylogenetic
analysis of the rbcL marker revealed the previously unknown species Grateloupia taiwanensis,
first reporting this non-native alga from the Gulf of Mexico, and it showed that the species
previously known as Grateloupia filicina in the Gulf of Mexico actually includes several species.
The organellar genomes of Grateloupia taiwanensis were also sequenced and annotated; both the
plastid and mitochondrial genome are typical of florideophyte red algae in size, gene content,
and structure. Mauve genome alignments demonstrated a pattern of genomic rearrangements
expected given the overall phylogeny of Rhodophyta.

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LIST OF ABBREVIATIONS
aa Amino acid(s)
BI Bayesian inference (of phylogeny)
bp Base pair(s)
cox1 Cytochrome c oxidase subunit alpha gene
DCJ Double-cut-and-join (in Mauve alignments)
e or e-value Expect value, used as a statistical significance threshold in BLAST results
GC Guanine-cytosine content
kb, kbp Kilobase-pairs (1,000 bp)
LCB Locally collinear block (in Mauve alignments)
ML Maximum likelihood (phylogenetic analysis)
ORF Open reading frame
rbcL Rubisco-1,5-bisphosphate carboxylase gene
s.l. Sensu lato; in the broad sense (in taxonomy)
s.s. Sensu stricto; in the strict sense (in taxonomy)
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ACKNOWLEDGMENTS

First and foremost, I would like to thank the chairperson of this dissertation, Juan López-
Bautista, for inspiring me with the fascinating world of algae since 2008, for his always useful
advice on algal and academic issues, and for his patience with me during the preparation of this
and other manuscripts. I thank my Ph.D. committee members—Debashish Bhattacharya, Phillip
Harris, Martha Powell, and Amy Ward—all of whom have provided unique perspectives and
recommendations for my graduate career, and from whom I have learned very much. I also thank
my labmates Gabriela Garcia-Soto, Daryl Lam, Trey Melton, Ana Tronholm, and David Ward,
who have graciously assisted me with collection trips, new samples of Grateloupia, and/or
phylogenetic and genomic analyses. I thank all of my past undergraduate assistants in the
PhycoLab, who spent many hours helping me with DNA extractions and reactions. Finally, I
sincerely thank Mike Wynne, for his friendly and very detailed advice on the taxonomy of
various groups of algae over the years.
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CONTENTS
ABSTRACT .................................................................................................................................... ii
LIST OF ABBREVIATIONS ........................................................................................................ iii
ACKNOWLEDGMENTS ............................................................................................................. iv
LIST OF TABLES ......................................................................................................................... vi
LIST OF FIGURES ...................................................................................................................... vii
1. Sequencing of the rbcL Marker Reveals the Nonnative Red Alga Grateloupia taiwanensis
(Halymeniaceae, Rhodophyta) in Alabama .....................................................................................1

2. Grateloupia filicina Revisited: Molecular Phylogenetics of Grateloupia (Halymeniaceae,
Rhodophyta) from the Gulf of Mexico ..........................................................................................14

3. The Plastid Genome of the Red Macroalga Grateloupia taiwanensis (Halymeniaceae) ..........27

4. The Mitochondrial Genome of Grateloupia taiwanensis (Halymeniaceae, Rhodophyta) and
Comparative Mitochondrial Genomics of Red Algae ...................................................................46

APPENDIX ....................................................................................................................................70
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LIST OF TABLES
1.1. List of Species Included in Phylogenetic Analysis of Grateloupia taiwanensis ....................13
2.1. List of Species Included in Phylogenetic Analysis of Grateloupia filicina ............................25
2.2. rbcL Pairwise Distances for Selected Grateloupia Species ...................................................26
3.1. Characteristics of Selected Red Algal Plastid Genomes .........................................................42
3.2. List of genes in G. taiwanensis Plastid Genome.....................................................................43
3.3. Comparison of tRNA Sequences in Red Algal Plastid Genomes ...........................................44
3.4. Novel ORFs in the G. taiwanensis Plastid Genome ...............................................................45
4.1 Characteristics of Selected Red Algal Mitochondrial Genomes ..............................................68
4.2. Comparison of Protein-coding Genes in Red Algal Mitochondrial Genomes .......................69

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LIST OF FIGURES
1.1. Phylogenetic Tree of G. taiwanensis and Related Taxa .........................................................11
1.2. Photograph of G. taiwanensis .................................................................................................12
2.1. Phylogenetic Tree of G. filicina and Related Taxa .................................................................24
3.1. Phylogenetic Tree of Division Rhodophyta ............................................................................39
3.2. G. taiwanensis Plastid Genome ..............................................................................................40
3.3. Pairwise Mauve Genome Alignments of Red Algal Plastid Genomes ...................................41
4.1. G. taiwanensis Mitochondrial Genome ..................................................................................62
4.2. Mauve Genome Alignment of G. taiwanensis and G. angusta ..............................................63
4.3. Mauve Genome Alignment of G. taiwanensis with Rhodymeniophycidae ...........................64
4.4. Mauve Genome Alignment of G. taiwanensis and Sporolithon durum ..................................65
4.5. Mauve Genome Alignment of G. taiwanensis and Pyropia haitanensis ................................66
4.6. Mauve Genome Alignment of G. taiwanensis and Cyanidioschyzon merolae ......................67
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Sequencing of the rbcL Marker Reveals the Nonnative Red Alga Grateloupia taiwanensis
(Halymeniaceae, Rhodophyta) in Alabama

ABSTRACT
Mobile Bay, AL has been the site for the introduction of several terrestrial and freshwater
invasive species, including red imported fire ants (Solenopsis invicta) and spike-topped apple
snails (Pomacea bridgesii). The Gulf of Mexico has also been invaded by several marine animal
species, such as zebra mussels (Dreissena polymorpha). To date, no invasive marine macroalga
has been reported in the Mobile Bay area. However, recent collections of an unusual species of
Grateloupia (Halymeniaceae, Rhodophyta) in Alabama indicate that an introduction has been
made. On the basis of phylogenetic analysis of the large subunit of ribulose-1,5-bisphosphate
carboxylase/oxygenase (rbcL) marker, the species has been identified as Grateloupia
taiwanensis S.M. Lin & H.Y. Liang. This is the first report of G. taiwanensis outside its native
range.
INTRODUCTION
Grateloupia C. Agardh is a genus of benthic marine red algae (Rhodophyta), currently
containing about 90 species (Guiry and Guiry, 2012). It is the largest genus in the family
Halymeniaceae. Species of this genus occur throughout the world in warm temperate to tropical
marine waters. Several Grateloupia species are known in the Gulf of Mexico, specifically
Grateloupia gibbesii Harvey and Grateloupia pterocladina (M.J. Wynne) S. Kawaguchi & H.W.
Wang, as well as many reports of unidentified Grateloupia species (see Fredericq et al., 2009).
Wynne (2011) listed a total of 11 species in the western Atlantic, including G. gibbesii, G.
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pterocladina, and Grateloupia filicina (J.V. Lamouroux) C. Agardh. Wynne, however,
remarked that past identifications of G. filicina in the western Atlantic, including the Gulf of
Mexico, are doubtful due to the results of De Clerck et al. (2005b), which indicated that true G.
filicina may be restricted to the Mediterranean Sea and Macaronesia. This suggests that tropical
collections of G. filicina actually belong to a different species.
The genus Grateloupia is known for having simple morphologies that make
distinguishing species difficult, and DNA sequencing has been instrumental in generic and
species-level circumscriptions (e.g. Wang et al., 2001; De Clerck et al., 2005b; Lin et al., 2008).
Specimens previously identified as G. filicina, frequently reported throughout the world, actually
show an unexpectedly high amount of genetic diversity and are therefore morphologically static.
As a result of G. filicina being demonstrated to be polyphyletic, several new species have been
split from the group (e.g., Kawaguchi et al., 2001; De Clerck et al., 2005b). Even in the past few
years, the genus has gained many new species, including Grateloupia huangiae S.-M. Lin & H.-
Y. Liang, Grateloupia dalianensis H.W. Wang & D. Zhao, and Grateloupia yinggehaiensis H.W.
Wang & R.X. Luan. The publications in which these species are described (Lin and Liang, 2011;
Zhao et al., 2012) include molecular phylogenetic analyses to more clearly delineate these taxa.
Additional taxonomic work is definitely necessary to continue to resolve systematic
inconsistencies and to account for unexpected, newly discovered diversity in the genus
Grateloupia.
In addition to these taxonomic concerns, it is important to consider that Grateloupia
contains species that are known to be aggressively invasive, most notably Grateloupia turuturu
Yamada. Grateloupia turuturu, along with several other Grateloupia species, has been
introduced in Italy (Cecere et al, 2011), New Zealand (D’Archino et al., 2007), Great Britain
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