Molecular diet study of Adélie penguins (Pygoscelis adeliae) and Weddell seals (Leptonychotes weddellii) in the Ross Sea using fecal DNA
- Alternative Title
- 분변 DNA를 이용한 로스해에 서식하는 아델리펭귄과 웨델해표의 분자식성 연구
- Abstract
- Antarctica’s Ross Sea is the biodiversity hotspot because of its vibrant and unique marine life and environment. The extreme marine environment of the Southern Ocean varies spatially and seasonally. Top predators play an essential role in a food web, and their diet can provide crucial information about even minor changes in the environment. DNA-based research is a non-invasive tool for overcoming traditional methods for analyzing dietary habits, which have devastating consequences on animals and their habitats. The diets of two top predators, Weddell seals (Leptonychotes weddellii) and Adélie penguins (Pygoscelis adeliae), were examined in this study using two molecular techniques, Next-Generation Sequencing (NGS) metabarcoding and quantitative PCR (qPCR) technology on fecal DNA. The study of the diet of P. adeliae was carried out using universal primers 18Sv9 and notothenioid-specific primers (miniFish) and species-specific assays. For penguins, 143 samples were collected from 7 stations. Two krill species (Euphausia superba and E. crystallorophias) and nototheniid fish were among the most abundant diets of Adélie penguin. Pleuragramma antarctica and Pagothenia borchgrevinki were the two most abundant species among the nototheniid preys. The results of the qPCR analysis showed a significant geographical difference in prey composition. The diet of Weddell seals was examined for total 93 scat samples were collected from 4 stations and for four years, from 2016 to 2019. The 18Sv9 rRNA metabarcoding results showed the main prey of L. weddellii belong to the phylum Chordata (fish and penguins) and Arthropoda (Crustacea), with minor contributions from Echinodermata and Cnidaria. The ‘miniFish’ metabarcoding revealed 22 fish species as the seal's diet, further classified into the families Nototheniidae, Artedidraconidae, Channichthyidae, Bathydraconidae, and Harpagiferidae. The use of qPCR assisted in determining the proportion of significant prey in different diets of the predator. Many parasites were detected in addition to the diet components. The penguin's diet was dominated by krills, whereas fish dominated the seal's diet. The research yielded a variety of intriguing, expected, and unexpected results, described in contrast to the literature and methodological limitations observed. The molecular diet analysis delivered crucial information on the role and behavior of generalist predators in various food webs and ecosystems. The data generated in this study could help monitor the world’s largest Marine Protected Area (MPA) in an Antarctic marine ecosystem.
- Issued Date
- 2022
- Awarded Date
- 2022. 2
- Type
- Dissertation
- Keyword
- Antarctica diet study Leptonychotes weddellii metabarcoding molecular analysis NGS Pygoscelis adeliae Ross Sea
- Publisher
- 부경대학교
- Affiliation
- Pukyong National University, Graduate school
- Department
- 대학원 4차산업융합바이오닉스공학과
- Advisor
- Hyun -Woo Kim
- Table Of Contents
- CHAPTER 1 General Introduction 1
1.1. Significance of fecal DNA in molecular ecological studies 2
1.2. Study of Fecal DNA using NGS metabarcoding technology and qPCR 3
1.3. Adélie penguins (Pygoscelis adeliae) and Weddell seals (Leptonychotes weddellii): Indicator species for monitoring Antarctic marine ecosystem 4
1.4. Relevance of the current study 6
1.5. The objectives of the study 8
1.6. Brief overview of the study 8
CHAPTER 2 Molecular diet study of Adélie penguins (Pygoscelis adeliae) in the Ross Sea region using fecal DNA 10
2.1. Abstract 11
2.2. Introduction 12
2.3. Materials and Methods 14
2.3.1. Sample collection and fecal DNA extraction 14
2.3.2. Primer design and PCR amplification of prey DNA 18
2.3.3. Library construction and NGS 20
2.3.4. Bioinformatic analysis 21
2.3.5. qPCR analysis 22
2.4. Results 23
2.4.1. Fecal DNA metabarcoding of Pygoscelis adeliae in the Ross Sea using 18Sv9 universal primers 23
2.4.2. Fecal DNA metabarcoding of Pygoscelis adeliae in the Ross Sea using miniFish primers 33
2.4.3. qPCR analysis of three main prey items 37
2.5. Discussion 43
CHAPTER 3 Molecular diet study of Weddell seals (Leptonychotes weddellii) in the Ross Sea region using fecal DNA 50
3.1. Abstract 51
3.2. Introduction 52
3.3. Materials and Methods 55
3.3.1. Sample collection and fecal DNA preparation 55
3.3.2. Primer design of prey items for metabarcoding and qPCR analysis of the diet of seal: Barcode for coverage and resolution 59
3.3.3. Library construction and Next Generation Sequencing (NGS) 64
3.3.4. Bioinformatic analyses 65
3.3.5. Quantitative PCR (qPCR) workflow and analysis 66
3.4. Results 68
3.4.1. Fecal DNA metabarcoding of Weddell seals (L. weddellii) in the Ross Sea using 18Sv9 universal primers 68
3.4.2. Fecal DNA metabarcoding of Weddell seals (L. weddellii) in the Ross Sea region using miniFish primers 70
3.4.3. The qPCR analysis for Pagothenia borchgrevinki, Pleuragramma antarctica, Trematomus spp., Channichthydae, Adélie Penguin, Notocrangon antarticus, and miniFish (Notothenioid) in seal’s diet. 80
3.4.4. qPCR for parasites Spirometra erinaceieuropaei and Contracaecum osculatum 82
3.4.5. Nonmetric multidimensional scaling NMDS plot of qPCR results 82
3.5. Discussion 89
3.5.1. Primers and metabarcoding results of 18sV9 and minifish assays 89
3.5.2. Cryo-pelagic fish Pagothenia borchgrevinki inhibiting underside sea ice 90
3.5.3. Distribution of Notothenioid fish in the Terra Nova Bay, the Ross Sea 92
3.5.4. Benthic preys: Notocrangon antarcticus (arthropod), Trematomus bernacchii, Pogonophryne sp., Gymnodraco sp. (notothenioids) 94
3.5.5. Non- detection of some major components of seal diet 95
3.5.6. Adélie penguin (Pygoscelis adeliae) are occasional prey 96
3.5.7. Factors responsible for diverse prey selection including season, prey abundance, seasonal availability, and foraging behavior 97
3.5.8. Quantification of main prey items 98
3.5.9. Parasites 99
3.6. Conclusion 100
CHAPTER 4 Discussion on combined Results 102
4.1. Diet profile, foraging habitat, prey distribution and behavior of Adélie penguin (Pygosceles adeliae) and Weddell seals (Leptonychotes weddellii) 103
4.2. Molecular marker selection for NGS metabarcoding and quantification 109
4.3. Limitations of the study 113
4.4. Conclusion and application of the study 115
CHAPTER 5 Future directions for fecal DNA metabarcoding 117
5.1. Strategies for samples collection and storage 118
5.2. Library preparation, PCR, and NGS sequencing platform 121
5.3. Bioinformatics process for high throughput sequencing data analysis of fecal DNA 123
Abbreviations 125
References 126
Abstract in the Korean Language 169
Acknowledgments 171
- Degree
- Doctor
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- 대학원 > 4차산업융합바이오닉스공학과
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