Using Omics and Behavioural Approaches to Understand Human Impacts on Octopods

Abstract

Our world is facing major environmental threats due to anthropogenic causes. Climate change and overfishing have put immense pressure on our ecosystems which can have irreversible consequences. Given our reliance on earth’s resources for our survival, this thesis investigates the fundamental biology and ecology of one key resource, the octopod, in a bid to understand, and thereby mitigate, the impacts of anthropogenic pressures on this charismatic and commercially important animal. Octopus berrima, the main study species in this thesis, constitutes a developing, small-scale fishery in southeast Australia. However, much information about this species remains unknown. I thus adopted an “omics” approach to examine their population genomic structure and their proteomic responses to ocean warming. I also studied their feeding behaviour to determine how adaptable they are to changing environmental conditions. To support sustainable fisheries management, I first genetically identified the octopod species harvested in South Australia as Octopus berrima and Octopus pallidus using the universal marker cytochrome oxidase III (COIII). I also found that these two morphologically similar species could be distinguished using morphometrics, which could be useful for fishers and fishery scientists. Then, using double digest restriction site-associated DNA sequencing (ddRADseq), I genetically compared intraspecific populations of these species across southeast Australia and delineated two metapopulations of O. pallidus and three metapopulations of O. berrima. This information is valuable for genomics-based management decisions to ensure the sustainability of octopod fisheries. Proteomics is a powerful yet under-utilised tool in ecology that can reveal how organisms respond to biological perturbations, such as disease and environmental stress. However, current gold standard methods for sampling and preparing tissues for proteomics analyses are not practical or feasible for many field-based ecological applications (i.e. immediately freezing samples at -80 °C). The lack of established protocols for the proteomic analyses of wild specimens, particularly obtained in remote areas, may be why proteomics is not widely used in ecology. Hence, I optimised the proteomics workflow by testing the suitability of RNAlater for the field and various parameters on wild-caught Octopus berrima tissue samples preserved on ice and in RNAlater. Proteins were detected using SDS-PAGE and liquid chromatography tandem mass spectrometry. I demonstrated that RNAlater is ideal for the field as it preserves protein integrity and I also successfully identified high numbers of proteins (> 3,500) that may contain potential biomarkers for future ecological research. While proteomic profiles were consistent across sexes and tissue types, homogenising tissues with steel beads yielded 10 % more proteins than with liquid nitrogen. These findings demonstrated the possibility of obtaining deep proteomic coverage on field-caught samples and serves as a first step to establishing proteomics protocols for studying the effects of environmental stressors on wild specimens. As a voracious and versatile predator, octopods feed on a wide variety of crustaceans, gastropods and bivalves that supports their exponentially high growth rates and “live fast, die young” life history strategy. Little research has explored behavioural plasticity with feeding in octopods despite its importance to a species’ ability to adapt to changing environmental conditions. Despite being commercially targeted, the preferred prey of Octopus berrima is not known, and neither do we know if their preference for a certain prey can be modified with imprinting. In an experimental setting, I allowed newly hatched octopods to select from three prey types to establish their baseline prey preference ranking. In a separate experiment, I exposed each group of eggs to visual and chemical stimuli of each of the three prey types, before testing their prey preference upon hatching. I found that they preferred isopods, followed by amphipods and then mussels. However, regardless of which prey type the embryos were exposed to, the resulting hatchlings retained their prey preference of isopods. These findings advance our understanding of behavioural plasticity in octopods which has ecological and evolutionary significance. With the predatory role of octopods in the food web as well as an increasing interest in utilising octopods as an alternative protein source to fish, studying the effects of ocean warming is important in understanding their capacity to adapt to future oceans, especially at a vulnerable phase of life in the embryonic stage. Hence, I exposed Octopus berrima embryos to different thermal conditions (19 °C – control, 22 °C – current summer temperature, and 25 °C – projected summer temperature in 2100) and analysed the proteome of the hatchlings using quantitative proteomics. For octopods exposed to 25 °C, I found significantly reduced levels of key proteins associated with the eyes, which suggests that future warming could impair octopod vision. I also found a corresponding increase in the abundance of proteins involved in the cellular stress response and decrease in those involved in non-essential processes like digestion, indicating severe signs of stress under future warming. This study is the first to examine proteomic responses of octopods to increasing temperatures and these findings suggest that an ecologically and commercially important species might be at risk in the face of climate change. Overall, this thesis has examined the population structures of octopods and their potential vulnerability to future oceans by using omics and behavioural approaches. New methods and insights developed from this research could assist future research on octopods, as well as conservation efforts to avoid overexploitation and extinction of these ecologically, commercially, and culturally important animals.