Browsing by Author "Koh, Yiling Eileen"

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Phototrophic Bacteria in Antarctic Sea Ice
(Te Herenga Waka—Victoria University of Wellington, 2011) Koh, Yiling Eileen; Ryan, Ken; Davy, Simon
Polar sea ice plays host to a number of life forms that are critical to the marine ecosystem and serve as important platforms supporting other organisms. Despite its austere appearance, sea ice is a complex habitat. Most of the prokaryotes in sea ice are heterotrophic bacteria and are known to play a significant role in sea ice carbon cycling and food web dynamics. Phototrophy plays an important biological role on Earth and only two mechanistically distinct processes exist for the harvesting and conversion of solar energy to chemical energy in bacteria. The first uses retinal-binding proteins that function as light-driven proton or chloride pumps. The second phototrophic mechanism is dependent upon photochemical chlorophyll-containing reaction centres in chlorophototrophs. The main focus of this research is the elucidation of phototrophic bacteria in the antarctic sea ice matrix, which has not been studied before. Environmental samples consisting of sea ice and brine were collected from five locations in the Ross Sea Region of Antarctica. DNA and RNA (the latter translated to cDNA) were extracted from these samples and clone libraries were constructed from the PCR-amplified genes using gene-specific oligonucleotide probes. The clones were sequenced and the sequence data was used to infer phylogenetic data. Cyanobacteria are common in many habitats on Earth but no positive cyanobacteria were detected in my sea ice samples; instead the bulk of sequences detected were of diatom origin. Temperature, salinity, nutrient requirements, growth and loss rates are major eco-physiological factors that may limit the survival of polar cyanobacteria. To the best of my knowledge, this study is the first to undertake a comprehensive molecular search for cyanobacteria in sea ice. Aerobic anoxygenic phototrophs (AAnPs) utilise bacteriochlorophyll-a as their main photosynthetic pigment. The gene segment pufM, which encodes a pigmentbinding protein subunit of the reaction centre complex, is the marker gene used to determine AAnPs diversity. This is the first report of AAnP bacteria present in the Ross Sea region, and particularly in the sea ice matrix. In particular, cDNA data suggested that AAnPs are present in a metabolically active state and might contribute to the energy budget within the sea ice. Proteorhodopsins (PR) are retinal binding bacterial integral membrane proteins that function as light-driven proton pumps. I demonstrated for the first time the presence of PR-bearing bacteria from the Bacteroides and Proteobacteria phyla within the sea ice matrix over a geographical and internal spatial scale. The PR gene was also detected as cDNA transcripts and this demonstrated that PR-bearng cells were metabolically active within upper middle and bottom sections of the sea ice. Thus the PR protein might be of considerable importance to the sea ice ecosystem, although further study is needed to confirm this prediction. Conversely, no positive Actinobacteria analogues of the rhodopsin sequences were detected, although preliminary amplification produced bands of the desired size. A snapshot of the bacterial diversity in my Terra Nova Bay samples was also obtained using the 16S rDNA genetic biomarker. All the clones with contiguous sequences were aligned to the Proteobacteria, Cytophaga-Flavobacteria-Bacteroides and Actinobacteria phyla while sequence fragments showing high percentage similarities to environmental, mammal, insect and human-associated sequences were also detected. No phototrophic bacteria were identified based on this 16S rDNA gene. In the course of my research I have, for the first time, showed the presence of phototrophic bacteria in annual antarctic sea ice. And most importantly, the photosynthetic genes were found to be actively expressed by bacteria at the time of sampling, suggesting that phototrophy does have an important role in the sustainment of the sea ice microbial communities. Previous paradigms holding that chlorophyll-based organisms solely carry out photosynthesis have been challenged. Rhodopsin-based and bacteriochlorophyll phototrophy are the ‘new kids on the block’, especially in the antarctic sea ice environment. Ultimately, more physiological and quantification experiments will be needed in order to establish the ecological roles of these light-harvesting bacteria in sea ice microbial assemblages in order to understand their basal functions and their contributions to the sea ice ‘microbial loop’.