Type of Document	Thesis
Author	Hunter, Evan
Author's Email Address	evanhunter@hotmail.com
URN	etd-03232006-134422
Title	Microbial Community Diversity Associated With Carbon and Nitrogen Cycling in Permeable Marine Sediments
Degree	Master of Science
Department	Oceanography, Department of
Advisory Committee	Advisor Name Title Heath Mills Committee Member Joel Kostka Committee Member Lee Kerkhof Committee Member Markus Huettel Committee Member
Keywords	SSU rRNA Microbial Community Analysis Nitrogen Cycle nosZ amoA Sandy Sediments
Date of Defense	2006-03-20
Availability	unrestricted
Abstract Though a large fraction of primary production and organic matter cycling in the oceans occurs on continental shelves dominated by sandy deposits, the microbial communities associated with permeable shelf sediments remain poorly characterized. Therefore, the primary objective of this study was to provide the first detailed characterization of microbial diversity in representative marine sands of the South Atlantic Bight (SAB) and the northeastern Gulf of Mexico (NEGOM) through analyses of SSU rRNA gene (Bacteria), nosZ (denitrifying bacteria), and amoA (ammonia-oxidizing bacteria) sequences. Communities were analyzed by DNA extraction, clone library construction, and terminal restriction fragment length polymorphism (T-RFLP) community fingerprinting. Sediment characteristics, geochemical parameters, and rate measurements were obtained in parallel with microbial community analysis. Microbiological and biogeochemical approaches were coupled, allowing the structure-function relationships of key microbial groups involved in carbon and nitrogen cycling in continental shelf sediments to be examined. In the SAB study (Ch. 1), clone libraries were constructed from both sediment core material and manipulated sediment within column experiments. Rapid organic matter degradation and coupled nitrification-denitrification were observed in column experiments at flow rates and oxygen concentrations resembling in situ conditions. Numerous SSU rRNA gene phylotypes were affiliated with the phyla Proteobacteria (classes Alpha-, Delta-, and Gammaproteobacteria), Planctomycetes, Cyanobacteria, Chloroflexi and Bacteroidetes. Detectable sequence diversity of nosZ and SSU rRNA genes increased in stratified redox-stabilized columns compared to in situ sediments, with the Alphaproteobacteria comprising the most frequently detected group. Alternatively, nitrifier communities showed a relatively low and stable diversity that did not co-vary with the other gene targets. In the NEGOM study (Ch. 2), high throughput techniques were developed and applied to extensively profile overall and denitrifying microbial communities in a large number of sediment samples over various sediment depth intervals, contrasting sites, and sampling periods. Cloning/sequencing and community fingerprinting (T-RFLP) approaches were applied in parallel to characterize microbial diversity and phylogenetic composition. Statistical estimators including species richness, Shannon-Weiner and 1/D indices, nucleotide diversity, gene diversity, evenness, and theta (pi) indicated little difference between four clone libraries constructed from selected depth intervals (0-2 cm, 18-20 cm) at each site in March. In contrast, T-RFLP profiles and robust phylogenetic analysis showed distinct trends in diversity according to site, depth, and time period sampled. The results elucidate predominant phylotypes that are likely to catalyze carbon and nitrogen cycling in marine sands. Several microbial groups (Deltaproteobacteria, Gammaproteobacteria, Planctomycetes) were confirmed as significant contributors to the microbial communities of permeable marine sediments in agreement with previous work. However, the robust sequence database of this study expanded current knowledge to reveal a large overall community diversity including additional groups (Alphaproteobacteria, Bacteriodetes/Chlorobi, and Cyanobacteria) that had not been previously recognized using cultivation-independent methods with inherently lower resolution. The Alphaproteobacteria, in particular, were shown to be relatively abundant in the overall and denitrifying communities at both SAB and NEGOM sites. Although overall diversity increased in response to redox stabilization and stratification in column experiments, the major phylotypes remained the same, indicating that the columns sufficiently mimic in situ conditions. While SSU rRNA gene phylotypes detected by clonal analysis were similar at the phylum level at all sites, the NEGOM site showed much higher species richness in comparison to SAB. At NEGOM, T-RFLP showed distinct differences in community diversity according to site, depth, and time. The sequence database from this thesis will facilitate the development of improved probes and primer sets to be used in quantifying the metabolically active members of permeable sand communities. Rapid community fingerprinting methods developed here should allow for more extensive comparisons across environmental gradients in order to better understand the factors controlling microbial diversity in permeable sediments.
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