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UC San Diego UC San Diego Electronic Theses and Dissertations UC San Diego UC San Diego Electronic Theses and Dissertations Title The chemical ecology of marine bacteria and the sediments they inhabit Permalink https://escholarship.org/uc/item/7594396g Author Tuttle, Robert Publication Date 2018 Peer reviewed|Thesis/dissertation eScholarship.org Powered by the California Digital Library University of California UNIVERSITY OF CALIFORNIA SAN DIEGO The chemical ecology of marine bacteria and the sediments they inhabit A dissertation submitted in partial satisfaction of the requirements for the degree Doctor of Philosophy in Marine Biology By Robert Nicholas Tuttle Committee in charge: Professor Paul Jensen, Chair Professor Gregory Rouse Professor Pieter Dorrestein Professor Chambers Hughes Professor Stuart Sandin Copyright Robert Nicholas Tuttle, 2018 All Rights Reserved The dissertation of Robert Nicholas Tuttle is approved, and it is acceptable in quality and form for publication on microfilm and electronically: ________________________________________________________________ ________________________________________________________________ ________________________________________________________________ ________________________________________________________________ ________________________________________________________________ Chair University of California San Diego 2018 iii Dedication To my grandfather, Dr. Harry J. Woehr You taught me that greatness often comes from humble beginnings. iv EPIGRAPH Nothing in life is to be feared, it is only to be understood. Now is the time to understand more, so that we may fear less. -Marie Curie v TABLE OF CONTENTS SIGNATURE PAGE ............................................................................................................................... …..iii DEDICATION ....................................................................................................................................... …..iv EPIGRAPH. .......................................................................................................................................... …...v TABLE OF CONTENTS .......................................................................................................................... …..vi LIST OF FIGURES ................................................................................................................................. …..ix LIST OF TABLES ................................................................................................................................... …..xi ACKNOWLEDGMENTS ........................................................................................................................ ….xii VITA .................................................................................................................................................... ….xv ABSTRACT OF THE DISSERTATION ..................................................................................................... ..xvii Chapter 1 Introduction ................................................................................................................... …..1 1.1 Introduction to chemical ecology ................................................................................... …..2 1.2 Marine chemical ecology – Eukaryotes .......................................................................... …..3 1.3 Marine chemical ecology – Prokaryotes ........................................................................ …..5 1.3.1 Nutrient limitation ...................................................................................... …..6 1.3.2 Prokaryote- prokaryote allelopathy............................................................ …..7 1.3.3 Prokaryote - Eukaryote allelopathy ............................................................ …..8 1.4 Induction of bacterial specialized metabolites ............................................................. …..9 1.4.1 Cell to cell contact ....................................................................................... …10 1.4.2 Nutrient supplementation .......................................................................... …11 1.4.3 Small molecules .......................................................................................... …11 1.5 Sediment bacteria – focusing on Salinispora ................................................................. …12 1.6 Advancements in mass spectrometry ............................................................................ …14 1.7 Summary of thesis .......................................................................................................... …16 1.8 References ...................................................................................................................... …17 Chapter 2 The Detection of Specialized Metabolites and Their Producers in Ocean Sediments .. …23 2.1 Abstract .......................................................................................................................... …24 2.2 Introduction .................................................................................................................... …25 2.3 Material and methods .................................................................................................... …26 2.3.1 Locations and samples ................................................................................ …26 2.3.2 Resin extractions and analysis .................................................................... …27 2.3.3 Staurosporine quantification ...................................................................... …31 2.3.4 C. elegans feeding deterrence .................................................................... …32 2.3.5 Brine shrimp lethality assay ........................................................................ …32 2.3.6 Bacterial community analysis ..................................................................... …33 2.3.7 Bacterial cultivation, identification, and metabolomics ............................. …34 2.3.8 Metagenomics ............................................................................................ …34 2.3.9 Phylogenetic analysis ........................................................ ............ ............ …35 2.4 Results .................................................................................................. ............ ............ …35 2.4.1 Sediment metabolomics ................................................... ............ ............ …35 2.4.2 Compound identification .................................................. ............ ............ …36 vi 2.4.3 Linking compounds to community composition .............. ............ ............ …44 2.4.4 Staurosporine quantification ............................................ ............ ............ …47 2.4.5 Metagenomic analysis ...................................................... ............ ............ …49 2.4.6 The ecological roles of staurosporine ............................... ............ ............ …51 2.5 Discussion ............................................................................................. ............ ............ …52 2.6 Acknowledgments ................................................................................ ............ ............ …56 2.7 References ............................................................................................ ............ ............ …56 2.8 Appendix ............................................................................................... ............ ............ …65 Chapter 3 Salinispora secondary metabolite mediated feeding deterrence .... ............ ............ …69 3.1 Abstract ................................................................................................ ............ ............ …70 3.2 Introduction .......................................................................................... ............ ............ …71 3.3 Methods ............................................................................................... ............ ............ …75 3.3.1 Salinispora strain selection and culturing ......................... ............ ............ …75 3.3.2 C. elegans maintenance and sterilization ......................... ............ ............ …76 3.3.3 Live Salinispora spp. assay setup ...................................... ............ ............ …77 3.3.4 Large scale cultivation and extraction .............................. ............ ............ …79 3.3.5 Salinispora extract feeding deterrence of C. elegans ....... ............ ............ …79 3.3.6 Bioactivity-guided compound isolation ............................ ............ ............ …80 3.3.7 Lomaiviticin C quantification and deterrence calculation ............ ............ …81 3.3.8 Ophryotrocha habitat preference assay ........................... ............ ............ …82 3.3.9 Marine nematode assays .................................................. ............ ............ …83 3.3.10 Statistical analyses ............................................................ ............ ............ …83 3.4 Results .................................................................................................. ............ ............ …84 3.4.1 Salinispora mediated C. elegans feeding preference ....... ............ ............ …84 3.4.2 Feeding assay using marine predators ............................. ............ ............ …92 3.5 Discussion ............................................................................................
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