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CHAPTER 1 Bacterial Diversity In miii UNIVERSITEIT GENT Faculteit Wetenschappen Laboratorium voor Microbiologie Biodiversity of bacterial isolates from Antarctic lakes and polar seas Stefanie Van Trappen Scriptie voorgelegd tot het behalen van de graad van Doctor in de Wetenschappen (Biotechnologie) Promotor: Prof. Dr. ir. J. Swings Academiejaar 2003-2004 ‘If Antarctica were music it would be Mozart. Art, and it would be Michelangelo. Literature, and it would be Shakespeare. And yet it is something even greater; the only place on earth that is still as it should be. May we never tame it.’ Andrew Denton Dankwoord Doctoreren doe je niet alleen en in dit deel wens ik dan ook de vele mensen te bedanken die dit onvergetelÿke avontuur mogelÿk hebben gemaakt. Al van in de 2 de licentie werd ik tijdens het practicum gebeten door de 'microbiologie microbe' en Joris & Ben maakten mij wegwijs in de wondere wereld van petriplaten, autoclaveren, enten en schuine agarbuizen. Het kwaad was geschied en ik besloot dan ook mijn thesis te doen op het labo microbiologie. Met de steun van Paul, Peter & Tom worstelde ik mij door de uitgebreide literatuur over Burkholderia en aanverwanten en er kon een nieuw species beschreven worden, Burkholderia ambifaria. Nu had ik de smaak pas goed te pakken en toen Joris met een voorstel om te doctoreren op de proppen kwam, moest ik niet lang nadenken. En in de voetsporen tredend van A. de Gerlache, kon ik aan de ontdekkingstocht van ijskoude poolzeeën en Antarctische meren beginnen... Jean was hierbij een uitstekende gids: bedankt voor je vertrouwen en steun. Naast de kritische opmerkingen en verbeteringen, gafje mij ook voldoende vrijheid om deze reis tot een goed einde te brengen. Joris was mijn metgezel van begin tot einde en dankzij zijn inspirerende inzichten en on uitputtelijke motivatie, kwam ik altijd weer op hetjuiste pad terecht wanneer ik dreigde te verdrinken in de onmetelÿke oceanen van de bacteriële taxonomie. Jouw boeiende verhalen over de polen ais de meest onherbergzame uithoeken van de wereld, zittend op een terrasje op het Leidse plein, deden me dan weer even wegdromen... I would like to thank the members of the MICROMAT-project and especially the persons who gathered field samples in the Vestfold Hills, the McMurdo Dry Valleys and the Larsemann Hills. I am also indebted to Dr. Tjhing-Lok Tan for the fruitful collaboration and the opportunity he gave me to study bacteria isolated from the Arctic and Antarctic seas. I am very grateful to the members of the examination committee for their critical reading and comments. Verder wil ik ook alle collega's van het Laboratorium voor Microbiologie bedanken voor de aangename werksfeer en de tijd die velen onder hen wisten vrij te maken om mij te helpen. In het bijzonder wil ik mijn 'reisgezellen' Margo en Sylvie bedanken voor hun inspanningen bij het isoleren van de honderden stammen uit de matstalen en deze veilig op te bergen voor mij in de R-collectie. Alle mensen van de 2 de fase (Margo, Klaas, Geert, Liesbeth, Robin, Dirk, Tom, Evie, Ilse, Fabiano, Cristiane & Sabri) wil ik bedanken voor de hulp bij verschillende praktische probleempjes en de 'interessante lunchgesprekken’ die voor de nodige verstrooiing zorgden. De thesisstudenten Lars Hulpio & Ilse Vandecandelaere verdienen voor hun harde werk en inspanningen een speciaal woordje van dank. Renata wil ik bedanken voor de hulp bij het aanleren van de Marmur-techniek en het vele sequentiewerk, Cindy en Ilse Cleenwerk voor hun raad bij de DNA-DNA hybridisaties en Liesbeth Lebbe voor de %GC bepalingen. Ook Karei Kersters dient te worden bedankt voor de hulp bij het nalezen van verschillende manuscripten. Annemie, Geert, Jeanine & Fernand wil ik bedanken voor hun administratieve en logistieke steun en ook Claudine verdient een speciaal woordje van dank voor haar geduld ais ik weer eens met één van mijn moeilijk te kweken beestjes langskwam die 'dringend' een LMG nummer nodig had. En dan rest mij nog de enthousiaste supporters van het thuisfront, familie en vrienden te bedanken voor hun interesse en de nodige ontspanning die ze boden. En vooral Klaas (my sweet instigator), bedankt voor je onvoorwaardelijke steun! Anyway the water turns You're always on my mind You made all this Because you made all this Stefanie Gent, 31 maart 2004 Contents Dankwoord Contents List of non-standard abbreviations Preface 15 PART 1: Overview of the literature Chapter 1 : Bacterial diversity in Antarctic lakes and polar seas 23 1.1 Antarctic lakes and polar seas 1.2 The Antarctic ice-free areas 1.2.1 McMurdo Dry Valleys 1.2.2 Vestfold Hills 1.2.3 Larsemann Hills 1.3 Polar microbial habitats 1.3.1 Heterotrophic bacteria in Antarctic lakes 1.3.1.1 The plankton in Antarctic lakes 1.3.1.2 The benthic community in Antarctic lakes 1.3.1.3 The food web in Antarctic lakes 1.3.2 Oligotrophic bacteria in polar seas 1.3.2.1 Different polar sea habitats 1.3.2.2 The food web in polar seas 1.3.2.3 The starvation-survival response 1.3.2.4 Psychrophily and oligotrophy 1.4 Bacterial diversity in polar habitats 1.4.1 Diversity of heterotrophic bacteria in Antarctic lakes 1.4.2 Bacterial diversity in polar seas 1.4.2.1 Diversity in the sea-ice community 1.4.2.2 Diversity in the Southern Ocean 1.4.2.3 Diversity in the Arctic Ocean 1.5 Importance of polar microorganisms 1.5.1 Industrial applications 1.5.1.1 PUFA-production 1.5.1.2 Cold-adapted enzymes 1.5.1.3 Bioremediation 1.5.1.4 Biocatalysis under low water conditions 1.5.1.5 Anti freeze proteins 1.5.1.6 Pigments 1.5.2 Exobiology 1.6 Conceptual framework PART 2: Experimental work Chapter 2: Diversity of 746 heterotrophic bacteria isolated from microbial mats 59 from ten Antarctic lakes (Syst Appl Microbiol 25: 603-610, 2002) Chapter 3: New taxa from Antarctic lakes within the Bacteroidetes 75 3.1 Polyphasic taxonomy of FAA-clusters 1 to 15 3.2 Flavobacterium gelidilacus sp. nov., isolated from microbial mats in Antarctic lakes (Int J Syst Evol Microbiol 53: 1241-1245, 2003) 3.3 Flavobacterium degerlachei sp. nov., Flavobacterium frigoris sp. nov. and Flavobacterium micromati sp. nov., novel psychrophilic bacteria isolated from microbial mats in Antarctic lakes (Int J Syst Evol Microbiol 54: 85-92, 2004) 3.4 Flavobacterium fryxellicola sp. nov. and Flavobacterium psychrolimnae sp.nov., novel psychrophila bacteria isolated from microbial mats in Antarctic lakes (Int J Syst Evol Microbiol, in press) 3.5 Gillisia limnaea gen. nov., sp. nov., a new member of the family Flavobacteriaceae isolated from a microbial mat in Lake Fryxell, Antarctica (Int J Syst Evol Microbiol 54: 445-448, 2004) 3.6 Algoriphagus antarcticus sp. nov., a novel psychrophile from microbial mats in Antarctic lakes (Int J Syst Evol Microbiol, in press) Chapter 4: New taxa from Antarctic lakes within the a-Proteobacteria 141 4.1 Polyphasic taxonomy of FAA cluster 41 4.2 Loktanella fryxellensis gen. nov., sp. nov., Loktanella vestfoldensis sp. nov. and Loktanella salsilacus sp. nov., new members of the Rhodobacter group, isolated from microbial mats in Antarctic lakes (Int J Syst Evol Microbiol, in press) Chapter 5: New taxa from polar seas within the y- Proteobacteria 165 5.1 Polyphasic taxonomy of FAA-clusters E, F and related strains 5.2 Alteromonas stellipolaris sp. nov.: novel budding and prosthecate bacteria from Antarctic seas (Int J Syst Evol Microbiol, in press) 5.3 Glaciecola polaris sp. nov., novel budding and prosthecate bacteria from the Arctic Ocean, and emended description of the genus Glaciecola (Int J Syst Evol Microbiol, in press) PART 3: Concluding remarks Chapter 6: Conclusions and future perspectives 201 6.1 Polyphasic taxonomy of polar bacteria 6.1.1 Fatty acid analysis 6.1.2 Rep-PCR fingerprinting 6.1.3 16S rDNA sequence analysis 6.1.4 DNA-DNA hybridizations 6.2 Bacterial diversity in Antarctic lakes and polar seas 6.3 Geographical distribution of bacterial taxa 6.4 Ecological considerations 6.5 Biotechnological applications 6.6 Future perspectives Summary 219 Samenvatting 225 References 231 Annex I List of strains from Antarctic lakes 253 Annex II List of strains from polar seas 267 Annex III Project descriptions 273 Annex IV Biotechnological exploitation of heterotrophic bacteria and 277 filamentous fungi isolated from benthic mats of Antarctic lakes Annex V Origin of bacterial isolates from Arctic and Antarctic seas 297 Annex VI Dendrograms of Antarctic lake and polar sea 301 isolates Curriculum vitae 307 List of non-standard abbreviations AA Arachidonic Acid ACAM Australian Collection of Antarctic Microorganisms, University of Tasmania, Hobart, Australia AFLP Amplified Fragment Length Polymorphism ARDRA Amplified rDNA Restriction Analysis ATCC American Type Culture Collection, Manassas, VA, USA AWI Alfred-Wegener-Institut für Polar- und Meeresforschung, Bremerhaven, Germany BCCM/ LMG Belgian Co-ordinated Collections of Microorganisms/ Laboratorium voor Microbiologie Gent, University of Ghent, Belgium CFU Colony Forming Units CIP Collection de l'Institut Pasteur, Paris, France CM-cellulose Carboxymethylcellulose DCM Deep Chlorophyl Maximum DGGE Denaturing Gradient Gel Electrophoresis DHA Docosahexaenic Acid DOM Dissolved Organic Matter DSMZ Deutsche Sammlung von Mikroorganismen und Zellkulturen, Braunschweig, Germany EMBL European Molecular Biology Laboratory, European Bioinformatics Institute, Cambridge, UK EPA Eicosapentaeonic Acid ERIC Enterobacterial Repetitive Intergenic Consensus FAA Fatty Acid Analysis HPLC High-performance Liquid Chromatography HTL Higher Trophic Levels 1AM Institute of Applied Microbiology, University
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