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Investigations of Fungal Diversity in Wooden Structures and Soils at Historic Sites on the Antarctic Peninsula1

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Investigations of Fungal Diversity in Wooden Structures and Soils at Historic Sites on the Antarctic Peninsula1 46 Investigations of fungal diversity in wooden structures and soils at historic sites on the Antarctic Peninsula1 Brett E. Arenz and Robert A. Blanchette Abstract: Investigations of microbial diversity in Antarctic are important to begin to understand ecosystem functioning and decomposition processes. This study documents fungi at 9 historic sites on the Antarctic Peninsula collected from wooden structures, other organic materials, and soils during a joint National Science Foundation and British Antarctic Sur- vey expedition in 2007. Many of these sites had wooden structures built by the British during the World War II Operation Tabarin, but others visited included the American ‘‘East Base’’ on Stonington Island and the Swedish hut on Snow Hill Is- land. Fungi were cultured on several different media and pure cultures were obtained and identified by DNA sequencing of the internal transcribed spacer region. Cadophora species previously found to attack historic wooden structures on Ross Island, Antarctica, were found at all but 1 location sampled in the Peninsula region. Fungi causing decay in the historic wooden structures and artifacts and those causing mold problems inside the structures are of great concern, and conserva- tion efforts are urgently needed to help preserve these important polar heritage structures. The results presented also ex- pand our knowledge on the identity of fungi present throughout the Antarctic Peninsula region and provide insights into the organisms responsible for decomposition and nutrient recycling. Key words: Antarctica, fungi, wood decay, biodeterioration, historic conservation. Re´sume´ : Les recherches portant sur la diversite´ microbienne de l’Antarctique sont importantes afin de commencer a` com- prendre le fonctionnement de l’e´cosyste`me et les processus de de´composition. Cette e´tude documente les populations de champignons de 9 sites historiques de la pe´ninsule Antarctique recueillis sur des structures de bois, sur d’autres mate´riaux organiques et dans le sol, au cours d’une expe´dition de la « National Science Foundation » et du « British Antarctic Sur- vey en 2007 ». Des structures de bois construites par les Britanniques lors de l’Ope´ration Tabarin pendant la deuxie`me guerre mondiale e´taient pre´sentes sur plusieurs sites, les autres sites visite´s comprenant la base ame´ricaine « East Base » de l’Iˆle de Stonington et les cabanes construites pas les Sue´dois sur l’Iˆle Snow Hill. Les champignons ont e´te´ cultive´s sur diffe´rents milieux, des cultures purifie´es ont e´te´ obtenues et identifie´es par se´quenc¸age d’ADN de la re´gion de l’espaceur interne transcrit (ITS). Des espe`ces appartenant a` Cadophora, tenues pre´ce´demment responsables de la de´te´rioration des structures de bois de l’Iˆle Ross en Antarctique, ont e´te´ trouve´es dans tous les sites e´chantillonne´sdelape´ninsule sauf un. Les champignons causant la pourriture des structures de bois historiques et des arte´facts et ceux qui causent des proble`mes de moisissure a` l’inte´rieur de ces structures sont pre´occupants et des efforts sont instamment requis pour aider a` pre´server cet he´ritage polaire d’importance. Les re´sultats pre´sente´se´largissent aussi nos connaissances sur l’identite´ des champignons pre´sents a` travers la re´gion de la pe´ninsule Antarctique et donnent un aperc¸u des organismes responsables de la de´compo- sition et du recyclage des nutriments. Mots-cle´s:Antarctique, pourriture du bois, de´te´rioration biologique, conservation historique. [Traduit par la Re´daction] Introduction middle part of the last century. The British in particular es- The Antarctic Peninsula has been the target of significant tablished a large presence in this region for not only scien- exploration over the past decades and many nations con- tific and geographic exploration, but also to reinforce their structed huts or relatively large bases there in the early and claims of territorial sovereignty. The buildings and materials left behind from these expeditions still exist and many have been classified as historic monuments by the Antarctic treaty Received 30 September 2008. Accepted 1 October 2008. system, but unfortunately deterioration and decay are caus- Published on the NRC Research Press Web site at cjm.nrc.ca on ing significant problems for their long term preservation. 30 January 2009. Past research on historic wooden structures in other re- B.E. Arenz2 and R.A. Blanchette.3 Department of Plant gions of Antarctica has found that they are affected by both Pathology, University of Minnesota, 495 Borlaug Hall, abiotic and biotic forms of degradation. Abiotic forms in- 1991 Upper Buford Circle, St. Paul, Minnesota 55108, USA. clude wind erosion, oxidation of metals (Otineo-Alego et al. 1This article is one of a selection of papers in the Special Issue 2000), and salt defibration of wood fibers (Blanchette et al. on Polar and Alpine Microbiology. 2002). Biotic deterioration is largely due to fungi that can 2Corresponding author (e-mail: [email protected]). cause both disfiguring mold growth on the surface of arti- 3Corresponding author (e-mail: [email protected]). facts and a soft-rot type of wood decay (Blanchette et al. Can. J. Microbiol. 55: 46–56 (2009) doi:10.1139/W08-120 Published by NRC Research Press Arenz and Blanchette 47 2004a). The fungi found to be responsible for decaying 0.001% cyclohexamide and 0.005% chloramphenicol added wood were several species of Cadophora including Cado- after autoclaving). Isolates were cultured at 8 and 20 8C. phora malorum, Cadophora luteo-olivaceae, and Cadophora Pure cultures were obtained after subsampling. fastigiata, as well as several previously undescribed Cado- DNA was extracted by a phenol–chloroform procedure. phora species. Although this past research took place in the This procedure was modified from Zhong and Steffenson Ross Sea region of Antarctica where it is relatively very (2001) in that the mycelium and the spores were collected cold and dry, environmental monitoring inside the huts has by scraping the surface of the agar plate containing the pure revealed that during the Austral summers conditions that are isolate, rather than filtering a liquid culture. Also, 700 mLof conducive to fungal growth (temperatures above 0 8C and the lysis buffer was used instead of 7 mL. Ribosomal DNA relative humidity above 80%) occur frequently (Held et al. was amplified via PCR with the primers ITS1 and ITS4 2005). A survey of fungal diversity at these Ross Sea his- (Gardes and Bruns 1993). PCR amplification was performed toric sites has found that the most abundantly isolated spe- using an Amplitaq Gold PCR Master-mix and a 1 mL tem- cies affecting the historic wood and artifacts were also plate DNA and following the manufacturer’s instructions isolated from soils around the structures and from more dis- (Applied Biosystems, Foster City, California). A MJ Re- tant and remote areas (Arenz et al. 2006). A number of cos- search PTC Minicycler (Watertown, Maryland) was used mopolitan species were also identified that had not been with the following profile: 94 8C for 5 min, 35 cycles of previously reported from Antarctica although these species 94 8C for 1 min, 50 8C for 1 min, and 72 8C for 1 min, fol- were found in relatively low frequency. lowed by a final extension step of 72 8C for 5 min. PCR Historic sites on the Antarctic Peninsula have not been the products of appropriate size (500–600 bp) were verified by subject of mycological investigations and little is known electrophoresis of the amplicons on a 1% agarose gel with a about the microorganisms present in these areas. If these po- SYBR green 1 (Molecular Probes, Eugene, Oregon) prestain lar heritage sites are to be adequately preserved, it is impor- and transilluminating with a Dark Reader DR45 (Clare tant for conservators to know what factors are contributing Chemical Research, Denver, Colorado). Sequencing was to the deterioration and decay of the wood and artifacts. performed for both primers using the ABI PRISM Dye Ter- Since the Antarctic Peninsula has a relatively warmer and minator Cycle Sequencing Ready reaction kit (Applied Bio- more humid climate than the Ross Sea region of Antarctica, systems) and an ABI Prism 377 automated DNA sequencer. degradation processes likely take place faster and a greater DNA sequence data were analyzed by Chromas software abundance of diverse fungi could be expected. This study (Technelysium Ltd., Helensvale, Australia) and assembled was done to determine fungal diversity present at 9 historic into a consensus sequence. The sequences were compared sites on the Antarctica Peninsula in woods, organic materi- with others in GenBank using BLASTn (Altschul et al. als, and soils, to make comparisons with other previous 1990) to find the best match. studies completed on the opposite side of Antarctica in the Selected wood samples were cut with a cryostat freezing Ross Sea region and to learn more about these organisms microtome and observed with a Hitachi S3500 scanning and their role in polar ecosystems. electron microscope (SEM). Materials and methods Results and discussion Nine sites on the Antarctic Peninsula were visited in Jan- A total of 186 samples were analyzed from all locations uary 2007 from the British Antarctic Survey (BAS) ship (Table 1), a large portion of which came from East Base be- HMS Endurance (Table 1). The sites consisted of 7 historic cause this was a very large historic site and the main focus British bases: Base A (Port Lockroy Station), Base D on of the deterioration assessments during the event. From Hope Bay, Base E on Stonington Island, Base F (Wordie these samples, 81 taxa were identified from 295 fungal iso- House), Base V (View Point Station), and Base W on De- lations (Table 2). Ascomycetes (58) were dominant among taille Island; and Base Y on Horseshoe Island (Fig.
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