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227504791.Pdf View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Scholar Commons | University of South Florida Research University of South Florida Scholar Commons School of Geosciences Faculty and Staff Publications School of Geosciences 2014 Searching for Cold-Adapted Microorganisms in the Underground Glacier of Scarisoara Ice Cave, Romania Alexandra Maria Hillebrand-Voiculescu Emil Racovita Institute of Speleology, [email protected] Corina Itcus Institute of Biology Bucharest Ioan Ardelean Institute of Biology Bucharest Denisa Pascu Institute of Biology Bucharest Aurel Persoiu Stefan cel Mare University See next page for additional authors Follow this and additional works at: https://scholarcommons.usf.edu/geo_facpub Part of the Earth Sciences Commons Scholar Commons Citation Hillebrand-Voiculescu, Alexandra Maria; Itcus, Corina; Ardelean, Ioan; Pascu, Denisa; Persoiu, Aurel; Rusu, Andreea; Brad, Traian; Popa, Elena; Onac, Bogdan P.; and Purcarea, Christina, "Searching for Cold-Adapted Microorganisms in the Underground Glacier of Scarisoara Ice Cave, Romania" (2014). School of Geosciences Faculty and Staff Publications. 1893. https://scholarcommons.usf.edu/geo_facpub/1893 This Article is brought to you for free and open access by the School of Geosciences at Scholar Commons. It has been accepted for inclusion in School of Geosciences Faculty and Staff Publications by an authorized administrator of Scholar Commons. For more information, please contact [email protected]. Authors Alexandra Maria Hillebrand-Voiculescu, Corina Itcus, Ioan Ardelean, Denisa Pascu, Aurel Persoiu, Andreea Rusu, Traian Brad, Elena Popa, Bogdan P. Onac, and Christina Purcarea This article is available at Scholar Commons: https://scholarcommons.usf.edu/geo_facpub/1893 COBISS: 1.01 Searching for cold-adapted microorganisms IN the underground glacier of Scarisoara Ice Cave, Romania Iskanje na mraz prilagojenih mikroorganizmov V podzemnem ledeniku Ledene jami Scarisoara (Romunija) Alexandra HILLEBRAND-VOICULESCU1#, Corina ITCUS2#, Ioan ARDELEAN2, Denisa PASCU2, Aurel PERŞOIU3, Andreea RUSU2, Traian BRAD4, Elena POPA2, Bogdan P. ONAC5,4 & Cristina PURCAREA2* Abstract UDC 551.444.6:579.26(498) Izvleček UDK 551.444.6:579.26(498) Alexandra Hillebrand, Corina Itcus, Ioan Ardelean, Denisa Alexandra Hillebrand, Corina Itcus, Ioan Ardelean, Denisa Pascu, Aurel Perşoiu, Andreea Rusu, Traian Brad, Elena Pascu, Aurel Perşoiu, Andreea Rusu, Traian Brad, Elena Popa, Bogdan P. Onac & Cristina Purcarea: Searching for Popa, Bogdan P. Onac & Cristina Purcarea: Iskanje na mraz cold-adapted microorganisms in the underground glacier of prilagojenih mikroorganizmov v podzemnem ledeniku Ledene Scarisoara Ice Cave, Romania jami Scarisoara (Romunija) Scarisoara Ice Cave (Romania) hosts one of world’s largest and old- študija obravnava pojavnost in časovno porazdelitev mikroorga- est underground glacier. While no studies were carried out on the nizmov v jamskem ledu v Ledeni jami Scarisoara (Romunija), v existence of microorganisms in this cave’s ice block, our interest is kateri je eden največjih in najstarejših jamskih ledenikov. Pojavnost to investigate the presence of microorganisms and their chronolog- mikroorganizmov nas zanima predvsem v povezavi s preteklimi ical distribution in the cave’s subterranean ice in relationship with klimatskimi spremembami. Vzorčili smo v različnih plasteh ledu, past climatic changes. Samples were collected from ice layers of starih od 900 let do recentnih in v njih določevali združbe mikro- different age (from present to ~900 cal. yrs. BP), and the diversity organizmov z klasičnim gojenjem in z molekularnimi metodami. of embedded microbial communities was assessed by classical cul- Mikroorganizme iz lednih sedimentov smo v temi in na svetlobi tivation and molecular techniques. The microorganisms from ice- gojili na temperaturah 5 °C in 15 °C. Epifluorescentna mikrosko- sediments were cultivated at 4 °C and 15 °C, in the presence and pija je pokazala prisotnost avtotrofnih prokariontov in evkariontov absence of light. Epifluorescence microscopy analysis indicates the v vzorcih ledu in vode, ki so bili izpostavljeni sončnemu obseva- presence of autotrophic prokaryotes and eukaryotes in sunlight- nju. Iz ledu smo izolirali celotno DNK ter bakterijske in evkarion- exposed ice and water samples. Total DNA was isolated from each t ske gene male ribosomske podenote (SSU-rRNA) namnožili s ice sample and the bacterial and eukaryotic SSU-rRNA genes were poli merazno verižno reakcijo (PCR). Kemično sestavo in orga- amplified by PCR. The chemical composition and organic content n­­ski delež globoko pokopanih (več kot 10 m globoko v ledu) in of both deeply buried (>10 m inside the ice block) and surface (su- površinskih (vodni bazen na površini ledu) habitatov smo pove- pra-glacial pond water) habitats were analyzed in relation to their zali z njihovo starostjo in organsko sestavo. študija prva poroča o age and organic composition. This study is the first to report on the prokariontskih in evkariontskih mikroorganizmih v podzemnem presence of both prokaryotic and eukaryotic microorganisms in ledu jame Scarisoara, ki uspevajo tako v organsko bogatih plasteh the subterranean ice block of Scarisoara Ice Cave, thriving in both ledu kot v prosojnem ledu. V ledu, izpostavljenem sončni svetlobi, organic-rich ice and clear ice layers. Phototrophic prokaryotes and smo odkrili prisotnost fototrofnih prokariontov in evkariontov. eukaryotes were identified in sun-exposed recent ice. The compo- Sestava na mraz prilagojenih mikroorganizmov v ledu se spremi- 1 Emil Racovita Institute of Speleology, Bucharest, Romania, e-mail: [email protected] 2 Institute of Biology Bucharest, Bucharest, Romania, e-mail: [email protected], [email protected], [email protected], [email protected], [email protected], e-mail: [email protected] 3 Stefan cel Mare University, Suceava, Romania, e-mail: [email protected] 4 Emil Racovita Institute of Speleology, Cluj-Napoca, Romania, e-mail: [email protected] 5 School of Geosciences, University of South Florida, USA, e-mail: [email protected] # authors with equal contribution * Corresponding author: Dr. Cristina PURCAREA, Department of Microbiology, Institute of Biology Bucharest, Romanian Academy, 296 Splaiul Independentei, 060031 Bucharest, Romania, e-mail: [email protected] Received/Prejeto: 20.05.2013 ACTA CARSOLOGICA 43/2–3, 319–329, POSTOJNA 2014 A. HILLEBRAND-VOICULESCU, C. ITCUS, I. ARDELEAN, D. PASCU, A. PERŞOIU, A. RUSU, T. BRAD4, E. POPA, B. P. ONAC & C.PURCAREA sition of cold-adapted ice embedded microbiota varied with the nja s starostjo in organsko sestavo habitata, na kar kaže tudi razlika habitat age and organic content, as resulting from dissimilarities in rastnih krivulj pri dveh različnih temperaturah. Prisotnost bakterij growth curve profiles at two different temperatures. The presence in evkariontov v vseh analiziranih vzorcih smo potrdili tudi z PCR of bacteria and eukaryotes in all the analyzed samples was asserted namnožitvijo genskih fragmentov SSU-rRNA. Opisane ugotovitve by PCR amplification of SSU-rRNA gene fragments.These findings bodo uporabne za rekonstrukcijo sprememb pestrosti mikroorga- can be further used to reconstruct changes in the microbial diver- nizmov v odvisnosti od okoljskih sprememb, zabeleženih v jam- sity over the past approximately 5000 years, in correlation with cli- skem ledu v zadnjih 5000 letih. matic and environmental changes recorded by the ice block. Ključne besede: podzemni ledenik, klimatski arhiv, mikroorga- Keywords: underground glacier, climate archives, microorga- nizmi, psihrofili, Ledena jama Scarisoara. nisms, psychrophiles, Scarisoara Ice Cave. INTRODUCTION The microbial diversity of cold environments was inves- 2006; Jungblut et al. 2010; Namsaraev et al. 2010; Uetake tigated from a large variety of exposed ice habitats (Pr- et al. 2010). Their presence and role in perennial ice de- iscu et al. 2007) such as polar ice sheets (Jungblut et al. posits from caves could be of particular interest for pal- 2010; Rehakova et al. 2010; Varin et al. 2010), alpine gla- aeoclimatic reconstruction. ciers (Tscherko et al. 2003) and frozen lakes (Felip et al. Recently, the discovery of liquid water and ice on 1995, Dieser et al. 2010), and Antarctic permanent lake Mars (Levin & Weatherwax 2003; Kerr 2010), as well as ice (Priscu et al. 1998, Murray et al. 2012). Compara- the theoretical demonstration of the possible existence tive geochemical and microbiological studies of ice cores of ice caves on Mars (Boston 2004; Williams et al. 2010), drilled in polar and high mountain areas (Miteva et al. increased the interest for such peculiar terrestrial envi- 2009) allowed for the identification of climate biomark- ronments and their biodiversity that mimic extrater- ers. In ice layers of different ages from Antarctic (Abyzov restrial conditions, thus giving an insight on exobiology et al. 1998; Xiang et al. 2005) and Alpine (Zhang et al. aspects (Jakosky et al. 2003; Rampelotto 2010), but also 2006) glaciers, the variation in microbial composition into possible earlier environments on Earth. and abundance was correlated with climatic changes. Scarisoara Ice Cave (Romania), containing one of However, very little is known to date about microor- the oldest and largest subterranean ice block (Perşoiu ganisms communities living in ice deposits from caves, 2011), presents an easily accessible and radiocarbon
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