No abstract

Arctic wintertime sea-ice cores, characterized by a temperature gradient of ؊2 to ؊20°C, were investigated to better understand constraints on bacterial abundance, activity, and diversity at subzero temperatures. With the fluorescent stains 4,6-diamidino-2-phenylindole 2HCl (DAPI) (for DNA) and 5-cyano-2,3-ditoyl tetrazolium chloride (CTC) (for O 2 -based respiration), the abundances of total, particle-associated (>3-m), freeliving, and actively respiring bacteria were determined for ice-core samples melted at their in situ temperatures (؊2 to ؊20°C) and at the corresponding salinities of their brine inclusions (38 to 209 ppt). Fluorescence in situ hybridization was applied to determine the proportions of Bacteria, Cytophaga-Flavobacteria-Bacteroides (CFB), and Archaea. Microtome-prepared ice sections also were examined microscopically under in situ conditions to evaluate bacterial abundance (by DAPI staining) and particle associations within the brine-inclusion network of the ice. For both melted and intact ice sections, more than 50% of cells were found to be associated with particles or surfaces (sediment grains, detritus, and ice-crystal boundaries). CTC-active bacteria (0.5 to 4% of the total) and cells detectable by rRNA probes (18 to 86% of the total) were found in all ice samples, including the coldest (؊20°C), where virtually all active cells were particle associated. The percentage of active bacteria associated with particles increased with decreasing temperature, as did the percentages of CFB (16 to 82% of Bacteria) and Archaea (0.0 to 3.4% of total cells). These results, combined with correlation analyses between bacterial variables and measures of particulate matter in the ice as well as the increase in CFB at lower temperatures, confirm the importance of particle or surface association to bacterial activity at subzero temperatures. Measuring activity down to ؊20°C adds to the concept that liquid inclusions in frozen environments provide an adequate habitat for active microbial populations on Earth and possibly elsewhere.The constraints on and sustainability of life in frozen environments are of considerable importance in a number of contexts, from polar microbial ecology and astrobiology to cryopreservation and other industrial applications (42). For example, a number of subzero environments, such as Antarctic and Arctic lakes (23,25,38), snow (3), glacial ice (46), and permafrost soils (41), have been investigated as Earth analogs for potential extraterrestrial habitats also at subzero temperatures. To date, fundamental questions underlying the behavior of bacteria in any frozen environment have not been adequately addressed: how do bacteria manage to persist and possibly remain active? At the lowest temperatures observed on Earth, what environmental factors enable and control bacterial survival and even sustained activity?This study focused on Arctic wintertime sea ice, the coldest marine habitat on Earth (temperature range of Ϫ2 to Ϫ35°C) (31) and an important component of polar climate and ecos...

Heterotrophic bacteria in sea ice play a key role in carbon cycling, but little is known about the predominant players at the phylogenetic level. In a study of both algal bands and clear ice habitats within summertime Arctic pack ice from the Chukchi Sea, we determined the abundance of total bacteria and actively respiring cells in melted ice samples using epifluorescence microscopy and the stains 4', 6'-diamidino-2-phenylindole 2HCl (DAPI) and 5-cyano-2,3-ditolyl tetrazolium chloride (CTC), respectively. Organic-rich and -poor culturing media were used to determine culturable members by plating (at 0 degrees C and 5 degrees C) and most-probable-number (MPN) analyses (at -1 degrees C). Total bacterial counts ranged from 5.44 x 10(4) ml(-1) in clear ice to 2.41 x 10(6) ml(-1) in algal-band ice samples, with 2-27% metabolically active by CTC stain. Plating and MPN results revealed a high degree of culturability in both types of media, but greater success in oligotrophic media (to 62% of total abundance) and from clear ice samples. The bacterial enumeration anomaly, commonly held to mean

Microbial processes that mineralize organic carbon and enhance solute production at the bed of polar ice sheets could be of a magnitude sufficient to affect global elemental cycles. To investigate the biogeochemistry of a polar subglacial microbial ecosystem, we analyzed water discharged during the summer of 2012 and 2013 from Russell Glacier, a land-terminating outlet glacier at the western margin of the Greenland Ice Sheet. The molecular data implied that the most abundant and active component of the subglacial microbial community at these marginal locations were bacteria within the order Methylococcales (59-100% of reverse transcribed (RT)-rRNA sequences). mRNA transcripts of the particulate methane monooxygenase (pmoA) from these taxa were also detected, confirming that methanotrophic bacteria were functional members of this subglacial ecosystem. Dissolved methane ranged between 2.7 and 83 lM in the subglacial waters analyzed, and the concentration was inversely correlated with dissolved oxygen while positively correlated with electrical conductivity. Subglacial microbial methane production was supported by d 13 C-CH 4 values between À 64% and À 62% together with the recovery of RT-rRNA sequences that classified within the Methanosarcinales and Methanomicrobiales. Under aerobic conditions, 498% of the methane in the subglacial water was consumed over B30 days incubation at B4 1C and rates of methane oxidation were estimated at 0.32 lM per day. Our results support the occurrence of active methane cycling beneath this region of the Greenland Ice Sheet, where microbial communities poised in oxygenated subglacial drainage channels could serve as significant methane sinks.