Nitrogen fixation on Arctic glaciers, Svalbard

Jon Telling,1 Alexandre M. Anesio,1 Martyn Tranter,1 Tristram Irvine‐Fynn,2

Andy Hodson,3 Catriona Butler,1 and Jemma Wadham1

Received 15 December 2010; revised 14 June 2011; accepted 6 July 2011; published 28 September 2011.

[1] Glacier surfaces contain a wide diversity of microorganisms and can host a range ofmicrobial activities. However, microbial nutrient cycling on glaciers is poorly understood.This study is the first to document nitrogen fixation (nitrogenase activity) on glaciers anddemonstrate its importance in supporting microbial growth. Rates of nitrogen fixation(nitrogenase activity) in cryoconite holes on three valley glaciers in Svalbard ranged from<2.0 to 99.9 mmol ethylene m−2 d−1 with rates inversely correlated to concentrationsof available inorganic nitrogen. Annual inputs of nitrogen by nitrogen fixation on a glaciercatchment scale are more than 2 orders of magnitude lower than the combined nitrogeninputs from snowmelt and rain. However, nitrogen fixation can be important forsupporting microbial growth on the glaciers during the middle to late melt season after thesnowline has retreated upslope.

Citation: Telling, J., A. M. Anesio, M. Tranter, T. Irvine‐Fynn, A. Hodson, C. Butler, and J. Wadham (2011), Nitrogen fixationon Arctic glaciers, Svalbard, J. Geophys. Res., 116, G03039, doi:10.1029/2010JG001632.

1. Introduction

[2] The surface of glaciers and ice sheets can be habitatsfor microbial life when liquid water is present [Hodson et al.,2008]. Ice covers ∼10% of the Earth’s surface in the currentinterglacial period, rising to 30% at the Last Glacial Maxi-mum [Paterson, 1994] and covered up to 100% of the globalsurface during glaciations in the Neoproterozoic [Hoffmanet al., 1998]. Research over the last decade has measuredsignificant pockets of microbial activity on glaciers and icesheets often associated with surface sediment (cryoconite)that melts into the ice surface to form cryoconite holes[Säwström et al., 2002; Anesio et al., 2009]. To date, therehave been few studies of how nutrient dynamics impact onmicrobial productivity and how nutrient dynamics changeover the melt season.[3] The study of nutrient dynamics on glaciers surfaces

has three broad objectives. The first is to determine howmicroorganisms survive and grow in the cold and nutrient‐poor supraglacial environment [Säwström et al., 2007;Stibal et al., 2008]. The second is to determine if nutrientsfrom glacier and ice sheet surfaces are important for sus-taining microbial communities in hydrologically connectedsubglacial and downstream environments [Hodson et al.,2008; Hood et al., 2009]. The third is to establish whetheror not anthropogenic pollution impacts on glacier and icesheet ecology. For example, anthropogenic nitrogen depo-sition on some Arctic glaciers has substantially increased

since preindustrial times and its effect on microbial com-munities is largely unknown [Hodson et al., 2009].[4] Nitrogen is often the most important nutrient for bal-

anced microbial growth [Redfield et al., 1963], yet ourunderstanding of nitrogen cycling on glacier surfaces is verylimited. The only study to report direct evidence of micro-bial nitrogen cycling on glacier or ice sheet surfaces used15N labeling experiments to demonstrate the rapid turnoverof ammonia in ice lidded cryoconite holes in the McMurdoDry Valleys of Antarctica [Tranter et al., 2004]. There is agreat deal of indirect evidence for supraglacial microbialnitrogen cycling, however, including (1) the suggestionof significant ammonia retention on Arctic valley glaciers[Hodson et al., 2005; Wynn et al., 2007], (2) the presence ofan additional source of nitrate on Arctic and Maritime Ant-arctic glaciers perhaps due to either nitrification or nitrogenfixation and subsequent mineralization [Hodson et al., 2005;Hodson, 2006], and (3) the demonstration of the potential fornitrogen fixation on glaciers and ice sheets by the identifi-cation of known nitrogen fixing microorganisms [Christneret al., 2003; Stibal et al., 2006; Edwards et al., 2011].[5] This study tests the hypothesis that nitrogen fixation

is important for supporting microbial communities onHigh Arctic valley glaciers. This allows us to evaluate howanthropogenic nitrogen deposition may have perturbed therequirement for in situ microbial nitrogen fixation.

2. Methods

2.1. Study Locations

[6] Nitrogen fixation assays and nutrient analysis werecarried out on cryoconite holes on three valley glaciers(Midtre Lovénbreen (ML), Vestre Brøggerbreen (VB),and Austre Brøggerbreen (AB)) situated in the NW of theSvalbard archipelago close to the scientific base atNy‐Ålesundat 79°N, 12°E (Figure 1). Themass balance of all three glaciers

1Bristol Glaciology Centre, School of Geographical Sciences,University of Bristol, Bristol, UK.

2Centre for Glaciology, Institute of Geography and Earth Science,Aberystwyth University, Aberystwyth, UK.

3Department of Geography, University of Sheffield, Sheffield, UK.

Copyright 2011 by the American Geophysical Union.0148‐0227/11/2010JG001632

JOURNAL OF GEOPHYSICAL RESEARCH, VOL. 116, G03039, doi:10.1029/2010JG001632, 2011

G03039 1 of 8

has been negative since the 1930s resulting in retreat and icethinning [Hagen et al., 2003; Nuth et al., 2007]. A total of40 cryoconite holes were assayed for in situ nitrogen fixationand sampled for water and cryoconite chemistry between8 July and 25 August 2009. Measurements encompassed boththe early melt season when cryoconite holes were still coveredby slush up until near the end of the main melt season whenthin ice lids on top of cryoconite holes were beginning topersist during the day.[7] Water samples for nutrient analysis from each cryo-

conite hole were taken with a 50 mL syringe and filteredthrough 0.45 mm Whatman™ cellulose nitrate filters into30 mL Nalgene™ HDPE containers. Cryoconite for nutrientanalysis from each hole was placed into sterile 15 mLcentrifuge tubes using plastic scoops. Samples were trans-ported chilled and in the dark to the laboratory within 8 h ofsampling, frozen at −20°C and transported frozen back toBristol for later chemical analysis.

2.2. Water and Cryoconite Chemistry

[8] Dissolved NH4+, NO3

− and NO2− (whose sum is termed

DIN) in the water phase of cryoconite holes were analyzedon a Bran and Luebbe Autoanalyzer 3. Detection limits were10 mg N L−1, 27.5 mg N L−1 and 3.5 mg N L−1 for NH4

+,

NO3−, and NO2

−, respectively (defined as 3s of six deionizedwater blanks). Precisions for analyses were 0.6%, 3.6%, and1.9%, respectively.[9] Exchangeable NH4

+, NO3− and NO2

− in cryoconitedebris (whose sum is termed TIN) were extracted using the1M KCL method of Maynard et al. [2007]. Extracts wereanalyzed on a Bran and Luebbe Autoanalyzer 3 as describedabove. The detection limits were 0.65, 0.55 and 0.03 mg Ng−1 dry sediment for NH4+, NO3

−, and NO2−, respectively,

with no nitrogen detected in KCl only controls.[10] Duplicate dried cryoconite samples were analyzed for

total carbon (TC) and total nitrogen (TN) on a EurovectorEA3000 Elemental Analyzer. Inorganic carbon (IC) wasdetermined using a Coulomat 720 analyzer, and total organiccarbon (TOC) was defined as (TC‐IC). Particulate organicnitrogen (PON) was defined as (TN‐TIN). Duplicate cryo-conite analyses were within 20% for both TOC and TN.The detection limits for TOC and TN were 100 mg C g−1 and100 mg N g−1, respectively.

2.3. Nitrogen Fixation Assays

2.3.1. In Situ Assays[11] Rates of nitrogen fixation (nitrogenase activity) were

measured using the acetylene assay [Stewart et al., 1967]

Figure 1. Location of field sites on Midtre Lovénbreen (ML), Vestre Brøggerbreen (VB), and AustreBrøggerbreen, situated in the NW of the Svalbard archipelago, close to the scientific base at Ny‐Ålesundat 79°N, 12°E. Location of early season slush covered cryoconite holes and main/late melt season cryo-conite holes used for in situ nitrogen fixation and nutrient analysis are shown. Note that two cryoconiteholes were sampled at each of the marked slush samples on VB and ML. The map is projected in uni-versal transverse Mercator (UTM) WGS84 with contours displayed at 100 m intervals.

TELLING ET AL.: NITROGEN FIXATION ON ARCTIC GLACIERS G03039G03039

2 of 8

using a field sampling technique based on the work of Stutzand Bliss [1973]. At each site (Figure 1) cryoconite wasadded to two 30 mL serum bottles (Wheaton) to give similarcryoconite thicknesses to that found in situ. Cryoconitewater (15 mL) was added to each of the bottles and thebottles closed with thick butyl stoppers (Bellco) andcrimped shut. An aliquot (1.5 mL) of 100% acetylene gas,generated in the field by adding water to technical gradecalcium carbide (Sigma), was then added via a syringe andneedle to one of the two bottles. The bottle of each pair withno acetylene addition was used to check for any naturalethylene (C2H4) production from the cryoconite or water.Both bottles were then shaken vigorously for 30 s andplaced at the bottom of the cryoconite holes for 24 h ± 3 h.In addition a total of 12 serum bottles (incorporating earlyand middle/late season sites on all three glaciers) were filledwith 15 mL of 0.2 mm filtered deionized water and incu-bated and analyzed as above to act as blank controls.[12] After incubation the bottles were removed from the

cryoconite holes and overpressurized by injecting 10 mL ofair into the bottles via a syringe and needle. Gas sampleswere then taken by attaching the incubation bottles via two‐way needles to preevacuated 5 ml serum bottles fitted withthick butyl stoppers (Bellco). After sampling, bottles werestored refrigerated for up to three months prior to analysison a Varian 3800 gas chromatograph (GC). Ethylene wasseparated from acetylene using a 2.0 m × 1/8″ Hayesep‐Tcolumn (80–100 mesh) and an isothermal temperature of85°C. Daily standards of 100 ppm ethylene (Sigma) gaveprecisions <8%. Standards of ethylene prepared in 5 mLserum vials and analyzed after three months storage alsogave values within 8% of 100 ppm. Ethylene results in ppmwere converted to mmol C2H4 m−2 d−1 by first accountingfor ethylene dissolution in the water using the equations ofBreitbarth et al. [2004] and then normalizing for the surfacearea of cryoconite in the bottles. The detection limit was2.0 mmol C2H4 m−2 d−1. Rates were converted into units

of mg N m−2 d−1 assuming a 3:1 molar ratio betweenethylene produced to nitrogen molecule fixed [Stewart et al.,1967] to allow comparison with previous estimates ofnitrogen fluxes on the glaciers.2.3.2. Laboratory Assays[13] Long incubations with acetylene can result in non

linear rates of ethylene production with time [David andFay, 1977]. Therefore laboratory experiments were carriedout to check the linearity of ethylene production over the24 h incubations used in this study. Triplicate 1 g cryoconitesamples from AB and VB with measurable rates of in situnitrogen fixation were used in laboratory experiments.Bottles with cryoconite and 15 mL 0.2 mm filtered deionizedwater were set up as described for the in situ incubationswith the exception that the bottles were left for 48 h in an icebath under fluorescent lights prior to starting the experiments inorder to counteract any inhibitory effects caused by freezingand thawing of the cryoconite [DuBois and Kapustka, 1983].Photosynthetically Active Radiation (PAR) was measurednext to the bottles using a Li‐Cor Li‐190 sensor and aver-aged 110 mmol m−2 s−1, similar to the mean of 121 mmolm−2 s−1 PAR measured at the bottom of cryoconite holes onthe three glaciers of this study (J. Telling, unpublished data,2009). After 48 h acetylene was added as described above.At 3 h intervals, subsamples (1 mL) of headspace gas weretaken with a gas tight syringe and analyzed for ethylene onthe GC as described above.

3. Results

3.1. Cryoconite Chemistry

[14] DIN was detected in seven of the 40 water samples,with six of the seven samples within the slush zone (Figure 2a).DissolvedNO3

− concentrations ranged from<27.5mgNL−1 to35.5 mg N L−1 and dissolved NH4

+ ranged from <10 mg N L−1

to 95.2 mg N L−1, with a maximum DIN of 132.5 mg N L−1

(Figure 2a).NO2−was not detected in any sample (<3.5mgNL−1).

[15] TIN was detected in 29 of the 40 cryoconite debrissamples (Figure 2b), ranging from <0.65 to 18.2 mg N g−1.In all 29 samples TIN was dominated by NH4

+ rather thanNO3

− with 94 ± 12% (1s) of TIN in the form of NH4+. The

four highest concentrations of TIN were from sampleswithin the slush zone (Figure 2b). Concentrations of TN incryoconite (2.2 to 3.8 mg N g−1) were substantially greaterthan TIN indicating that 99.9 ± 1.4% of TN was in the formof PON (Figure 3a). There was no significant correlationbetween PON and TIN if all samples were included (r =0.255, p = 0.117, n = 40, two‐tailed t test) (Figure 3a).However, there was a significant correlation between PONand TIN if slush zone samples were excluded (r = 0.529,p < 0.01, n = 30, Spearman’s correlation, two‐tailed t test)with no significant correlation between PON and TIN ofslush only samples (r = 0.018, p = 0.960, n = 10, Spearman’scorrelation, two‐tailed t test). The dominance of PON overTIN is consistent with the strong correlation between TOCand TN (r = 0.939, p < 0.001, n = 40, Spearman’s correlation,two‐tailed t test; Figure 3b).

3.2. Nitrogen Fixation

3.2.1. In Situ Nitrogen Fixation[16] Nitrogen fixation rates in bottles amended with cryo-

conite ranged from <2.0 mmol C2H4 m−2 d−1 to 99.9 mmol

Figure 2. In situ measurements of nitrogen fixation (N2

fixation) versus inorganic nitrogen in cryoconite holes.(a) Ethylene (C2H4) production versus dissolved inorganicnitrogen (DIN) in the water phase of cryoconite holes. Con-centrations of TIN and DIN below detection are plotted aszero. (b) Ethylene (C2H4) production versus exchangeableinorganic nitrogen in cryoconite (TIN). Open circles areearly melt season (slush zone) samples; solid circles aremiddle/late melt season samples. Dotted lines show thedetection limit (<2.0 mmol ethylene (C2H4) m

−2 d−1).

TELLING ET AL.: NITROGEN FIXATION ON ARCTIC GLACIERS G03039G03039

3 of 8

C2H4 m−2 d−1 (Figure 2). Nitrogen fixation rates wereundetectable in all ten cryoconite holes analyzed during theearly melt season on the three glaciers (8–17 July). Nitrogenfixation was detected however in 16 of the 30 cryoconiteholes analyzed after the snowline had retreated upslope inthe middle to late melt season (3–25 August) (Figure 2).Ethylene production in control bottles with no acetyleneadded was 0.17 ± 0.28 mmol C2H4 m

−2 d−1 (1s). Nitrogenfixation was significantly negatively correlated to DIN (r =0.463, p = 0.003, n = 40, Spearman’s correlation, two‐tailedt test; Figure 2a) and to TIN (r = 0.731, p < 0.001, n = 40,Spearman’s correlation, two‐tailed t test; Figure 2b). Therewas no significant correlation between nitrogen fixation andPON (r = 0.056, p = 0.734, n = 40, Spearman’s correlation,two‐tailed t test).3.2.2. Laboratory Nitrogen Fixation[17] Nitrogen fixation rates in both the AB and VB

experiments were linear over 24 h (r2 = 0.99 for AB, and r2 =0.98 for VB, n = 8, slopes of regression lines in bothexperiments significantly different from zero, p < 0.001;Figure 4). This indicates that the relatively long incubationperiod of 24 h used for in situ incubations is not likely to haveadversely affected rate measurements. The precision (C.V.)of triplicate rate measurements after 24 h were 47% and 40%for the AB and VB experiments, respectively.

4. Discussion

4.1. Importance of Nitrogen Fixation for the NitrogenBudget of Svalbard Glaciers

[18] This is the first study to give direct evidence thatmicrobes can fix nitrogen on glacier surfaces (Figure 2).Here we estimate the importance of nitrogen fixation rela-tive to other nitrogen inputs to the glaciers. Previous esti-mates of the nitrogen inputs from snow, rain and ice melt toAB and ML (but not VB) on a glacier catchment scale havebeen made by Hodson et al. [2005] for the 1998/1999season (ML only) and 1999/2000 season (ML and AB). We

convert the nitrogen fixation rates of this study to the unitsof kg N km−2 d−1 used by Hodson et al. [2005] using

N2 f ixationannual ¼ N2 f ixationdaily � coverage� time; ð1Þ

where N2 fixationannual is the annual input of nitrogenfrom nitrogen fixation onto the glacier (kg N km−2 yr−1),N2 fixationdaily is the range of daily nitrogen fixation ratein cryoconite holes of this study (mean ±1s) in units of

Figure 3. (a) Particulate organic nitrogen (PON) versus TIN of cryoconite debris. Open circles are earlymelt season (slush zone) samples; solid circles are middle/late melt season samples. There was no signif-icant correlation between PON and TIN when all samples were included. However, there was a significantcorrelation between PON and TIN when slush zone samples were excluded (r = 0.529, p < 0.01, n = 30,Spearman’s correlation, two‐tailed t test). (b) Total organic carbon (TOC) versus total nitrogen (TN) ofcryoconite debris.

Figure 4. Time course experiments testing the linearity ofethylene (C2H4) production over 24 h using cryoconite fromAustre Brøggerbreen (open circles) and Vestre Brøggerbreen(solid circles). Each experiment used three replicate labora-tory incubations at 0.1°C under fluorescent lights. All incuba-tion bottles in these experiments were preincubated underfluorescent lights for 48 h prior to commencing the experi-ments to ameliorate any adverse effects due to freezing.The r2 values were determined by linear regression.

TELLING ET AL.: NITROGEN FIXATION ON ARCTIC GLACIERS G03039G03039

4 of 8

kg N km−2 d−1, coverage is the estimated areal cryoconitecoverage of the glaciers as a fraction of total surface area(0.01 [Hodson et al., 2007]), and time is the typicallength of a melt season on the glaciers (60 days [Hodsonet al., 2007]).[19] The nitrogen inputs to the glaciers are dominated by

snowmelt DIN, particulate nitrogen (PN) (the latter of whichmay be dominantly PON [Hodson et al., 2005]), rainfall,and to a lesser extent DON (Figure 5a). The high degree ofvariation in rainfall TIN estimates (Figure 5) reflects ananomalously high input of DIN from early summer rainfallon ML in 1998/1999 due to the rapid transport of anthropo-genic pollution from western Europe [Hodson et al., 2009].This extreme nitrogen deposition event resulted in inputs ofrainfall DIN on ML (82.5 kg N km−2 yr−1) exceeding inputsfrom snowmelt DIN (26.6 kg N km−2 yr−1) [Hodson et al.,2005]. The 1999/2000 melt season likely represents a moretypical year when inputs of snowmelt DIN on ML andAB (16.3 ± 0.7 kg N km−2 yr−1) exceeded those of rainfallDIN (12.5 ± 2.40 kg N km−2 yr−1) [Hodson et al., 2005].Using the more typical data from 1999/2000, snowmelt andrainfall nitrogen inputs (DIN, PN and DON) comprised99.1% of TN inputs to the glaciers while ice melt contrib-uted only 0.34 ± 0.11% (Figure 5). The mean estimatednitrogen fixation input of this study (0.04 ± 0.11 kg N km−2

yr−1; Figure 5) is equivalent to 0.1 ± 0.3% of the mean TNinputs of the 1999/2000 season. Inputs of nitrogen vianitrogen fixation are therefore more than 2 orders of mag-nitude smaller than previous estimates of precipitation onSvalbard valley glaciers, but within an order of magnitude ofprevious estimates of nitrogen input from ice melt.

4.2. Importance of Nitrogen Fixation for Supporting InSitu Microbial Growth on Glaciers

[20] The detection of nitrogen fixation in over half of thesampled cryoconite holes during the middle to late melt

season (16 of 30 holes; section 3.2.1) demonstrates thatnitrogen fixation can be a source of assimilatory nitrogen tosupraglacial microbial communities when alternative sour-ces of nitrogen become limited. The following calculationsestimate whether nitrogen fixation rates are sufficiently largeto sustain a significant fraction of microbial growth incryoconite holes. We compare the nitrogen fixation rates ofthis study with previously determined rates of bacterialproduction (BP) and net ecosystem production (NEP) on thethree valley glaciers. NEP is the net balance between grossphotosynthesis and community respiration, with positivevalues indicating a net increase in carbon (and hence alsopresumably nitrogen [Redfield et al., 1963]) assimilated bythe microbial community as a whole [Telling et al., 2010].In contrast, BP is typically a measure of heterotrophic, butnot phototrophic, microbial growth [Anesio et al., 2011].Rates of NEP and BP on the three glaciers have beenreported as −1.3 ± 5.7 mg C g−1 d−1 and 200.1 ± 208.5 ng Cg−1 d−1, respectively [Anesio et al., 2011; Telling et al.,2010]. We convert these daily microbial rates into annualareal rates via equation (2) to allow comparison with thenitrogen fixation rates and other nitrogen inputs shown inFigure 5:

microbial growthannual ¼ microbial growthdaily � 14

12� 1

6:6� mass � time; ð2Þ

where microbial growthannual is the estimated annual assim-ilatory nitrogen requirement of BP or NEP (kg N km−2 yr−1),microbial growthdaily is the range of BP or NEP rates incryoconite holes (from Anesio et al. [2011] and Telling et al.[2010], respectively) converted to units of kg C km−2 d−1,14/12 converts moles of carbon to moles of nitrogen, 1/6.6 isthe typical nitrogen:carbon ratio during balanced microbialgrowth [Redfield et al., 1963], mass is the typical cryoconite

Figure 5. Nitrogen inputs to Svalbard valley glacier surfaces compared to the estimated nitrogendemands of microbial growth: (a) all data and (b) magnified section showing only ice melt and microbialrate data. Data for snow, rain, and ice are based on previous data from Midtre Lovénbreen and AustreBrøggerbreen [Hodson et al., 2005]. Nitrogen inputs from nitrogen fixation rates were derived fromequation (1) (see text). Nitrogen demands for rates of microbial net ecosystem production (NEP) andbacterial production (BP) were derived from equation (2) (see text). Open circles are mean values; errorbars are ±1s.

TELLING ET AL.: NITROGEN FIXATION ON ARCTIC GLACIERS G03039G03039

5 of 8

mass distribution on the glacier surfaces (10,600 kg−2 km−2

[Hodson et al., 2007]), and time is the typical length of amelt season on the glaciers in days (60 days [Hodson et al.,2007]).[21] The estimated annual nitrogen demands of BP and

NEP on the glaciers are 0.02 ± 0.02 kg N km−2 yr−1 and−0.15 ± 0.64 kg N km−2 yr−1. These are a similar order ofmagnitude to nitrogen inputs from nitrogen fixation (0.05 ±0.21 kg N km−2 yr−1; Figure 5b). These calculations there-fore demonstrate the potential for nitrogen fixation to supporta significant fraction of estimated microbial assimilatorynitrogen requirements when other forms of nitrogen on theglaciers become limited.

4.3. Controls on Nitrogen Fixation Activity

[22] Nitrogen fixation is significantly negatively corre-lated to both DIN and TIN (Figure 2), indicating thatavailable inorganic nitrogen is a strong control on thepresence or absence of nitrogen fixation in cryoconite holes.The negative correlation between nitrogen fixation andavailable nitrogen is consistent with nitrogen fixation inother aquatic and terrestrial environments [Howarth et al.,1988] as nitrogen fixing microbes preferentially take upfixed forms of nitrogen rather than fixing nitrogen owing tothe high relative energetic cost of the latter [Gutschick,1978].[23] It is highly likely however that organic nitrogen

provides additional recycled nitrogen to cryoconite micro-bial communities. PON dominates the nitrogen chemistry ofcryoconite debris (Figure 3a), and cryoconite holes in theDry Valleys of Antarctica can have DON concentrationsthat exceed DIN by an order of magnitude [Tranter et al.,2004]. Unfortunately, DON was not measured on theSvalbard glaciers of this study. However, nitrogen fixationwas only active when TIN was <1.3 mg N g−1 and DINbelow detection (Figure 2). This suggests that DON wasinsufficient to support all microbial assimilatory require-ments in cryoconite holes where nitrogen fixation wasdetected, and hence that labile DON was depleted eithercoincidentally or before the DIN was fully utilized.[24] Snowmelt DIN is a likely source for the measured

concentrations of DIN in early season cryoconite holes. Themaximum concentrations of NO3

−(aq) and NH4

+(aq) (35.5 mg N

L−1 and 95.2 mg N L−1, respectively) were within the rangeof those previously reported for snowmelt on ML (77 ±98 mg N L−1 and 64.4 ± 47.6 mg N L−1 for NO3

−(aq) and

NH4+(aq), respectively [Wynn et al., 2007]). Significant

retention of snowmelt derived NH4+ in the early melt season is

also a likely source of TIN (Figure 2b). Changes in supra-glacial stream chemistry have indicated that the uptake ofNH4

+ from snowmelt principally occurs during the early meltseason [Wynn et al., 2007; Hodson et al., 2009], consistentwith the highest TIN concentrations of this study occurringwithin slush zone samples (Figure 2b).[25] There may be an additional source of inorganic

nitrogen into cryoconite holes from the remineralization oforganic matter, with the organic matter derived either fromwithin snowmelt (PN or DON; Figure 5) or from withincryoconite debris (Figure 3b). The remineralization oforganic matter offers one explanation for the weak but sig-nificant (p < 0.01) positive correlation between cryoconitePON and TIN inmain to late melt season samples (Figure 3a).

A second equally plausible explanation for the positivecorrelation between PON and TIN, however, is that NH4

+

derived from snowmelt, ice melt or rain may preferentiallyadsorb onto organic matter [Seitzinger et al., 1991].[26] A third potential source of inorganic nitrogen to

cryoconite holes is nitrogen fixation. However, the signifi-cant negative relationship between TIN and nitrogen fixation(Figure 2b) indicates that TIN does not accumulate in regionsof nitrogen fixation. Instead NH4

+ produced by nitrogen fix-ation is more likely rapidly incorporated into organic matter,which may then provide an important source of labilerecycled organic nitrogen for other microorganisms.

4.4. Temporal Variation in Nitrogen Fixation

[27] The data in Figure 2 suggest that nitrogen fixation isnot likely in cryoconite holes during the early melt seasonwhen samples are covered in slush, however nitrogen fixa-tion may be active in cryoconite holes during the middle tolate melt season. The apparent temporal variation in nitrogenfixation can be explained by the migration of the snowlineupslope during the middle to late melt season, resultingin supraglacial water chemistry on the lower slopes of theglaciers becoming increasingly dominated by inputs fromice melt rather than snowmelt [Hodson et al., 2005; Wynnet al., 2007]. Snowmelt dominated cryoconite holes in theearly melt season have sufficient DIN and/or TN to preventnitrogen fixation (Figure 2). In contrast cryoconite holes inthe middle to late melt season, with nitrogen inputs likelydominated by ice melt, can have both DIN and TN con-centrations below detection (Figure 2). The transition fromsnowmelt to ice melt dominated supraglacial water can beaccelerated by changes in hydrological routing upslope onthe glaciers. For example snowmelt at higher altitude on MLdoes not reach the lower slopes of the glacier as is routeddown moulins into subglacial channels [Wynn et al., 2007].The greater potential for nitrogen fixation in regions whereice melt dominates the water chemistry is consistent withestimates of the relative inputs of nitrogen from ice melt andsnowmelt on ML [Hodson et al., 2005]. While previousestimates of nitrogen inputs from snowmelt are more than 2orders of magnitude greater than the estimated microbialnitrogen assimilatory requirements on the glaciers (Figure 5),previous estimates of annual nitrogen inputs from ice meltare slightly lower than the highest annual nitrogen require-ments of NEP (Figure 5) and the same order of magnitude asthe estimated mean annual nitrogen requirements of BP(Figure 5).

4.5. Impact of Anthropogenic Nitrogen Deposition onSupraglacial Ecology

[28] The deposition of NO3− and NH4

+ on Svalbard valleyglaciers have increased 65% and 20% since preindus-trial times, respectively, owing to anthropogenic pollution[Kekonen et al., 2005]. By dividing the mean previousestimated NO3

− and NH4+ inputs from snowmelt and rainfall

on AB and ML (data from Hodson et al. [2005]) by 1.65 ×and 1.2 ×, respectively, it can be calculated that 23% (mean of1998/1999 and 1999/2000 data) or 19.5% (1999/2000 dataonly) of TN inputs were likely anthropogenic. Increasesin NO3

− deposition due to anthropogenic pollution havein some cases been even greater elsewhere in the world.Deposition of NO3

− on some Alpine glaciers, the Greenland

TELLING ET AL.: NITROGEN FIXATION ON ARCTIC GLACIERS G03039G03039

6 of 8

Ice Sheet and some central Asian glaciers has increased by90%, 100%, and 130% since preindustrial times, respec-tively [Döscher et al., 1995; Olivier et al., 2006; Fischeret al., 1998], while NH4

+ deposition has increased by 100%on Himalayan glaciers and by 200% on Alpine glaciers[Kang et al., 2002; Döscher et al., 1996].[29] Data from this study show a very clear and significant

relationship between the TIN availability in cryoconite holesand nitrogen fixation. We hypothesize that the large increasesin anthropogenic TIN deposition since preindustrial timesmay have had the potential to reduce the extent of micro-bial nitrogen limitation within cryoconite holes and hencereduced annual rates of nitrogen fixation on glaciers.

5. Conclusions

[30] Microorganisms can fix nitrogen on Svalbard valleyglacier surfaces when other sources of nitrogen becomelimited. Annual inputs of nitrogen by nitrogen fixation onSvalbard valley glaciers are estimated to be more than 2orders of magnitude lower than nitrogen inputs from snow-melt and rain. However, nitrogen fixation may be importantfor supporting microbial growth in cryoconite holes in themiddle to late melt season after the snowline has retreatedupslope. The increase in anthropogenic nitrogen depositionon Svalbard glaciers since preindustrial times may haveaffected microbial growth and activity by reducing themicrobial requirement for nitrogen fixation.

[31] Acknowledgments. This work was funded by grants awarded toA.A. and A.H. from NERC (NE/G00496X/1 and NE/G006253/1). Wewould like to thank Nick Cox for his logistical help at the NERC Arcticstation, Ny‐Ålesund, and Jon Hawkings for his assistance in field sampling.We are also grateful for helpful comments on the draft manuscript byM. Stibal and three anonymous reviewers.

ReferencesAnesio, A. M., A. J. Hodson, A. Fritz, R. Psenner, and B. Sattler (2009),High microbial activity on glaciers: Importance to the global carboncycle, Global Change Biol., 15, 955–960, doi:10.1111/j.1365-2486.2008.01758.x.

Anesio, A. M., B. Sattler, C. Foreman, J. Telling, A. Hodson, M. Tranter,and R. Psenner (2011), Carbon fluxes through bacterial communitieson glacier surfaces, Ann. Glaciol., 51(56), 32–40, doi:10.3189/172756411795932092.

Breitbarth, E., M. M. Mills, G. Friedrichs, and J. LaRoche (2004), TheBunsen gas solubility coefficient of ethylene as a function of temperatureand salinity and its importance for nitrogen fixation assays, Limnol.Oceanogr. Methods, 2, 282–288, doi:10.4319/lom.2004.2.282.

Christner, B. C., B. H. Kvitko, and J. N. Reeve (2003), Molecular identi-fication of Bacteria and Eukarya inhabiting an Antarctic cryoconite hole,Extremophiles, 7, 177–183, doi:10.1007/s00792-002-0309-0.

David, K. A., and P. Fay (1977), Effects of long‐term treatment with acet-ylene on nitrogen‐fixing microorganisms, Appl. Environ. Microbiol., 34,640–646.

Döscher, A., H. W. Gäggeler, U. Schotterer, and M. Schwikowski (1995),A 130 year deposition record of sulphate, nitrate and chloride from a highalpine glacier, Water Air Soil Pollut., 85, 603–609, doi:10.1007/BF00476895.

Döscher, A., H. W. Gäggeler, U. Schotterer, and M. Schwikowski (1996),A historical record of ammonium concentrations from a glacier in theAlps, Geophys. Res. Lett., 23, 2741–2744, doi:10.1029/96GL02615.

DuBois, J. D., and L. A. Kapustka (1983), Freeze recovery physiology ofnitrogenase activity in terrestrial Nostoc sp. colonies, Appl. Environ.Microbiol., 46, 773–778.

Edwards, A., A. M. Anesio, S. M. Rassner, B. Sattler, B. P. Hubbard, W. T.Perkins, M. Young, and G. W. Griffith (2011), Possible interactionsbetween bacterial diversity, microbial activity and supraglacial hydrology

of cryoconite holes in Svalbard, ISME J., 5, 150–160, doi:10.1038/ismej.2010.100.

Fischer, H., D. Wagenbach, and J. Kipfstuhl (1998), Sulfate and nitrate firnconcentrations on the Greenland ice sheet: 2. Temporal anthropogenicdeposition changes, J. Geophys. Res., 103, 21,935–21,942, doi:10.1029/98JD01886.

Gutschick, V. P. (1978), Energy and nitrogen fixation, BioScience, 28,571–575, doi:10.2307/1307512.

Hagen, J. O., J. Kohler, K. Melvold, and J. G. Winther (2003), Glaciers inSvalbard: Mass balance, runoff and freshwater flux, Polar Res., 22,145–159, doi:10.1111/j.1751-8369.2003.tb00104.x.

Hodson, A. J. (2006), Biogeochemistry of snowmelt in an Antarctic gla-cial ecosystem, Water Resour. Res., 42, W11406, doi:10.1029/2005WR004311.

Hodson, A. J., P. N. Mumford, J. Kohler, and P. M. Wynn (2005), TheHigh Arctic glacial ecosystem: New insights from nutrient budgets,Biogeochemistry, 72, 233–256, doi:10.1007/s10533-004-0362-0.

Hodson, A., et al. (2007), A glacier respires: Quantifying the distributionand respiration CO2 flux of cryoconite across an entire Arctic supraglacialecosystem, J. Geophys. Res., 112, G04S36, doi:10.1029/2007JG000452.

Hodson, A., A. M. Anesio, M. Tranter, A. Fountain, M. Osborn, J. Priscu,J. Laybourn‐Parry, and B. Sattler (2008), Glacial ecosystems, Ecol.Monogr., 78, 41–67, doi:10.1890/07-0187.1.

Hodson, A., T. J. Roberts, A.‐C. Engvall, K. Holmén, and P. Mumford(2009), Glacier ecosystem response to episodic nitrogen enrichment inSvalbard, European High Arctic, Biogeochemistry, 98, 171–184,doi:10.1007/s10533-009-9384-y.

Hoffman, P. F., A. J. Kaufman, G. P. Halverson, and D. P. Schrag (1998),A Neoproterozoic snowball Earth, Science , 281 , 1342–1346,doi:10.1126/science.281.5381.1342.

Hood, E., J. Fellman, R. G. M. Spencer, P. J. Hernes, R. Edwards,D. D’Amore, and D. Scott (2009), Glaciers as a source of ancient and labileorganic matter to the marine environment, Nature, 462, 1044–1047,doi:10.1038/nature08580.

Howarth, R. W., R. Marino, J. Lane, and J. C. Cole (1988), Nitrogen fixationin freshwater, estuarine, and marine ecosystems: 1. Rates and importance,Limnol. Oceanogr., 33, 669–687, doi:10.4319/lo.1988.33.4_part_2.0669.

Kang, S., P. A. Mayewski, D. Qin, Y. Yan, D. Zhang, S. Hou, and J. Ren(2002), Twentieth century increase of atmospheric ammonia recorded inMount Everest ice core, J. Geophys. Res., 107(D20), 4595, doi:10.1029/2001JD001413.

Kekonen, T., J. Moore, P. Peramaki, R. Mulvaney, E. Isaksson, V. Pohjola,and R. S. W. van de Wal (2005), The 800 year long ion record from theLomonosovfonna (Svalbard) ice core, J. Geophys. Res., 110, D07304,doi:10.1029/2004JD005223.

Maynard, D. G., Y. P. Kalra, and J. A. Crumgaugh (2007), Nitrate andexchangeable ammonium nitrogen, in Soil Sampling andMethods of Anal-ysis, 2nd ed., edited by M. R. Carter and E. G. Gregorich, pp. 25–38, CRCPress, Boca Raton, Fla.

Nuth, C., J. Kohler, H. F. Aas, O. Brandt, and J. O. Hagen (2007), Glaciergeometry and elevation changes on Svalbard (1936–90): a baseline dataset,Ann. Glaciol., 46, 106–116, doi:10.3189/172756407782871440.

Olivier, S., C. Blaser, S. Brütsch, N. Frolova, H. W. Gäggeler, K. A.Henderson, A. S. Palmer, T. Papina, andM. Schwikowski (2006), Tempo-ral variations of mineral dust, biogenic tracers, and anthropogenic spe-cies during the past two centuries from Belukha ice core, Siberian Altai,J. Geophys. Res., 111, D05309, doi:10.1029/2005JD005830.

Paterson, W. S. B. (1994), The Physics of Glaciers, 3rd ed., 480 pp.,Butterworth‐Heinemann, Oxford, U. K.

Redfield, A. C., B. H. Ketchum, and F. A. Richards (1963), The influenceof organisms on the composition of seawater, in The Sea, vol. 2, editedby M. H. Hill, pp. 26–77, John Wiley, New York.

Säwström, C., P. Mumford, W. Marshall, A. Hodson, and J. Laybourn‐Parry (2002), The microbial communities and primary productivity ofcryoconite holes in an Arctic glacier (Svalbard 79°N), Polar Biol., 25,591–596, doi:10.1007/s00300-002-0388-5.

Säwström, C., J. Laybourn‐Parry, W. Granéli, and A. M. Anesio (2007),Heterotrophic bacterial and viral dynamics in Arctic freshwaters: Resultsfrom a field study and nutrient‐temperature manipulation experiments,Polar Biol., 30, 1407–1415, doi:10.1007/s00300-007-0301-3.

Seitzinger, S. P., W. S. Gardner, and A. K. Spratt (1991), The effect ofsalinity on ammonium sorption in aquatic sediments: Implications forbenthic nutrient recycling, Estuaries Coasts, 14, 167–174, doi:10.2307/1351690.

Stewart, W. D., G. P. Fitgeral, and R. H. Burris (1967), In situ studies onN2 fixation using the acetylene reduction technique, Proc. Natl. Acad.Sci. U. S. A., 58, 2071–2078, doi:10.1073/pnas.58.5.2071.

Stibal, M., M. Šabacká, and K. Kaštovská (2006), Microbial communitieson glacier surfaces in Svalbard: Impact of physical and chemical proper-

TELLING ET AL.: NITROGEN FIXATION ON ARCTIC GLACIERS G03039G03039

7 of 8

ties on abundance and structure of cyanobacteria and algae, MicrobialEcol., 52, 644–654, doi:10.1007/s00248-006-9083-3.

Stibal, M., M. Tranter, J. Telling, and L. G. Benning (2008), Speciation,phase association and potential bioavailability of phosphorus on aSvalbard glacier, Biogeochemistry, 90, 1–13, doi:10.1007/s10533-008-9226-3.

Stutz, R. C., and L. C. Bliss (1973), Acetylene reduction assay for nitrogenfixation under field conditions in remote areas, Plant Soil, 38, 209–213,doi:10.1007/BF00011228.

Telling, J., A. M. Anesio, J. Hawkings, M. Tranter, J. L. Wadham, A. J.Hodson, T. Irvine‐Fynn, and M. L. Yallop (2010), Measuring rates ofgross photosynthesis and net community production in cryoconite holes:A comparison of field methods, Ann. Glaciol., 51(56), 135–144.

Tranter, M., A. G. Fountain, C. H. Fritsen, W. B. Lyons, J. C. Priscu, P. J.Statham, and K. Welch (2004), Extreme hydrochemical conditions in

natural microcosms entombed within Antarctic ice, Hydrol. Processes,18, 379–387, doi:10.1002/hyp.5217.

Wynn, P. M., A. J. Hodson, T. H. E. Heapon, and S. R. Chenery (2007),Nitrate production beneath a High Arctic glacier, Svalbard, Chem. Geol.,244, 88–102, doi:10.1016/j.chemgeo.2007.06.008.

A. M. Anesio, C. Butler, J. Telling, M. Tranter, and J. Wadham, BristolGlaciology Centre, School of Geographical Sciences, University of Bristol,Bristol BS8 1SS, UK. (jon.telling@bristol.ac.uk)A. Hodson, Department of Geography, University of Sheffield, Sheffield

S10 2TN, UK.T. Irvine‐Fynn, Centre for Glaciology, Institute of Geography and Earth

Science, Aberystwyth University, Aberystwyth SY23 3DB, UK.

TELLING ET AL.: NITROGEN FIXATION ON ARCTIC GLACIERS G03039G03039

8 of 8