bioRxiv preprint doi: https://doi.org/10.1101/115808; this version posted April 11, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 1 ASSESSING THE CHEMISTRY AND BIOAVAILABILITY OF DISSOLVED ORGANIC MATTER FROM GLACIERS AND ROCK 2 GLACIERS 3 4 1,2 1,3 3 1,4 5 Fegel, Timothy , Boot, Claudia M. , Broeckling, Corey D. , Hall, Ed K. 6 1. Natural Resource Ecology Laboratory, Colorado State University 2. U.S. Forest Service, 7 Rocky Mountain Research Station, 3. Department of Chemistry, Colorado State University 4. 8 Proteomics and Metabolomics Facility, Colorado State University 4. Department of Ecosystem 9 Science and Sustainability, Colorado State University 10 11 Tim Fegel [email protected] 12 13 14 Key Points: 15 Bioavailability of organic matter released from glaciers is greater than that of rock glaciers in the 16 Rocky Mountains. 17 The use of GC-MS for ecosystem metabolomics represents a novel approach for examining 18 complex organic matter pools. 19 Both glaciers and rock glaciers supply highly bioavailable sources of organic matter to alpine 20 headwaters in Colorado. 1 bioRxiv preprint doi: https://doi.org/10.1101/115808; this version posted April 11, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 21 Abstract 22 As glaciers thaw in response to warming, they release dissolved organic matter (DOM) to 23 alpine lakes and streams. The United States contains an abundance of both alpine glaciers and 24 rock glaciers. Differences in DOM composition and bioavailability between glacier types, like 25 rock and ice glaciers, remain undefined. To assess differences in glacier and rock glacier DOM 26 we evaluated bioavailability and molecular composition of DOM from four alpine catchments 27 each with a glacier and a rock glacier at their headwaters. We assessed bioavailability of DOM 28 by incubating each DOM source with a common microbial community and evaluated chemical 29 characteristics of DOM before and after incubation using untargeted gas chromatography mass 30 spectrometry based metabolomics (GC-MS). Prior to incubations, ice glacier and rock glacier 31 DOM had similar C:N ratios and chemical diversity, but differences in DOM composition. 32 Incubations with a common microbial community showed DOM from ice glacier meltwaters 33 contained a higher proportion of bioavailable DOM (BDOM) and resulted in greater bacterial 34 growth efficiency (BGE). After incubation, DOM composition from each source was statistically 35 indistinguishable. This study provides an example of how MS based metabolomics can be used 36 to assess effects of DOM composition on differences in bioavailability of DOM. Furthermore, it 37 illustrates the importance of microbial metabolism in structuring composition of DOM. Even 38 though rock glaciers had significantly less BDOM than ice glaciers, both glacial types still have 39 potential to be important sources of BDOM to alpine headwaters over the coming decades. 40 2 bioRxiv preprint doi: https://doi.org/10.1101/115808; this version posted April 11, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 41 1.Introduction 42 Ice glaciers (hereafter glaciers) are perennial ice structures that move and persist in areas where 43 annual snow accumulation is greater than annual snow ablation at decadal or longer time spans. 44 Rock glaciers are flowing bodies of permafrost, composed of coarse talus and granular regolith 45 both bound and lubricated by interstitial ice (Berthling, 2011). Both glaciers and rock glaciers 46 integrate atmospherically deposited chemicals with weathering products and release reactive 47 solutes to adjacent surface waters (Williams et al., 2007; Dubnick et al., 2010; Fellman et al., 48 2010; Stibal et al., 2010; Stubbins et al., 2012). On average, alpine glaciers appear to be 49 responsible for the release of a larger flux of carbon (0.58 +/- 0.07 Tg C) from melting ice 50 annually when compared to continental glaciers (Hood et al., 2015). Dissolved organic matter 51 (DOM) from large alpine glaciers of the European Alps and Alaska has been shown to be 52 bioavailable and thus can support microbial heterotrophy (Hood et al., 2009; Singer et al., 53 2012). In the contiguous United States, both glaciers and rock glaciers, each with distinct 54 geophysical attributes (Fegel et al. 2016), are common to many alpine headwaters, with rock 55 glaciers being an order of magnitude more abundant than glaciers. However, little is known 56 about the quantity or quality of DOM being released from these smaller glaciers and rock 57 glaciers in mountain headwater ecosystems of the western U.S. (Woo et al., 2012; Fegel et al., 58 2016). 59 Assessment of the bioavailability of aquatic DOM to date has largely relied on bioassays that 60 measure the rate and amount of DOM consumed over time (e.g. Amon and Benner, 1996; del 61 Giorgio and Cole, 1998; Guillemette and del Giorgio, 2011). Bioassays present difficulties in 62 assessing DOM consumption because each assessment requires an independent incubation, 63 creating the potential for confounding effects due to differences in the microbial communities 3 bioRxiv preprint doi: https://doi.org/10.1101/115808; this version posted April 11, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 64 among incubations. Broad molecular characterizations of DOM (Benner, 2002; Berggren and 65 del Giorgio, 2015) have also previously been applied to estimate bioavailable DOM (BDOM), 66 though these analyses often only address differences in bioavailability of either coarse 67 categories of organic molecules or individual model compounds rather than the composition of 68 the DOM pool as a whole (e.g., del Giorgio and Cole, 1998). Recently the research community 69 has begun to explore high resolution analytical chemistry to assess the molecular 70 characteristics of environmental DOM (e.g., Kujawinski, 2002; Kellerman et al., 2014, 2015; 71 Andrilli et al., 2015). Whereas no single method can identify the entire spectrum of compounds 72 present in an environmental DOM pool (Derenne and Tu, 2014), mass spectrometry (MS) has 73 the ability to define structural characteristics of thousands of individual compounds contained 74 within a single DOM sample. 75 There is a suite of challenges in trying to concurrently understand the composition and 76 bioavailability of DOM. Difficulties in experimentally linking the molecular characterization of 77 DOM pools to the proportion of BDOM are partly due to the highly diverse constituent 78 compounds that compose natural DOM and the inherent challenges in characterizing that 79 diversity (Derenne and Tu, 2014). In addition, different microbial communities may access 80 different components of the same DOM pool making it challenging to compare how composition 81 affects bioavailability among ecosystems. However, even in the face of these challenges some 82 broad patterns of the relationship between the composition and bioavailability of DOM are 83 beginning to emerge. For example, studies from glacial, estuarine, and marine environments 84 have shown a clear correlation between proteinaceous DOM and a high proportion of BDOM 85 (Andrilli et al., 2015). Understanding which constituent molecules influence the bioavailability 86 of DOM is an important step in understanding how DOM pools contribute to heterotrophy 4 bioRxiv preprint doi: https://doi.org/10.1101/115808; this version posted April 11, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 87 among aquatic ecosystems and how different components of DOM alter the residence time of 88 that DOM pool in the environment. 89 We hypothesized that differences in the origin of DOM between glaciers and rock glaciers 90 would result in differences in molecular composition of DOM between glacier types with the 91 potential to alter the proportion of BDOM. DOM derived from glaciers originates from in situ 92 microbial activity via phototrophy on the glacier surface, and byproducts of heterotrophic 93 bacterial activity in the glacier porewaters and subsurface (Anesio et al., 2009; Hood et al., 2009; 94 Singer et al., 2012; Fellman et al., 2015; Fegel et al., 2016). In addition, atmospheric deposition 95 of organic matter onto the glacier surface can be an important source of DOM to the glacier 96 meltwaters (Stubbins et al., 2012). DOM from rock glaciers is an amalgam of compounds 97 originating both from multicellular vegetation growing on the rock glacier surface and microbial 98 processing within the rock glacier itself (Wahrhaftig and Cox, 1959; Williams et al., 2007). 99 Previous research has shown that DOM released from glaciers and rock glaciers in the western 100 United States differ in their optical properties, with glaciers having higher amounts of protein- 101 like DOM than rock glaciers, as identified through fluorescence spectroscopy (Fegel et al., 102 2016). 103 In this study, we assessed differences in the composition of DOM between glaciers and rock 104 glaciers and asked which of those differences (if any) correspond to differences in the proportion 105 of BDOM from each glacier type.