Modern calibration of Poa flabellata (Tussac grass) as a new paleoclimate proxy in the South Atlantic

Terrestrial paleoclimate records are rare in the South Atlantic, limiting opportunities to provide a prehistoric context for current global changes. The tussock grass, Poa flabellata, grows abundantly along the coasts of the Falkland Islands and other sub-Antarctic islands. It forms extensive peat records, providing a promising opportunity 10 to reconstruct high-resolution regional climate records. The isotopic composition of leaf and root tissues deposited in these peats has the potential to record variation in precipitation, temperature, and relative humidity over time, but these relationships are unknown for P. flabellata. Here, we measured the isotopic composition of P. flabellata and precipitation and explore relationships with seasonal temperature and humidity variations across 4 study locations in the Falkland Islands. We reveal that inter-seasonal differences in carbon and oxygen stable isotopes of leaf α-cellulose 15 of living P. flabellata correlated with monthly mean temperature and relative humidity. The carbon isotope composition of leaf α-cellulose (δ13Cleaf) records the balance of CO2 supply through stomata and the demand by photosynthesis. The positive correlation between δ13Cleaf and temperature and negative correlation between between δ13Cleaf and relative humidity suggest that photosynthetic demand for CO2 relative to stomatal supply is enhanced when conditions are warm and dry. Further, the positive correlation between δ13Cleaf and δ18Oleaf (r = 0.88, p < 0.001, 20 n = 24) indicates that stomatal closure during warm dry periods explain seasonal variation in δ13Cleaf. We observed significant differences between winter and summer seasons for both δ18Oleaf and δ13Cleaf, and among study locations for δ18Oleaf, but not δ13Cleaf. δ18O values of monthly composite precipitation were similar between seasons and among study locations, yet characteristic of the latitudinal origin of storm tracks and seasonal winds. The weak correlation between δ18O in monthly composite precipitation and δ18Oleaf further suggests that relative humidity is the main driver 25 of the δ18Oleaf. The oxygen isotopes in root α-cellulose did not reflect, or only partially reflected (at one study location), the δ18O in precipitation. Overall, this study supports the use of peat records formed by P. flabellata to fill in a significant gap in our knowledge of the long-term trends in Southern Hemisphere climate dynamics.

animals in the Southern Ocean , and warming of the western South Atlantic is projected to alter the distribution of island plants as well (Jones et al., 2013;Upson et al., 2016). Meteorological measurements 35 from the Falkland Islands date back to 1874 and are not continuous (Lister and Jones, 2015). This means we lack critical information on the long-term patterns and whether these are novel conditions. Paleoecological archives, such as high-resolution lake sediments and tree rings, can provide useful long-term records documenting and quantifying changes (Dietl et al., 2015;Dietl and Flessa, 2011;Willis et al., 2010), but such records are lacking for the South Atlantic. The absence of trees and deep lakes across many sub-Antarctic islands especially limits high-resolution, 40 independent paleoclimate reconstructions, which are essential for detecting past abrupt climate change. However, many sub-Antarctic islands support widespread communities of peat-forming C3 tussock grasses (Poa flabellata), which provide important habitat and shelter for breeding marine animals such as seals and seabirds. Peat records formed by P. flabellata present a promising avenue for paleoclimate reconstructions similar to other peatland vegetation types in mid-to high latitudes (Amesbury et al., 2015;Chambers et al., 2012;Pendall et al., 2001). 45 P. flabellata grasslands in the South Atlantic generate substantial amounts of peat (Smith and Clymo, 1984), and have the highest carbon accumulation rates of any peatland globally (Payne et al., 2019). Endemic to the South Atlantic, P. flabellata only occurs on Tierra del Fuego, the Falkland Islands, Gough Island, and South Georgia. P. flabellata grasslands were once widespread throughout the Falkland Islands, but are now greatly reduced because of land-use change and introduced grazers (Strange et al., 1988;Wilson et al., 1993). The term "tussock" is used to describe the 50 clumping growth form of P. flabellata, while the species itself is commonly known as "tussac." Several factors support the utility of P. flabellata peats as a paleoclimate proxy. P. flabellata peatlands are formed by a nearly single-species community of P. flabellata, which allow very little light or space for other plants to co-occur in the absence of disturbance. Tillers of P. flabellata grow on top of a pedestal of decaying roots and leaves that can reach 4 meters high (Fig. 1e) (Smith and Clymo, 1984); mature plants thus likely primarily use water from 55 precipitation, as they are not rooted in the soil directly, or from evaporatively enriched water in the decaying pedestal. Smith and Prince (1985) established radiocarbon ( 14 C) dates for a P. flabellata pedestal and estimated an age of 250 to 330 years. P. flabellata grass forms extensive peat deposits of up to 13.3-m deep, with carbon accumulation rates of 139 g C m -2 yr -1 (Payne et al., 2019;Smith and Clymo, 1984), far greater than peatlands of similar latitude in the Northern Hemisphere (18.6 g C m -2 yr -1 ), the tropics (12.8 g C m -2 yr -1 ) or Patagonia (22 g C m -2 yr -1 ) (Yu et al., 60 2010). Subfossil P. flabellata leaves are abundant in these peats, and readily can be separated from root subfossils.
Grasses exhibiting the tussock growth form often have evergreen leaves and exhibit a profligate/opportunistic water 65 use strategy, due to the high evaporative conditions and pulses of water availability in semi-arid habitats (Moreno-Gutiérrez et al., 2012;Sala et al., 1989;Schwinning and Ehleringer, 2001). The growth phenology of P. flabellata is such that it mainly increases in height in summer (~ 39 cm per year) while in winter an increase in basal area occurs with the production of new tillers at the base of the pedestal (Stanworth and Upson, 2013).
Stable isotopes of oxygen (δ 18 O), hydrogen (δD), and carbon (δ 13 C) in the cellulose of plant tissues (roots, shoots, and 70 leaves) can reliably record the climate signal related to environmental growing conditions (Araguás-Araguás et al., 2000). δ 13 C and δ 18 O record species' water-use strategies in water-limited environments because of physiological responses such as changes in stomatal conductance and assimilation rates (Farquhar and Sharkey, 1982;Moreno-Gutiérrez et al., 2012). Tussock grasses typically occur in water-limited environments where conservative water use strategies are common functional traits that allow tussock grasses to take advantage of pulses of water (Moreno-75 Gutiérrez et al., 2012). Correlations between δ 18 O of plant cellulose and air temperature and humidity provide information on environmental conditions in the season the cellulose tissue was formed. The δ 18 O of leaf water is a primary driver of δ 18 O in leaf cellulose, and is influenced by the δ 18 O value of plant source water, temperature, humidity, and plant physiology (Helliker and Ehleringer, 2002;Roden and Ehleringer, 1999). The δ 18 O of source water often correlates with temperature of the environment (Libby et al., 1976). Apart from water source, δ 18 O of 80 cellulose can also be influenced by internal exchange among organic molecules and plant water pools (Sternberg et al., 1986). The δ 13 C value of leaf biomass in C3 plants records δ 13 C of source CO2 and the expression of fractionation effects associated with CO2 diffusion into and through leaf tissue and carboxylation (Farquhar et al., 1982). The net discrimination against 13 C during photosynthesis is driven by changes in the supply of CO2 through stomatal pores and demand for CO2 by photosynthetic biochemistry (Cernusak et al., 2013;Farquhar et al., 1982;Ferrio and Voltas, 85 2005). The δ 13 C value of roots tends to be 1-3 per mil (‰) higher than that of leaves due to a number of postphotosynthetic biochemical fractionations and C allocation pathways (Cernusak et al., 2009).
Plant species vary in the way they isotopically record precipitation and temperature; therefore, peat comprised of a single species is more desirable over a mixture of species (van Geel and Middeldorp, 1988). To test the potential of P. flabellata peatlands as a paleoclimate proxy, we conducted a modern calibration study. We measured δ 18 O and δ 13 C 90 from living P. flabellata leaf tissues (α-cellulose) collected monthly at four sites across the Falkland Islands (51° S, 59° W; Fig. 1b-c). We aim to improve our understanding of Southern Hemisphere climate dynamics with a new paleoclimate proxy that leverages the unique properties of P. flabellata.

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The Falkland Islands (Fig. 1a) are located approximately 500 km east of southern South America, between 51˚0.5' S to 52˚28.0' S and 61˚22.0' W to 57˚40.5' W. The cool-temperate (mean temperature : January 9.4 °C and July 2.2 °C) climate of the Falkland Islands is driven by the cold Antarctic Circumpolar Current, the waters surrounding the Antarctic Peninsula, the Falklands Current, and the Andes of southern Patagonia to the west (Turner and Pendelbury, 2000). The persistent winds of the southwesterly wind belt average 8.5 m s -1 (30 km h -1 ), with gale 100 force winds averaging 70 days per year and annual precipitation generally ranges between 400 to 600 mm (Jones et al., 2013;Lister and Jones, 2015). Study sites were selected to reflect 1) climatic diversity, and 2) the availability of volunteers to collect monthly samples for one year. We ultimately selected four sites (Fig. 1b-c): Bleaker Island, Cape Dolphin, Surf Bay, and West Point Island.

Precipitation, temperature and relative humidity
Precipitation was collected at each site using a Palmex monthly composite collection sampler (Palmex d.o.o., Zagbreb, Croatia). The Palmex collector is designed to prevent evaporation and evaporative enrichment of 18 O in precipitation samples without the use of paraffin oil (Gröning et al., 2012), and has been recommended by the Global Network of Isotopes in Precipitation (GNIP). δ 18 O and δD stable isotope ratios of water samples were measured at the University of Wyoming Stable Isotope Facility (UWSIF) using a high-temperature conversion elemental analyzer (TCEA) 110 connected to a Thermo Scientific Delta V Plus that is run in continuous flow mode via a ConFlo IV. The technique used injections of 1 µl of water into the TCEA column filled with glassy carbon heated to 1420 °C. Precipitation samples were purified using cryogenic vacuum distillation (Ehleringer and Osmond, 1989) prior to TCEA analysis to remove aeolian debris, including marine salts. Internal QA/QC working standards calibrated against IAEA international standards Vienna Standard Mean Ocean Water (VSMOW) and Standard Light Antarctic Precipitation 115 (SLAP) and spanning the range of measured values in our study were analyzed with each batch of samples with analytical precision typically better than 0.3 and 2.5 ‰ for δ 18 O and δD, respectively. Isotope values are reported with respect to VSMOW. Explanations of methods for daily average temperature and relative humidity measurements, as well as seasonal wind speed, wind direction, and back trajectory models to determine origins of air masses are found in the Supplemental Text 1.

Poa flabellata field collection and cellulose extraction, and isotope analyses
Poa flabellata plants were collected at the start of each month at each site from October 2015 through September 2016, from relatively uniform habitats that were undisturbed by grazing or tilling. Up to six P. flabellata plant tillers (leaves, stem, and roots) were collected near each of the four stations each month. Whole plants were stored in paper envelopes stored in a cool, dark, dry location until frozen. Samples collected between September 2015 to February 2016 were 125 frozen in February 2016 and samples collected in March 2016 to August/September 2016 were frozen in August/September 2016. Samples were frozen for eight days at the Falkland Islands Department of Agriculture to comply with U.S. Department of Agriculture permitting to prevent the spread of pests. For leaf material, the inner developing (youngest) leaves were collected and assumed to represent the past month of growth. There was no indication that leaves were morphologically different between summer and winter. Only coarse roots were used, fine 130 roots were excluded. Variation in environmental conditions during the growth of a leaf blade can lead to isotopic variations along the gradient of a single leaf as has been shown with δ 18 O of cellulose Ehleringer, 2000, 2002); therefore, whole-leaf plant samples were homogenized by drying at 50 °C and pulverizing using a Retsch ball mill at the University of Maine. For each sample, we used 20 mg of pulverized and homogenized leaf or root material for extraction and purification of α-cellulose, following an adapted procedure of (Brendel et al., 2000). Samples were 135 vortexed throughout extraction and purification for homogenization and were visually inspected for purity. Further indicators of purity include low amounts of nitrogen content (0.13 % to 0.16 % N), and analysis of carbon content (42.1 % to 42.8 % C) in cellulose. As an internal quality control, one leaf sample was selected for extraction and purification of α-cellulose throughout the sample processing in batches of 10 to 12 samples. The δ 13 C and δ 18 O of leaf cellulose for the internal quality control samples varied by < 0.1 ‰ and 0.3 ‰, respectively. 140 δ 18 O and δ 13 C stable isotope ratios of α-cellulose samples were measured at the University of Wyoming Stable Isotope Facility (UWSIF). Oxygen was analyzed using a TCEA coupled to a Thermo Delta V IRMS; δ 18 O values are expressed relative to VSMOW (Craig, 1961;Gonfiantini, 1978). Values were normalized to the VSMOW scale using USGS-42 (8.6 ‰ accepted value) and IAEA-601 cellulose (31.9 ‰ accepted value) quality control standard reference materials for oxygen isotopic composition. Analytical precision was ± 0.3 ‰ for δ 18 O based on repeated analysis of internal 145 standards, and samples loaded into silver capsules had weights ca. 0.25 mg. Carbon isotope composition of the cellulose samples were determined using a Costech 4010 Elemental Analyzer coupled to a Thermo Delta Plus XP-IRMS. Analytical precision was ± 0.1 ‰ for δ 13 C based on repeated internal standards. Quality control standard reference material for carbon isotopic composition included USGS-40 glutamic acid (-28.3‰ accepted value), USGS-41 glutamic acid (24.4 ‰ accepted value), and internal UWSIF α-cellulose (-24.9 ‰ accepted value). Carbon samples 150 weighed ca. 2 mg and were loaded into tin capsules.

Statistical analysis
For both δ 18 O and δ 13 C, we analysed the average of three to four plant leaf samples per month for summer (DJF) and winter (JJA) season at each of the four sites, and the average of up to eight plant root samples (Supplementary Data 1 and 2). Because samples were collected at the start of each month, we define summer as the months of DJF and winter 155 as the months JJA. We used Pearson's correlation coefficient, r, to detect associations between δ 18 O of cellulose and precipitation samples to test whether δ 18 O samples reflects the isotopic value of precipitation. Using Pearson's correlation coefficient, we tested for a relationship between δ 13 C and δ 18 O values of cellulose, temperature, and relative humidity. We tested for a significant difference between summer and winter δ 18 O and δD in precipitation using a ttest (n = 24). A separate one-way analysis of variance (ANOVA) to test for significant differences among sites 160 included δ 18 O and δD in precipitation samples from the entire year (n = 47).
A two-way ANOVA compared the main effects of season (summer vs. winter) and the four study locations on the δ 18 O and δ 13 C stable isotopes of α-cellulose of P. flabellata leaves and roots grown in the summer versus winter, followed by a post hoc test (Tukey's multiple comparison of means). P-values < 0.05 are considered significant.
Descriptive and multivariate analyses were conducted with SigmaPlot 12.5. 165 3 Results

Environmental measurements
Across all sites, summer daily average temperatures ranged from 3.5 °C to 15.6 °C (mean = 10.0 °C) and relative humidity ranged from 64.2 % to 98.1 % (mean = 81.1 %). Winter daily average temperatures ranged from -1.8 °C to 7.6 °C (mean = 3.7 °C), and relative humidity ranged from 73.6 % to 100 % (mean 94.3 %). Seasonal temperature 170 (°C) and relative humidity (%) minimum and maximum ranges for individual study locations are found in Table S1.

Wind
The wind rose (Fig. 1d) shows that winter winds at Bleaker Island primarily blew from the west and northwest. In 175 winter, two spokes in the west and NNW direction comprise >30% of the total recorded 15-minute wind directions.
In summer, three spokes in the west, WSW, and SW directions comprise >45% of all 15-minute wind directions. The wind rarely blew from the east, SE, or north. Examining winds from the west in winter, >10% of wind speeds recorded were between 5 and 10 m s -1 (18 and 36 km h -1 ), and the frequency of strongest winds came from the NNE. In summer, >20% winds from the SW were between 5 and 10 m s -1 , and there was a higher frequency of 10 to 15 m s -1 180 (36 to 54 km h -1 ) wind speeds than in winter. Seasonal wind variation deviated from the long-term average . Reanalysis data (ERA Interim; Fig. S2) indicated that the wind speeds during summer (DJF 2015 to 2016) were stronger over the Falkland Islands (5 to 6 m s -1 ) (18 to 21 km h -1 ) and weaker during winter (JJA 2016).

Seasonal HYSPLIT air mass trajectory analyses
The daily back trajectory HYSPLIT analysis indicated that during the summer, 89% of the air masses originated (n = 185 344 individual trajectories) west of the Falkland Islands. Approximately 11% of summer air masses originated south of the Falkland Islands near the Antarctic Peninsula. In winter, 79% of the air mass back trajectories (n = 332 individual trajectories) were from the west, NW, and SW, while 21% of air masses had backward trajectories south of the Falkland Islands near the Antarctic Peninsula (Fig. 3).

Discussion
Significant inter-seasonal differences in δ 13 Cleaf and δ 18 Oleaf indicate that P. flabellata tissues record high-resolution patterns of environmental change, supporting the use of P. flabellata peat records as a paleoenvironmental proxy. Ferrio and Voltas (2005) established a positive correlation between δ 13 Cleaf and vapor pressure deficit suggesting stomatal conductance is sensitive to atmospheric moisture conditions. The observed positive correlation between 240 δ 13 Cleaf and temperature suggests higher temperatures led to an increased assimilation rate and reduced discrimination against δ 13 C as shown in other vascular plant studies (Ferrio and Voltas, 2005;Ménot and Burns, 2001). Variation in δ 13 Cleaf is driven by changes in the ratio of CO2 partial pressure in the leaf and that of the ambient air, and can be explained by a greater influence of either stomatal conductance or increased photosynthetic capacity (Scheidegger et al., 2000). As plant stomata close in response to low humidity and/or high evaporative conditions like high wind 245 speeds in the Falkland Islands, the internal partial pressure of CO2 decreases and the δ 13 Cleaf increases (Farquhar et al., 1982, p.198).
The δ 18 Oleaf is influenced by soil water, leaf water enrichment of 18 O from transpiration, and biochemical fractionations.
Leaf water enrichment of 18 O due to transpiration, which is reflected in δ 18 Oleaf (Deniro and Epstein, 1979;Roden and Ehleringer, 1999;Sternberg et al., 1986, p.198;Yakir, 1992), depends on relative humidity (Helliker and Ehleringer, 250 2002). However, the relationship between relative humidity and δ 18 Oleaf deteriorated at relative humidity > 90 % in one C3 species (Helliker and Ehleringer, 2002). Diffusion limitation by stomatal resistance is primarily driven by relative humidity (White et al., 1994). The δ 18 Oleaf and relative humidity (> 60 %) were negatively correlated, which is consistent with other studies showing that δ 18 Oleaf increases as relative humidity decreases (Barbraud et al., 2012;Helliker and Ehleringer, 2002). At high relative humidity the leaf will more strongly record variation in the isotopic 255 composition of atmospheric vapor, however we have no direct measurements of δ 18 O in water vapor.
Previous work on leaf water and cellulose in grasses demonstrated that the atmospheric-leaf vapor conditions are a strong predictor of δ 18 Oleaf (Helliker and Ehleringer, 2002;Lehmann et al., 2018). The δ 18 O of the leaf water is captured in the cellulose isotopes and can reflect the effect of changing environmental conditions during the growth of the leaf (Helliker and Ehleringer, 2002;Lehmann et al., 2017). The δ 18 Oleaf can also depend on physiological effects, the type 260 of plant anatomical feature used, and stage of development (Lehmann et al., 2017;Liu et al., 2017). Because cellulose records environmental variation along a gradient during leaf growth, we collected and homogenized whole leaves to avoid the complications of δ 18 O enrichment (Helliker and Ehleringer, 2002). Our work supports the finding that atmospheric-leaf vapor conditions are reflected in δ 18 Oleaf, and expands the use of such paleoclimate proxies to peatforming tussock grasses, which opens up new possibilities for reconstructing paleoclimates across the South Atlantic and beyond. At the higher relative humidity range, the δ 18 Oleaf is more of a reflection of source water, while the δ 18 Oleaf at low humidity differed greatly from source waters because of evaporative enrichment of 18 O.
The positive correlation between δ 13 Cleaf and δ 18 Oleaf (Pearson's r = 0.88, p < 0.001, n = 24; Table 1; Fig. 5 suggests that stomatal conductance is the driving force acting on these two proxies, which is a likely scenario when water is not limiting (Saurer et al., 1997;Scheidegger et al., 2000). According to the Scheidegger et al.(2000) model, the 270 decline in stomatal conductance was much more strongly expressed than photosynthetic capacity (maximum net photosynthesis). When air humidity increases, stomatal conductance is assumed to increase. In our study, stomatal conductance is likely driving both δ 13 Cleaf and δ 18 Oleaf due to relative humidity (Fig. 7). This pattern fits well with the Barbour and Farquhar (2000) model. Measurements of relative humidity allowed us to determine that stomatal conductance was more influential as a possible cause of change in partial pressure of CO2 within the leaf.

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In the Falkland Islands, precipitation amount is not highly seasonal, but tends to vary the most in the summer ( Fig. S3; data from Jones et al., 2013). Over the year of our study, summer δ 18 O in precipitation varied more than winter δ 18 O and tended to be more depleted. This is supported by the wind rose from Bleaker Island, which indicated prevailing winds from the SW (Fig. 1d). In winter, precipitation tended to be more enriched in 18 O, and most prevailing winds came from the west and NW where 18 O in equatorward precipitation would be more enriched than 18 O in high latitude 280 meteoric sources (Fig. 4b). The significant negative correlation between δ 18 O in precipitation and δ 18 Oroot at Cape Dolphin (Pearson's r = 0.868, p = 0.025, n = 6; Table 1) is consistent with the latitudinal origin of storm tracks (Fig.   3) and seasonal wind data (Fig. 1d). The pattern found at Bleaker Island is less clear, and warrants further investigation.
The observed lack of correlations between δ 18 Oleaf and δ 18 O values of precipitation (Table 1) (Table 1). At Cape Dolphin, greatest enrichment of 18 Oroot occurred in summer when δ 18 O precipitation was relatively low. In contrast, at Cape Dolphin δ 18 Oroot was lowest during the winter months when δ 18 O of precipitation was high. Despite the strong relationship between δ 18 O precipitation and δ 18 Oroot at Cape Dolphin, we 295 propose that P. flabellata peatlands may not be ombrotrophic, and may potentially source water from fog, sea-spray, groundwater, or a mix. However, the relationship found at Cape Dolphin warrants further analysis of δ 18 Oroot and source waters. We also consider that the δ 18 Oroot records a signal of leaf humidity, and is influenced by source water and humidity effects on leaves; sugars produced in leaves are transported to roots to form cellulose, and about half of the oxygen atoms in root cellulose originate from the leaf water signal. Roots of P. flabellata may have represented greater temporal integration of δ 18 O from precipitation into cellulose due to difficulty in distinguishing new growth in roots, like Empodisma in New Zealand (Amesbury et al., 2015).
We did not find a relationship between δ 18 O in precipitation and the δ 18 Oleaf in our one-year study. It may be that because the δ 18 O in leaf water is controlled by source water and humidity, any changes in humidity confound a direct relationship between source water δ 18 O values and δ 18 O of leaf water and δ 18 Oleaf. Still, the δ 18 O in plant water pools 305 and δ 18 Oleaf are primarily influenced by δ 18 O in precipitation. It may also be possible that the variation in source area did not affect the δ 18 O enough for us to detect a significant impact on the δ 18 Oleaf. Based on this work, the source area could influence δ 18 Oleaf in peat records if source area changes were greater than what we observed. Without experimentation we can not disentangle temperature and humidity using δ 13 C and δ 18 O; further work is needed to understand the relationship between δ 18 Oroot, δ 18 O of precipitation, and that of root and leaf waters. Identifying sources 310 of potential water would also add value, especially considering anecdotes of local differences in fog in the Falkland Islands, which may be an unappreciated source of water for P. flabellata. Across the geographic range in the South Atlantic, P. flabellata may record a larger latitudinal gradient of isotopes in precipitation, as well as temperature and humidity, than recorded in the Falkland Islands.
Despite this limitation, establishing the seasonal patterns recorded by P. flabellata cellulose in the Falkland Islands 315 does enable us to test paleoclimate hypotheses regarding the climate dynamics in the South Atlantic (e.g. Turney et al., 2016) and Southern Hemisphere westerly wind behavior within the regions where P. flabellata occurs. As with any paleoenvironmental reconstruction, inferences are constrained by the temporal grain and resolution (Jackson, 2012). While P. flabellata is sensitive to inter-seasonal differences and forms highly productive peat records, we stress that paleoclimate reconstructions from P. flabellata peats will represent an integrated signal of broader climate trends, 320 and not annual-scale or seasonal records. Our calibration study is based on a modern snapshot of environmental conditions influencing δ 13 Cleaf and δ 18 Oleaf values in P. flabellata, so investigators conducting downcore peat reconstructions using δ 18 Oleaf must consider that changes in δ 18 Oleaf are modified by precipitation source and changes in humidity through effects on stomatal conductance. We recommend measuring δ 13 Cleaf and δ 18 Oleaf of subsamples with comparable time-averaging (leaf, root, bulk subfossils; Fig. 8) by incorporating a large sampling of multiple leaf 325 fragments (n ³ 10) from each horizon (e.g. 1 cm intervals), and to interpret this signal to integrate multiple years (as determined by the sediment accumulation rate; Fig. 8). Time-averaging within a given 1 cm horizon of even highly productive peat records with sedimentation rates ~ 20 to 30 yr cm -1 would be greater than the age of an individual tussac leaf.
Peat-based reconstructions may be limited to identifying periods of warm/dry or cold/wet conditions that are similar 330 to (or more extreme than) the observed seasonal variations we report here. Thus, we believe that δ 13 Cleaf and δ 18 Oleaf time-averaged values can reliably be used to identify the timing of transitions between warmer and drier conditions to colder and wetter conditions that correlate with major hemispheric drivers in climate. Resolving temperature and moisture signals independently would likely require growth chamber or warming experiments, which was beyond the scope of this study, but which could help develop this proxy further. For now, this proxy remains a qualitative indicator, though it has potential to become a quantitative reconstruction if evaluated via experimentation or in tandem with other plant or microbial biomarkers.

Conclusion
The scarcity of terrestrial paleoclimate records in the South Atlantic has limited our understanding of past and future climate change and its impacts on ecosystems and people. We found that δ 13 C and δ 18 O stable isotope values in P.

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flabellata tissues are correlated with seasonal differences in temperature and moisture, providing a promising new avenue for paleoclimate reconstructions in the South Atlantic. P. flabellata peats have high accumulation rates, contain abundant leaves, and date back to at least 12,500 14 C years, with the potential to provide decadal-scale records of temperature, precipitation, and moisture source. Future work is needed to determine whether δ 18 O and δ 13 C of cellulose from P. flabellata macrofossils complement other regional proxies for changes in atmospheric temperature and 345 relative humidity during the Holocene. Troublingly, these coastal peatlands are currently threatened by sea level rise and over-grazing, and their reductions means we are losing vital information about past environments in a time when paleoclimate records are needed to provide context for modern climate change in the South Atlantic.

Data availability
Datasets for monthly stable isotopes in precipitation, average temperature, and average relative humidity have been 350 submitted to the Global Network of Isotopes in Precipitation (https://nucleus.iaea.org/wiser) and will be publicly available upon acceptance for publication. Datasets for leaf and root stable isotopes of cellulose can be found at http://dx.doi.org/10.5281/zenodo.3104573, hosted at Zenodo upon acceptance for publication.

Author contribution
DG, DW, and JG designed the experiments and DG carried them out. DG performed laboratory analyses. DG prepared 355 the manuscript with contributions from all co-authors.

Competing interests
The authors declare that they have no conflict of interest.

Acknowledgements
We gratefully thank the citizen scientists who collected samples:        Table 1 for Pearson's r and p-values corresponding to correlations for each site.