Novel Environmental Proxies Based On Bacterial 3-hydroxy Fatty Acids And Their Applications In Stalagmite

Global climate change has gained more and more concerns. People need to understand the climate changes of Earth history and predict the future climate change. Stalagmites have become a key terrestrial archive in palaeoclimatic reconstruction and gained lots of attention in the 21st century due to their ability to yield continuous and undisturbed records, precise and absolute chronologies and their global terrestrial distribution. Numbers of palaeoclimate proxies have been established and applied to speleothem archives by researchers from domestic and abroad. These proxies include: oxygen isotopes, carbon isotopes, clumped isotope, annual growth banding, trace elements, magnetic information, inclusions and biomarkers. Among these environmental proxies, oxygen isotopes are the most widely applied and are effectively the’master’or standard approach for stalagmite analysis. However, oxygen isotope records inherently encode a mix of climatic signals, including at the regional scale: temperature changes, isotopic composition of source waters, precipitation as well as complex site-specific factors. Thus interpretation of oxygen isotope speleothem records requires site-specific considerations and deconvolution of a complex interplay of factors.Abundance of biomarkers have been detected in stalagmite samples. Although the relatively complex sources of biomarkers in stalagmites, some compounds have simple or single source, especially lipids from microbial membrane. For example, the 3-hydroxy fatty acids (3-OH-FAs) are very sensitive to environmental changes and well preserved in spelethems, thus have great potential in palaeoclimate reconstruction.3-OH-FAs are characteristic compounds of Lipid A, the lipid component of the lipopolysaccharides (LPS) located in the outer membrane of Gram-negative bacteria. 3-OH-FAs are linked to large molecules via amide bonds or ester bonds, thus they can not be easily degraded and could be well preserved in sediments. Abundance of 3-OH-FAs have been found in speleothem in the previous research, but the application of 3-OH-FAs in palaeoclimate reconstruction is rare. Here we aim to extract 3-OH-FAs from soil and stalagmite samples based on an optimized method, to establish new environmental proxies based on 3-OH-FAs and apply them into stalagmites. The main results are shown as following:1. Novel temperature and pH proxies based on 3-OH-FAs from soilsEstablished novel pH and temperature proxies based on 3-OH-Fas from soil samples.3-OH-FAs in surface soils collected from an altitudinal transect on Mt. Shennongjia were examined to explore their relationships with environmental parameters. The ratio of the total sum of iso and anteiso 3-OH-FAs to the total amount of normal 3-OH-FAs (RIAN), the ratio of summed branched homologues to the sum of all 3-OH-FA homologues (Branched Index) and the ratio of summed wo to summed normal 3-OH-FA homologues (REST) are highly correlated with soil pH. Furthermore, the RAN15 and RAN15 ratios exhibit significant correlations with MAAT and MAP. As precipitation is not likely to be an ecologically limiting factor in the moist-humid environment of Mt. Shennongjia, we assume that MAAT is the dominant control. Notably, the 3-OH-FA based temperature proxies RAN15 and RAN17, are not pH dependent, which should be an advantage in environments where pH is highly variable and could be a confounding variable. Our discovery of new independent proxies for pH and MAAT from an altitudinal transect of surface soils from Mt. Shennongjia has profound significance for palaeoclimatic and environmental studies. Multi-proxy terrestrial reconstructions of pH and temperature could be established by comparing 3-OH-FAs with GDGT based proxies.At the meantime, detectable and abundance 3-OH-FAs were found in Tianchi Lake sediment, marine sediment from Baltic Sea, stalagmite sample from Heshang Cave, which shows potential wide application of 3-OH-FAs proxies. The variations in the 3-OH-FA signatures between the different settings are likely due to controls by environmental and climatic parameters on membrane lipid production by bacteria. Moreover, monounsaturated 3-OH-FAs with even carbon numbers (C12, C14,C16,C18) were uniquely found in the Tianchi Lake sediment, suggesting a unique source of 3-OH-FAs in that lake environment, thus specific calibrations are likely required for applications to a diverse range of palaeoclimatic archives.2. Palaeotemperature and palaeohydrology reconstruction based on 3-OH-FAs proxiesThe temperature in the middle reaches of Yangtze River over the last 9 ka BP was reconstructed for the first time by appling soil based temperature proxy RAN15 in HS4 stalagmite samples. The average distributions of hydroxy fatty acids (OH-FAs) in the overlying soils, cave surface sediments and the HS4 stalagmite samples are investigated. The overall similarity in the distribution patterns of the three sample sets is notable. However, we note the distinctive features of the 3-OH-FA distribution in the HS4 samples, specifically the greater relative abundance of the n-C12 and i-C17 3-OH-FAs compared to both overlying soils and cave sediments. This suggests an additional contribution of 3-OH-FAs, derived from niche micro-organisms dwelling on the surface of the actively-forming stalagmite, which are sequestered and trapped in the calcite matrix.Temperature changes in the middle reaches of Yangtze River during the last 9 kyr BP was reconstructed by the ratio of a-Cis to n-C15 3-OH-FAs (RAN15) from HS4 stalagmite. The reconstructed temperature varied from 17.8 to 20.2℃ during the Holocene with a relative warm period in the early to middle Holocene (8.6-5.2 ka BP), and then since 5.2 ka BP, a trend towards cooler conditions. The reconstructed temperature is very close to the modern annual temperature outside the cave (18℃).The hydrological condition in the middle reaches of Yangtze River over the last 9 ka BP was reconstructed by appling C15 hydro index in HS4 stalagmite samples. Because the small variation range of pH in cave sediments, the novel pH proxies based on Shennongjia Mountain soils is not suitable for cave system. We find an empirical relationship between the ratio of i-C15 to a-C15 3-OH-FA (C15 hydro index) in the overlying soils and cave surface sediments and pH (R2= 0.65, p<0.001). Based on the semi quantitative relationship between C15 Hydro index and hydrological conditions from our cave investigation, we gained a new precipitation record during the last 9 ka BP by applying the new precipitation proxy in HS4 stalagmite samples. The precipitation record inferred from C15 Hydro index reveals two relatively long wet periods and one relative dry period in central China,8.1-6.0 ka BP,3.5-0 ka BP and 6.0-3.5 ka BP respectively, superimposed on this general pattern are several pronounced dry events around 8.2,3.8 and 0.5 ka BP.3. Palaeovegetation reconstruction inferred from the carbon isotope of fatty acids from stalagmiteThe palaeovegetation in the middle reaches of Yangtze River over the last 9 ka BP was reconstructed by the analysis of compound specific carbon isotope of fatty acids from HS4 stalagmite. The overall distribution pattern of fatty acid in cave overlying soil, cave surface sediment and HS4 stalagmite is similar. The carbon number of fatty acids ranges from C12 to C32, showing significance even/odd predominance and maximizing at Ci6. The source of short chain fatty acids in stalagmite is complex. The high plants, soil microbial and cave microbia all contribute short chain fatty acids to stalagmite.The carbon isotope of C16 fatty acid (δ13C16 FA) varied from -27.5%o to -25.2%o, the carbon isotope of C18 fatty acid (δ13C18 FA) varied from -27.0%o to -23.4%o. The same variation trend between δ13C16FA and δ13C18FA indicates both of the fatty acids sharing the same sources. The carbon isotope variation of the long chain fatty acids (δ13C22 FA and δ13C24 FA) shows larger variation amplitude than the short chain fatty acids (δ13C16FA and δ13C18FA), with δ13C22FA varing from -32.9%o to -25.7%o and δ13C24FA varing from-33.5‰ to -25.2‰。According to the δ13C24 FA from HS4 stalagmite and the average carbon isotope value of C24 fatty acid from C3 and C4 plants, palaeovegetation was reconstructed in the middle reaches of Yangtze River over the last 9 ka BP. The variation of palaeovegetation was divided into three stages. Stage 1, increase of C3 plants and decrease of C4 plants between 8.6-6.8 ka BP. Stage 2, decrease of C3 plants and increase of C4 plants between 6.8-3.3 ka BP. Stage 3, cyclic percentage variation of C3 and C4 plants between 3.3-0 ka BP. The variation of C3 and C4 plants is mainly affected by temperature, as well as the hydrological condition.In summary, novel temperature and pH proxies based on 3-OH-FAs were established, and be well applicated into speleothem samples. We revealed the temperature and hydrological changes over the last 9 ka BP in the middle reaches of Yangtze River. The specific carbon isotope of fatty acid from stalagmite were analized for the first time and palaeovegetation was reconstructed in the middle reaches of Yangtze River over the last 9 ka BP based on the δ13C24 FA record from stalagmite. We hope the work opens up new avenues of research on 3-OH-FAs and compound specific isotope analysis of lipids from stalagmite.