Controls on brGDGT Production in the Seasonally Anoxic Water Column and Sediments of Rotsee (Lake Rot)

Reliable paleotemperature proxies are essential for reconstructing past climate. To refine interpretation of the MBT' 5ME index, based on bacterial brGDGT lipids, a year‐long study was conducted in Rotsee, Switzerland, a seasonally stratified lake with a 4–21°C temperature range. Suspended particulate matter was collected monthly from the epilimnion and the hypolimnion, complemented by surface sediments and surrounding soils. Both intact polar (IPL) and core lipid (CL) brGDGTs were analyzed alongside 16S rRNA gene data to disentangle environmental (temperature, dissolved oxygen, and pH) and biological (microbial community) controls on brGDGT compositions. In the stratified epilimnion, MBT' 5ME values showed a muted response to summer warming ( r = 0.59, p < 0.1), whereas the isomer ratio (IR) correlated more strongly with temperature ( r = 0.68, p < 0.05). MBT' 5ME and IR were also significantly correlated ( r = 0.93, p < 0.0001), providing a novel diagnostic tool to identify sedimentary GDGTs derived from surface waters. In the seasonally anoxic hypolimnion, MBT' 5ME correlated with pH ( r = 0.79, p < 0.01) and IR with dissolved oxygen ( r = −0.65 and p < 0.05). Microbial DNA analysis revealed low Acidobacterial abundances (<0.4% of reads), suggesting MBT' 5ME patterns are not solely driven by this phylum. Instead, hypolimnion IPL‐brGDGTs correlated with gene abundance of several other bacteria, indicating broader microbial contributions. Surface sediments reflected an integrated water column signal, while also showing evidence for additional in situ IPL‐brGDGT production. Overall, findings demonstrate that stratification onset drives MBT' 5ME variability, while epilimnion temperature exerts stronger control on IR, refining their application in paleoclimate proxies.