The degree of methylation and cyclization of bacteria-derived branched glycerol dialkyl glycerol tetraether (GDGT) membrane lipids in soils depends on temperature and soil pH. Expressed in the methylation index of branched tetraethers (MBT) and cyclization ratio of branched tetraethers (CBT), these relationships are used to reconstruct past annual mean air temperature (MAT) based on the distribution of branched GDGTs in ancient sediments; the MBT-CBT proxy. Although it was shown that the best correlation of this proxy is with annual MAT, it remains unknown whether a seasonal bias in temperature reconstructions could occur, such as towards a seasonal period of 'optimal growth' of the, as yet, unidentified soil bacteria which produce branched GDGTs. To investigate this possibility, soils were sampled from eight different plots in the USA (Minnesota and Ohio), The Netherlands (Texel) and the UK (Devon) in time series over 1 year and analyzed for their branched GDGT content. Further analyses of the branched GDGTs present as core lipids (CLs; the presumed fossil pool) and intact polar lipids (IPLs; the presumed extant pool) were undertaken for two of the investigated soil plots. The amount of IPL-derived branched GDGTs is low relative to the branched GDGT CLs, i.e. only 6-9% of the total branched GDGT pool. In all soils, no clear change was apparent in the distribution of branched GDGT lipids (either core or IPL-derived) with seasonal temperature change; the MBT-CBT temperature proxy gave similar temperature estimates year-round, which generally matched the mean annual soil temperature. In addition to a lack of coherent changes in relative distributions, concentrations of the branched GDGTs did not show clear changes over the seasons. For IPL-derived GDGTs these results suggest that their turnover time in soils is in the order of 1 year or more. Thus, our study does not provide evidence for seasonal effects on the distribution of branched GDGTs in soils, at least at mid-latitudes, and therefore, no direct evidence for a bias of MBT-CBT reconstructed temperatures towards a certain season of optimal growth of the source bacteria. If, however, there is a slight seasonal preference of branched GDGT production, which can easily be obscured by natural variability due to the heterogeneity of soils, then a seasonal bias may potentially still develop over time due to the long turnover time of branched GDGTs.

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