Revised Manuscript Em05d-100 Traditional Cattle Manure Application Determines Abundance, Diversity and Activity of Methanogenic Archaea in Arable European Soil Running Title: Methanogenic Soil Archaea 21 22 23

27 Based on lipid analyses, 16S rRNA/rRNA gene single strand conformation polymorphism 28 fingerprints and methane flux measurements, influences of the fertilisation regime on 29 abundance and diversity of archaeal communities were investigated in soil samples from the 30 long-term (103 years) field trial in Bad Lauchstädt, Germany. The investigated plots followed 31 a gradient of increasing fertilisation beginning at no fertilisation and ending at the 'cattle 32 manure' itself. The archaeal phospholipid etherlipid (PLEL) concentration was used as an 33 indicator for archaeal biomass and increased with the gradient of increasing fertilisation, 34 whereby the concentrations determined for organically fertilized soils were well above 35 previously reported values. Methane emission, although at a low level, were occasionally only 36 observed in organically fertilized soils, whereas the other treatments showed significant 37 methane uptake. Euryarchaeotal organisms were abundant in all investigated samples but 16S 38 rRNA analysis also demonstrated the presence of Crenarchaeota in fertilised soils. Lowest 39 molecular archaeal diversity was found in highest fertilized treatments. Archaea 40 phylogenetically most closely related to cultured methanogens were abundant in these 41 fertilized soils, whereas Archaea with low relatedness to cultured microorganisms dominated 42 in non-fertilized soils. Relatives of Methanoculleus spp. were found almost exclusively in 43 organically fertilised soils or cattle manure. Methanosarcina related microorganisms were 44 detected in all soils as well as in the cattle manure, but soils with highest organic application 45 rate were specifically dominated by a close phylogenetic relative of Methanosarcina 46 thermophila. Our findings suggest that regular application of cattle manure increased archaeal 47 biomass, but reduced archaeal diversity and selected for methanogenic Methanoculleus and 48 Methanosarcina strains, leading to the circumstance that high organic fertilised soils did not 49 function as a methane sink at the investigated site anymore.