Supplementary Figure S1

Transcription

1 Supplementary Figure S1 KRO STO R-p R-p3 RUD R-nt L-n L-h L-l ph <6 6-7 >7 WEON [mg/kg dw soil] <1 1-3 >3 WEOC [mg/kg dw soil] <1 1-3 >3 Soil type sandy sandy loam silty clay GSF B77V B77T E16 Figure S1: (Geo)Graphical representation of soil samples used in this study, including some of their chemical characteristics as given in Table S1.

2 Supplementary Figure S a 9, 7, AOB amoa copies reaction -1 8, 7, 6,, 4, 3,, 1, RUD STO AOA amoa copies reaction -1 6,, 4, 3,, 1, RUD STO bead beating time for cell lysis [s] bead beating time for cell lysis [s] b dilution series of N. multiformis amoa standard +KRO or RUD DNA dilution series of fosmid 4d9 + RUD DNA C(t) DNA + RUD DNA + KRO DNA Log quantity C(t) DNA + RUD DNA Log quantity Figure S: Controls for amoa qpcr. a: influence of different bead beating times during DNA cell lysis on AOB and AOA amoa copy numbers in two different soils using the Griffiths method 6. DNA from RUD (light grey) yielded most AOB amoa copies at 1 s beat beating time while copy numbers of STO derived DNA (dark grey) were highest at 18 s (left graph). Different results were obtained for AOA amoa, for which highest values were measured at 3 s for RUD, while STO revealed constant copy numbers over treatment time (right graph). b: influence of inhibitory substances on amoa copies in the two soils that were most heavily contaminated by humic acids. Ten ng (4 µl) of soil DNA preparation from KRO or RUD, respectively, was added to diluted standard DNA, in this case from N. multiformis. While no inhibition was observed for sample KRO, some inhibition was observed in heavily contaminated soil RUD, reducing the determined amoa copy number by up to %, (between log and log 8 C(t)-values are slightly higher where soil DNA was added to standard). Inhibition tests on AOA amoa qpcr are shown with RUD soil (light grey) and fosmid 4d9 (dark grey) as a standard dilution series. C(t)-values indicated similar inhibition as in AOB amoa qpcr. C(t) = cycle threshold in qpcr, log quantity = log of template copy numbers.

3 Supplementary Figure S3 Soil amoa Lawn GRASS3 Pasture KRO1 Sandy soil Pasture KRO Sandy soil RUD cdna RUD1 cdna 3 OKR-C-16 Lawn GRASS1.1 OKR-C-14 9/9/9 7/37/83 99/1/1 Sargasso Sea AACY17168 OKR-C- 13 Austrian Alps ALPINE3 Austrian Alps ALPINE 1 cm 6 7 cm 4 8/6/1 Both 1 & 6 7 cm 78/68/ /7/9 OKR-C /1/1 Nitrosopumilus maritimus Sargasso Sea AACY Marine amoa Figure S3: Phylogeny of 7 derived crenarchaeal amoa sequences recovered from two distinct depths ( to 1 cm and 6 to 7 cm) of a soil profile sampled at Rotböll, Germany. Sequences from other surveys included those from Rotböll, Germany 8 (RUD: accession numbers DQ3488-DQ34814); Alpine soil, Austria (R: DQ DQ34699); Public lawn, Scotland (GRASS: DQ347, DQ3474); Pasture, Norway (KRO: DQ3481, DQ34816), Tennessee soil, USA (OKR: DQ14887, DQ148876, DQ and DQ14888), Sargasso Sea 1 (AACY17168 and AACY143967) and Nitrosopumilus maritimus 11 (DQ898). Values at tips indicate the number of identical sequences represented by the individual branch [orange: all from -1 cm; blue: all from 6-7 cm; green: from both -1 and 6-7 cm (and +/- from other surveys); black: from other surveys only]. Maximum likelihood distances were calculated from 169 aligned amino acid positions and values at major nodes represent bootstrap support from protein distance/protein maximum-parsimony/dna distance analyses (expressed as a percentage of 1 replicates). LogDet DNA distances were calculated from an initial alignment of 7 nucleotides after all 3rd codon positions were excluded. Scale bar indicates.1 changes per site.

4 Supplementary Figure S4 16S amoa Archaea Arch F/98R 19F/643R AOB CTO 189F/64R A189/R 1F*/R 1 = 1 ng, = 1 ng, 3 = 1 pg, 4 = 1 pg, = 1 pg, 6 = 1 fg, 7 =1 fg DNA Figure S4: Typical results of an MPN PCR experiment with different dilutions of DNA extracted from the L-h top soil. Of the five different primer pairs used, two targeted 16S rrna genes and three targeted amoa genes (indicated bottom right of each panel).

5 Supplementary Figure S Crenarchaeol (µg g -1 soil) R = amoa (copies 1 7 g -1 soil) Figure S: Bivariance plot of crenarchaeol abundance (µg g -1 dry soil) versus AOA amoa copies (1 7 g -1 dry soil). The high coefficient of correlation (R =.88; n = 8) indicates a common origin of crenarchaeol from ammonia oxidizers. GDGT and crenarchaeol concentrations determined for samples from managed grassland KRO and STO were excluded, due to deviations in archaeal lipid composition, which was strongly dominated by non-isoprenoidal GDGT (see chromatogram in Fig. b). In managed grasslands caldarchaeol comprised >7% and crenarchaeol <1% of isoprenoidal GDGT, whereas in the pristine grasslands and arable soils compositions were <6% caldarchaeol and >3% crenarchaeol (see also Supplementary Table S1).

6 Supplementary Figure S6 Caldarchaeol GDGT 4 GDGT 1 GDGT GDGT GDGT 6 GDGT 3 GDGT 7 Crenarchaeol Figure S6: Structures of glyceroldialkylglycerol tetraethers encountered in grassland and arable soil. Names and numbers refer to labels in chromatograms (Fig. ).