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Supplementary information 1

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Table S1: Seed samples used in this study 3

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Scientific name ID Common name Taxonomy ID Genotype Provider Sample Production Region

Harvest Year TSW (g)

H0 (number of seeds)

H24 (number of germinating seeds)

H96 (number of seedlings)

PBS Tween 20 (0.05%) Conditions of incubation

Brassica oleracea var. capitata S01 cabbage 3716 Guardian HM-Clause C50634 Chile 2004 6.68 924 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica oleracea var. capitata S02 cabbage 3716 Guardian HM-Clause C32216 France 2010 4.83 1289 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica oleracea var. capitata S03 cabbage 3716 Guardian HM-Clause D48989 Chile 2012 4.59 1427 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica oleracea var. capitata S04 cabbage 3716 Count HM-Clause C39271 France 2010 4.11 1563 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm



Brassica oleracea var. capitata S07 cabbage 3716 Negus HM-Clause C41073 France 2010 6.57 975 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica oleracea var. capitata S08 cabbage 3716 Negus HM-Clause C43102 Italia 2010 4.85 1282 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica oleracea var. capitata S09 cabbage 3716 Negus HM-Clause D22352 Chile 2011 7.41 885 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica oleracea var. botrytis S10 cauliflower 3715 Clovis Vilmorin K31987 France 2012 5.47 1147 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica oleracea var. italica S11 broccoli 36774 Verdia Vilmorin J93783 Japan 2011 4.44 1050 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica rapa subsp. rapa S12 turnip 51350 Armand HM-Clause D56281 France 2012 2.86 1094 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica rapa subsp. rapa S13 turnip 51350 Plessis HM-Clause D45368 Chile 2012 1.72 1790 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica rapa subsp. rapa S14 turnip 51350 Declic HM-Clause D44795 Chile 2012 2.82 887 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica rapa subsp. rapa S15 turnip 51350 Clovis HM-Clause D59599 France 2012 1.67 1750 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica napus var. napus S16 canola 138011 Darmor Bzh IGEPP

France 2011 3.92 1105 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Brassica napus var. napus S17 canola 138011 Yudal IGEPP


Raphanus sativus var. sativus S18 radish 240228 Flamboyant5 Vilmorin


Diplotaxis tenuifolia S19 garden rocket 264416 Nature Vilmorin K45457 France 2012 0.37 1721 200 100 25 ml / 1000 seeds 2h30 RT, 140 rpm

Arabidospis thaliana S20 thale-cress 3702 SF2 IRHS


Medicago truncatula S21 barrel clover 3880 A17 IRHS

France 2012 4.14 1030 200 100 25 ml / 1000 seeds o/n 4°C, 140 rpm

Phaseolus vulgaris S22 bean 419294 Flavert Psp Vilmorin K22374 France 2012 178.4 982 200 100 445 ml / 1000 seeds o/n 4°C, 140 rpm

Phaseolus vulgaris S23 bean 419294 Flavert Vilmorin K36604 France 2012 191.4 961 200 100 477 ml / 1000 seeds o/n 4°C, 140 rpm

Phaseolus vulgaris S24 bean 419294 Flavert Vilmorin K38580 Chili 2013 172.5 1003 200 100 445 ml / 1000 seeds o/n 4°C, 140 rpm

Daucus carota S25 carrot 79200 Presto Vilmorin J89925 New Zealand 2012 1.85 1488 200 100 10 ml/ 1000 seeds o/n 4°C, 140 rpm

Daucus carota S26 carrot 79200 Maestro Vilmorin K01057 USA 2012 1.87 1685 200 100 10 ml / 1000 seeds o/n 4°C, 140 rpm

Solanum lycopersicum S27 tomato 4081 Retana Vilmorin K10554 China 2011 1.88 1007 200 100 10 ml / 1000 seeds o/n 4°C, 140 rpm

Solanum lycopersicum S28 tomato 4081 Superpro Vilmorin K35846 China 2012 1.13 1044 200 100 10 ml / 1000 seeds o/n 4°C, 140 rpm

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Table S2: Summary of the different sequencing runs performed in this study 6

molecular markers runs index pairs of reads quality sequences

OTUs aOTUs (> 0.1% of sequences per sample)

Sequences belonging to aOTUs

16S rRNA gene (V4 region) 1 single 3,683,321 2,084,455 11,811 1,017

2,023,266

16S rRNA gene (V4 region) 2 single 8,771,960 4,977,994 4,838,019

ITS1 1 single 2,825,933 2,658,269 7,421 448

2,539,261

ITS1 2 single 6,814,684 6,052,730 5,756,919

gyrB 3 dual 3,275,542 2,159,392 27,860 1,940 2,039,520

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The table summarizes information regarding the 3 three independent sequencing runs performed with the Illumina MiSeq v. 2.0 platform (250 bp paired-8

end reads). The number of pair of reads, quality sequences, operational taxonomic units (OTUs), aOTUs and sequences belonging to aOTUs are indicated in 9

each column. 10

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Table S3: Bacterial strains used to construct the artificial bacterial community sample. 12

CFBP Species Type strain Genome sequenced Family 16S copies

1955 Bacillus pumilus N N Bacillaceae 6

4227 Bacillus licheniformis Y Y Bacillaceae 7

3031 Empedobacter brevis Y Y Flavobacteriaceae 1

1430 Erwinia amylovora Y Y Enterobacteriaceae 7

6723 Ochrobactrum anthropi Y Y Brucellaceae 4

6623 Agrobacterium sp. biovar 1 N N Rhizobiaceae 4

6715 Mesorhizobium loti Y Y Phyllobacteriaceae 2

2534 Xanthomonas axonopodis pv. phaseoli Y (pathotype) Y Xanthomonadaceae 2

5241 Xanthomonas campestris pv. campestris Y (pathotype) Y Xanthomonadaceae 2

7087 Pseudomonas savastanoi pv. phaseolicola N Y Pseudomonadaceae 5

2049 Clavibacter michiganensis subsp. sepedonicus Y Y Microbacteriaceae 2

2425 Acidovorax avenae subsp. avenae Y Y Comamonadaceae 3

2227 Burkholderia cepacia Y Y Burkholderiaceae 6

2401 Rhodococcus fascians Y N Nocardiaceae 5

4223 Cellulomonas flavigena Y Y Cellulomonadaceae 2

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An artificial community sample was prepared by mixing equal amount of genomic DNA of 15 bacterial strains. Bacterial strains were provided from 14

Collection For Plant-associated Bacteria (CIRM-CFBP, IRHS, 42 rue Georges Morel, 49070 Beaucouzé Cedex, France). Accession numbers are indicated in the 15

first column, while the number of 16S rRNA gene per genome is indicated on the last column. 16

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Table S4: Factors influencing the structure of the microbial community associated to seeds, germinating seeds and seedlings. 18

Factors 16S aOTUs (97%) gyrB aOTUs (98%) ITS1 aOTUs (97%)

Fs p-value Fs p-value Fs p-value

2004-2010 1.46536 0.188 0.537712 0.895 3.32829 0.002

2004-2011 1.82538 0.072 1.13381 0.314 2.19849 0.024

2004-2012 0.810669 0.583 0.592975 0.952 2.10447 0.024

2004-2013 3.11742 0.073 0.740876 0.666 7.41677 ≤0.001

2010-2011 5.1418 ≤0.001 3.45549 ≤0.001 5.24959 ≤0.001

2010-2012 2.23036 0.036 1.74296 0.034 6.94314 ≤0.001

2010-2013 3.57399 0.005 1.20973 0.277 5.77593 ≤0.001

2011-2012 2.40716 0.013 1.5319 0.065 2.02932 0.022

2011-2013 4.71099 ≤0.001 2.11737 0.023 4.97353 ≤0.001

2012-2013 2.91625 0.009 1.31244 0.166 3.56567 0.002

Chile-France 6.27247 ≤0.001 1.82256 0.045 4.99039 ≤0.001

H0-H24 5.50175 ≤0.001 1.23593 0.221 4.74451 ≤0.001

H0-H96 13.5318 ≤0.001 2.28088 0.008 8.64584 ≤0.001

H24-H96 7.93636 ≤0.001 1.63601 0.047 7.82317 ≤0.001

Brassicaceae vs other 1.68907 0.090 1.40559 0.112 7.69972 ≤0.001

Brassica vs other 2.0475 0.029 1.19421 0.244 7.25763 ≤0.001

B. oleracea vs other 2.28002 0.030 1.37505 0.141 5.35931 ≤0.001

B.oleracea var. capitata vs other 3.22188 0.002 1.59455 0.083 6.54833 ≤0.001

Technical replicate 0.0287207 1 na na 0.0156882 1

Seed, germinating seed and seedling samples were clustered according to the structure of their microbial community (as assessed by Bray-Curtis 19

dissimilarities matrix of 16S rRNA gene, gyrB and ITS sequences). The influence of harvesting year, production region, physiological stage, family, genus, 20

species and varieties on the structure of the bacterial (16S rRNA gene and gyrB sequences) and fungal communities (ITS1) was assessed by analysis of 21

molecular variance (AMOVA). Fs values and p-values are reported for each molecular marker. Factors are considered as significant when p-value ≤ 0.001 22

(highlighted in bold). 23

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Table S5: Microbial core community associated to seed samples 24

core aOTUs Taxonomy All seeds (28) Brassicaceae (20) Brassica (17) B.oleracea (11) B.oleracea capitata (9)

16S rRNA gene

Otu00001 Pseudomonas (100) 100 100 100 100 100

Otu00002 Pantoea (84) 100 100 100 100 100

Otu00007 Xanthomonas (87) 100 100 100 100 100

Otu00013 Sphingomonas (100) 93 100 100 100 100

Otu00020 Sphingomonas (99) 89 100 100 100 100

Otu00003 Pseudomonas (100) 82 100 100 100 100

gyrB

Otu00001 Pantoea agglomerans (100) 100 100 100 100 100

ITS1 Otu1047 Mycosphaerellaceae (100) 100 100 100 100 100

Otu1020 Cladosporium (99) 96 100 100 100 100

Otu0379 Alternaria (100) 96 95 94 91 100

Otu1343 Alternaria infectoria 95 95 94 91 100

Otu1273 Cryptococcus (98) 88 90 94 91 100

Otu0361 Pleosporaceae (100) 86 88 91 91 100

Otu0267 Filobasidium (96) 82 90 94 91 100

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Number of aOTUs systematically associated to all seed samples, seeds related to Brassicaceae, Brassica, B. oleracea and B. oleracea var. capitata. Number of 26

samples associated to each class is indicated in parentheses. Frequency (expressed as percentage) of aOTUs are shown in each column. 27

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Table S6: List of aOTUs significantly enriched during emergence. 28

aOTU Taxonomy

logF

C (

H9

6 v

s H

0)

FDR

(H

96

vs

H0

)

16S rRNA gene

Otu00060 Bacillales (100) 2,22 1,5E-08

Otu00005 Paenibacillus (100) 5,08 4,4E-17



Otu00557 Oxalobacteraceae (100) 2,44 1,9E-09

Otu00018 Duganella (87) 2,29 7,2E-04

Otu00050 Gammaproteobacteria (100) 3,61 2,5E-15

Otu00048 Enterobacteriaceae (100) 4,13 2,1E-16

Otu00068 Enterobacteriaceae (100) 2,04 1,5E-07

Otu00012 Pseudomonas (99) 4,16 2,9E-07




gyrB

Otu01019 Bacillaceae (100) 2,4 2,E-04

Otu00531 Bacillus amyloliquefaciens (100) 2,8 1,E-05

Otu02826 Bacillus mojavensis (100) 3,1 1,E-05

Otu00141 Bacillus pumilus (100) 2,9 6,E-04

Otu01110 Paenibacillus (100) 3,0 2,E-05

Otu00066 Paenibacillus (82) 3,3 2,E-06

Otu00456 Paenibacillus polymyxa (92) 6,1 5,E-10

Otu27596 Actinomycetales (91) 2,2 8,E-04

Otu04503 Sphingomonadaceae (100) 2,6 1,E-04

Otu00573 Sphingomonas (100) 4,2 2,E-07

Otu01462 Lautropia mirabilis (100) 2,3 4,E-04

Otu01084 Oxalobacteraceae (100) 2,5 1,E-04

Otu00082 Duganella zoogloeoides (99) 3,7 4,E-05

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Otu00739 Massilia (100) 3,4 4,E-06

Otu00176 Massilia (99) 6,2 8,E-10

Otu00039 Massilia alkalitolerans (100) 5,3 4,E-09



Otu00261 Massilia timonae (100) 3,9 6,E-07




Otu00118 Epsilonproteobacteria (100) 2,1 6,E-04

Otu00165 Epsilonproteobacteria (94) 7,2 5,E-12

Otu00081 Enterobacteriaceae (100) 7,6 3,E-11

Otu00037 Pantoea (100) 6,7 1,E-09

Otu00040 Pantoea (100) 7,5 2,E-10

Otu00045 Pantoea (100) 5,8 1,E-07

Otu00173 Pantoea agglomerans (100) 2,3 2,E-04

Otu00559 Acinetobacter calcoaceticus (100) 3,6 1,E-06

Otu00555 Acinetobacter pittii (100) 2,3 5,E-04

Otu00018 Pseudomonas (100) 5,3 1,E-05










Otu00048 Pseudomonas fluorescens (100) 7,5 2,E-11



Otu00149 Pseudomonas oleovorans (100) 4,3 4,E-07

Otu00296 Pseudomonas oleovorans (94) 2,5 4,E-04

Otu00114 Pseudomonas viridiflava (100) 5,8 1,E-09

ITS1

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Otu1342 Ascomycota(100) 2,71 1,7E-04

Otu0748 Alternaria (100) 2,73 1,8E-04

Otu1462 Alternaria (83) 3,34 3,4E-13

Otu2419 Alternaria (85) 2,78 4,5E-11

Otu1198 Alternaria (98) 2,26 1,7E-09

Otu4462 Alternaria chartarum (85) 2,47 4,9E-05

Otu1272 Cladosporium cucumerinum (95) 2,47 4,2E-10

Otu1130 Chaetomiaceae (100) 3,29 5,0E-13

Otu0767 Chaetomiaceae (80) 2,62 1,8E-10

Otu1110 Chaetomium globosum (93) 6,04 2,4E-20

Otu1222 Clonostachys (99) 3,75 1,5E-14

Otu0199 Wickerhamomyces (100) 2,46 7,8E-10

Otu0146 Basidiomycota (84) 2,10 2,2E-08

Otu0544 Basidiomycota(80) 2,47 4,6E-08



Otu5074 Ceratobasidiaceae (85) 2,84 2,6E-11

Otu0706 Cryptococcus (91) 2,93 7,9E-06



Otu0720 Daedaleopsis confragosa (100) 6,05 2,4E-20

Otu0737 Debaryomyces hansenii(100) 4,21 1,3E-09

Otu0191 Filobasidiaceae (84) 2,87 1,9E-11

Otu1175 Filobasidiaceae (98) 3,55 1,7E-14

Otu0267 Filobasidium (96) 2,73 4,9E-06

Otu0661 Fuscoporia ferrea (100) 2,92 1,3E-11

Otu0435 Hypocreales (82) 5,83 7,3E-20

Otu0314 Leothiomycetes (100) 5,51 3,7E-21

Otu0229 Leptospora rubella (100) 4,10 1,7E-15

Otu2057 Meruliaceae (86) 4,71 3,5E-17

Otu0342 Penicillium (100) 4,02 1,3E-16



Otu0599 Penicillium brevicompactum (100) 2,27 2,4E-05

Otu0594 Penicillium commune (96) 5,33 6,4E-11

Otu0775 Peniophora piceae (93) 6,71 2,3E-15

Otu1369 Pleosporaceae (100) 4,13 1,3E-15

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Otu0860 Saccharomycetales (94) 2,31 3,3E-09

Otu0197 Trametes (100) 4,01 1,0E-16

Otu1093 Trechisporales (87) 3,31 4,4E-13

Otu0143 Tremellales (80) 3,35 8,4E-14

Otu1435 Tremellomycetes (80) 2,47 1,5E-04



Otu1033 Trichocomaceae (98) 2,57 2,9E-05

Otu0190 Trichoderma viride (94) 7,45 9,0E-24

Otu0610 Rhizopus oryzae (99) 5,69 1,6E-21

Otu0193 Unclassified 2,49 6,2E-10

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Differences in relative abundance of aOTUs between samples were assessed with the R package edgeR (1) using exact binomial tests corrected for multiples 30

inferences with the Benjamini-Hochberg method (2). aOTUs were defined as significantly enriched at a corrected pvalue < 0.001 and a log2 fold change ≥ 2. 31

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Figure S1: Influence of biotic and abiotic factors on bacterial and fungal richness 33

Number of aOTUs (y axis) observed in seed samples (H0). Each sample is represented by a green line, while the grey area represented the distribution. 34

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Figure S2: Comparison of gyrB genetic distance to gANI 36

Pairwise distances between gyrB regions were computed and sequences were grouped at different genetic 37

distance (from 0.01 to 0.10). These gyrB groups were then compared to cliques obtained with whole-genome 38

based average nucleotide identity (gANI) values available at http://ani.jgi-psf.org. These gANI cliques can be 39

used as a proxy for species delineation (N. Varghese, S. Mukherjee, N. Ivanova, K. Konstantinidis, K. 40

Mavrommatis, N. Kyrpides and A. Pati, submitted for publication). Sensitivity, precision and F1 score were 41

calculated at each genetic distance. 42

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References: 45

1. Robinson MD, McCarthy DJ, Smyth GK. 2010. edgeR: a Bioconductor package for differential 46 expression analysis of digital gene expression data. Bioinformatics 26:139-140. 47

2. Benjamini Y, Hochberg Y. 1995. Controlling the false discovery rate: A practical and powerful 48 approach to multiple testing. J R Stati Soc Series B 57:289-300. 49

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http://ani.jgi-psf.org/

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