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2 3 Table S1: Seed samples used in this study 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 ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S02 cabbage 3716 Guardian HM-Clause C32216 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S03 cabbage 3716 Guardian HM-Clause D48989 Chile ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S04 cabbage 3716 Count HM-Clause C39271 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S05 cabbage 3716 Count HM-Clause C12471 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S06 cabbage 3716 Count HM-Clause C16799 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S07 cabbage 3716 Negus HM-Clause C41073 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S08 cabbage 3716 Negus HM-Clause C43102 Italia ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. capitata S09 cabbage 3716 Negus HM-Clause D22352 Chile ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. botrytis S10 cauliflower 3715 Clovis Vilmorin K31987 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica oleracea var. italica S11 broccoli Verdia Vilmorin J93783 Japan ml / 1000 seeds 2h30 RT, 140 rpm Brassica rapa subsp. rapa S12 turnip Armand HM-Clause D56281 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica rapa subsp. rapa S13 turnip Plessis HM-Clause D45368 Chile ml / 1000 seeds 2h30 RT, 140 rpm Brassica rapa subsp. rapa S14 turnip Declic HM-Clause D44795 Chile ml / 1000 seeds 2h30 RT, 140 rpm Brassica rapa subsp. rapa S15 turnip Clovis HM-Clause D59599 France ml / 1000 seeds 2h30 RT, 140 rpm Brassica napus var. napus S16 canola Darmor Bzh IGEPP France ml / 1000 seeds 2h30 RT, 140 rpm Brassica napus var. napus S17 canola Yudal IGEPP France ml / 1000 seeds 2h30 RT, 140 rpm Raphanus sativus var. sativus S18 radish Flamboyant5 Vilmorin France ml / 1000 seeds 2h30 RT, 140 rpm Diplotaxis tenuifolia S19 garden rocket Nature Vilmorin K45457 France ml / 1000 seeds 2h30 RT, 140 rpm Arabidospis thaliana S20 thale-cress 3702 SF2 IRHS France ml / 1000 seeds 2h30 RT, 140 rpm Medicago truncatula S21 barrel clover 3880 A17 IRHS France ml / 1000 seeds o/n 4 C, 140 rpm Phaseolus vulgaris S22 bean Flavert Psp Vilmorin K22374 France ml / 1000 seeds o/n 4 C, 140 rpm Phaseolus vulgaris S23 bean Flavert Vilmorin K36604 France ml / 1000 seeds o/n 4 C, 140 rpm Phaseolus vulgaris S24 bean Flavert Vilmorin K38580 Chili ml / 1000 seeds o/n 4 C, 140 rpm Daucus carota S25 carrot Presto Vilmorin J89925 New Zealand ml/ 1000 seeds o/n 4 C, 140 rpm Daucus carota S26 carrot Maestro Vilmorin K01057 USA ml / 1000 seeds o/n 4 C, 140 rpm Solanum lycopersicum S27 tomato 4081 Retana Vilmorin K10554 China ml / 1000 seeds o/n 4 C, 140 rpm 4 5 Solanum lycopersicum S28 tomato 4081 Superpro Vilmorin K35846 China ml / 1000 seeds o/n 4 C, 140 rpm 2

3 6 Table S2: Summary of the different sequencing runs performed in this study 7 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 2,023,266 11,811 1,017 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 2,539,261 7, 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, The table summarizes information regarding the 3 three independent sequencing runs performed with the Illumina MiSeq v. 2.0 platform (250 bp paired- end reads). The number of pair of reads, quality sequences, operational taxonomic units (OTUs), aotus and sequences belonging to aotus are indicated in each column. 11 3

4 12 Table S3: Bacterial strains used to construct the artificial bacterial community sample. CFBP Species Type strain Genome sequenced Family 16S copies Bacillus pumilus N N Bacillaceae Bacillus licheniformis Y Y Bacillaceae Empedobacter brevis Y Y Flavobacteriaceae Erwinia amylovora Y Y Enterobacteriaceae Ochrobactrum anthropi Y Y Brucellaceae Agrobacterium sp. biovar 1 N N Rhizobiaceae Mesorhizobium loti Y Y Phyllobacteriaceae Xanthomonas axonopodis pv. phaseoli Y (pathotype) Y Xanthomonadaceae Xanthomonas campestris pv. campestris Y (pathotype) Y Xanthomonadaceae Pseudomonas savastanoi pv. phaseolicola N Y Pseudomonadaceae Clavibacter michiganensis subsp. sepedonicus Y Y Microbacteriaceae Acidovorax avenae subsp. avenae Y Y Comamonadaceae Burkholderia cepacia Y Y Burkholderiaceae Rhodococcus fascians Y N Nocardiaceae Cellulomonas flavigena Y Y Cellulomonadaceae An artificial community sample was prepared by mixing equal amount of genomic DNA of 15 bacterial strains. Bacterial strains were provided from Collection For Plant-associated Bacteria (CIRM-CFBP, IRHS, 42 rue Georges Morel, Beaucouzé Cedex, France). Accession numbers are indicated in the first column, while the number of 16S rrna gene per genome is indicated on the last column. 17 4

5 18 Table S4: Factors influencing the structure of the microbial community associated to seeds, germinating seeds and seedlings Factors 16S aotus (97%) gyrb aotus (98%) ITS1 aotus (97%) Fs p-value Fs p-value Fs p-value Chile-France H0-H H0-H H24-H Brassicaceae vs other Brassica vs other B. oleracea vs other B.oleracea var. capitata vs other Technical replicate na na Seed, germinating seed and seedling samples were clustered according to the structure of their microbial community (as assessed by Bray-Curtis dissimilarities matrix of 16S rrna gene, gyrb and ITS sequences). The influence of harvesting year, production region, physiological stage, family, genus, species and varieties on the structure of the bacterial (16S rrna gene and gyrb sequences) and fungal communities (ITS1) was assessed by analysis of molecular variance (AMOVA). Fs values and p-values are reported for each molecular marker. Factors are considered as significant when p-value (highlighted in bold). 5

6 24 Table S5: Microbial core community associated to seed samples core aotus Taxonomy All seeds (28) Brassicaceae (20) Brassica (17) B.oleracea (11) 16S rrna gene Otu00001 Pseudomonas (100) Otu00002 Pantoea (84) Otu00007 Xanthomonas (87) Otu00013 Sphingomonas (100) Otu00020 Sphingomonas (99) Otu00003 Pseudomonas (100) gyrb Otu00001 Pantoea agglomerans (100) B.oleracea capitata (9) ITS1 Otu1047 Mycosphaerellaceae (100) Otu1020 Cladosporium (99) Otu0379 Alternaria (100) Otu1343 Alternaria infectoria Otu1273 Cryptococcus (98) Otu0361 Pleosporaceae (100) Otu0267 Filobasidium (96) Number of aotus systematically associated to all seed samples, seeds related to Brassicaceae, Brassica, B. oleracea and B. oleracea var. capitata. Number of samples associated to each class is indicated in parentheses. Frequency (expressed as percentage) of aotus are shown in each column. 6

7 logfc (H96 vs H0) FDR (H96 vs H0) 28 Table S6: List of aotus significantly enriched during emergence. aotu Taxonomy 16S rrna gene Otu00060 Bacillales (100) 2,22 1,5E-08 Otu00005 Paenibacillus (100) 5,08 4,4E-17 Otu00029 Paenibacillus (99) 7,79 5,7E-25 Otu00062 Paenibacillus (100) 2,94 3,4E-09 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 Otu00044 Pseudomonas (81) 2,30 6,8E-09 Otu00051 Pseudomonas (95) 3,94 1,4E-15 Otu00101 Pseudomonas (99) 2,23 1,4E-08 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 7

8 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 Otu00127 Massilia alkalitolerans (100) 5,2 5,E-09 Otu00212 Massilia alkalitolerans (100) 5,7 1,E-09 Otu00261 Massilia timonae (100) 3,9 6,E-07 Otu00898 Massilia timonae (100) 3,2 5,E-06 Otu00080 Massilia timonae (90) 5,2 7,E-09 Otu00235 Massilia timonae (95) 3,8 7,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 Otu00023 Pseudomonas (100) 3,8 5,E-04 Otu00046 Pseudomonas (100) 5,4 3,E-08 Otu00047 Pseudomonas (100) 3,8 8,E-05 Otu00061 Pseudomonas (100) 6,4 3,E-10 Otu00095 Pseudomonas (100) 2,5 2,E-04 Otu00101 Pseudomonas (83) 5,3 3,E-07 Otu00103 Pseudomonas (100) 3,3 5,E-06 Otu00137 Pseudomonas (100) 3,4 4,E-06 Otu00185 Pseudomonas (100) 4,1 3,E-07 Otu00048 Pseudomonas fluorescens (100) 7,5 2,E-11 Otu00164 Pseudomonas fluorescens (100) 3,3 5,E-06 Otu00342 Pseudomonas fluorescens (100) 3,0 2,E-05 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 8

9 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 Otu7237 Basidiomycota(87) 3,97 3,6E-15 Otu4974 Basidiomycota(99) 2,87 1,8E-07 Otu5074 Ceratobasidiaceae (85) 2,84 2,6E-11 Otu0706 Cryptococcus (91) 2,93 7,9E-06 Otu0474 Cryptococcus (98) 3,02 3,5E-05 Otu0685 Cryptococcus (99) 2,23 8,5E-04 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 Otu0962 Penicillium (100) 4,35 1,1E-07 Otu0512 Penicillium (99) 2,97 7,7E-12 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 9

10 Otu2343 Pleosporaceae (100) 2,16 1,2E-08 Otu2925 Pleosporaceae (100) 2,62 1,8E-10 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 Otu1457 Tremellomycetes (88) 4,74 2,9E-17 Otu1265 Tremellomycetes (93) 2,72 7,2E-11 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 Differences in relative abundance of aotus between samples were assessed with the R package edger (1) using exact binomial tests corrected for multiples inferences with the Benjamini-Hochberg method (2). aotus were defined as significantly enriched at a corrected pvalue < and a log 2 fold change 2. 10

11 Figure S1: Influence of biotic and abiotic factors on bacterial and fungal richness 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. 11

12 Figure S2: Comparison of gyrb genetic distance to gani Pairwise distances between gyrb regions were computed and sequences were grouped at different genetic distance (from 0.01 to 0.10). These gyrb groups were then compared to cliques obtained with whole-genome based average nucleotide identity (gani) values available at These gani cliques can be used as a proxy for species delineation (N. Varghese, S. Mukherjee, N. Ivanova, K. Konstantinidis, K. Mavrommatis, N. Kyrpides and A. Pati, submitted for publication). Sensitivity, precision and F1 score were calculated at each genetic distance References: 1. Robinson MD, McCarthy DJ, Smyth GK edger: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics 26: Benjamini Y, Hochberg Y Controlling the false discovery rate: A practical and powerful approach to multiple testing. J R Stati Soc Series B 57:

Mem. Faculty. B. O. S. T. Kindai University No. 38 : 1 10 (2016)

Mem. Faculty. B. O. S. T. Kindai University No. 38 : 1 10 (2016) 1 2 Memoirs of The Faculty of B. O. S. T. of Kindai University No. 38 2016 In recent years, several papers were published on microflora