Supplementary Material for “Fungi Are More Dispersal-Limited Than Bacteria 2 Among Flowers” 3
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1 Supplementary Material for “Fungi are more dispersal-limited than bacteria 2 among flowers” 3 4 5 6 7 Supplemental Figure S1 8 Thrips (Frankliniella occidentalis) were first fed a diet containing the antibacterial 9 chloramphenicol. Thrips were sterilized in batches through repeated washings inside 10 pipette chambers. Washing consisted of vortexing at maximum rpm and plunging out 11 solution with ethanol, water, bleach, and water three more times. The assay layout used 12 included 1/3 of wells beginning with microbes and waiting 24 hours for thrips to disperse 13 microbes. 14 15 16 17 18 Supplementary Figure S2—Relationship between cell number in suspension (plated) 19 and CFU detection on a) Reasoner’s Agar (R2A) and b) and yeast agar media (YMA) 20 plates, using the bacteria Acinetobacter nectaris and the yeast Metschnikowia reukaufii 21 for R2A and YMA plates, respectively. 22 23 24 A) Delpat ● Diccap ● 25 26 0.6 27 Colspa ● Asccor ● Staaju ● 28 0.4 Clapur ● R Mimaur ● ColhetTrilax● ● AscfasCyngra● ● ● 29 Penhet ● Delnud ● Erical 0.2 Trilan ● 30 Casocc ● PhacamSalcoa● ● Clacon ● Claung ● Amsmen ● Calocc Monvil● ● ● ● Asceri ● Aescal ScrcalChlpom● Clawil ● Dicvol ● Salmic ● 31 Lepcal ● Delhes ● 32 0.0 Antvex ● Salspa ● 33 Phacal ● 34 0 10 20 30 40 index 35 B) Diccap ● 36 0.8 Delpat ● 37 Asccor ● 38 0.6 Cyngra ● 39 40 0.4 Mimaur ● Colspa ● R ColhetAescal● ● Clapur ● Trilax ● Staaju ● Delnud ● 41 Penhet ● Erical ● Toxfre ● Trilan ●Salspa ● 0.2 ClaungMonvil● ● Scrcal ● Amsmen ● 42 Chlpom ● Phacam ● Salmic ● Ascfas ● Salcoa ● Clacon ● Lepcal ● Delhes ● 43 Calocc ● Asceri ● Casocc ● 0.0 Dicvol ● 44 Clawil ● Antvex ● Phacal ● 45 0 10 20 30 40 46 index 47 48 Supplementary Figure S3. Within-species correlation coefficients for Pearson 49 correlations between bacterial and fungal A) presence absence and B) abundance. 50 Points are colored black when the correlation is significant at FDR >0.05 and labeled 51 with genus and species abbreviations. Note that all but four correlation coefficients are 52 positive (above 0). 53 54 A) Bee Bio Stebbins 0.8 2016 0.4 variable 0.0 BactPres YeastPres 0.8 P(microbes) 2017 0.4 0.0 100 150 200 100 150 200 Julian 55 B) Bee Bio Stebbins 56 6 57 2016 4 58 2 59 variable 0 60 LogTotBact 61 6 LogTotFungi 2017 62 Log10(CFU+1) 4 63 2 64 0 100 150 200 100 150 200 65 Julian 66 Supplementary Figure S4. Seasonal patterns in the a) incidence of and b) 67 abundance of colony forming units on Reasoner’s agar (R2A) plates (bacteria, 68 blue) and colonies on yeast media agar plates (YMA, fungi, in orange) found in 69 floral nectar from two sampling sites in Northern California. In panel b) points are 70 only shown for non-zero values. Sites were sampled through the duration of the 71 main spring flowering period at both sites during 2016 and 2017. Lines represent 72 loess fit with 95% confidence values. Points represent individual nectar samples. 73 74 75 76 77 78 Supplementary Figure S5. Summary of taxa detected in pooled nectar samples. 79 Bacterial data were recovered from N=7 samples for 16S and N=17 samples for ITS 80 sequences. Chloroplast contamination accounted for 37% of reads in the ITS reads and 81 30% of reads in the 16S dataset. 82 83 84 85 Supplementary Figure S6. Frequency of detection for most abundant microbial genera 86 found in floral nectar and identified by MALDI, plotted by flower morphology of plant 87 species from which it was isolated. Blue bars indicate that taxa was more frequently 88 detected than expected by chance and red bars indicate that taxa was less frequently 89 detected than expected by chance. Bar thickness is proportional to the number of times 90 this species was detected in each flower types. P-value indicates significance based on 91 an independence model for a contingency table. 92 B Exposed Within_corolla_short Within_corolla_mid Within_corolla_long Pearson residuals: 3.5 Acinetobacter Bacillus 2.0 Candida A Erwinia 0.0 Metschnikowia -2.0 Micrococcus -2.8 p-value = 1.9685e-10 Pseudomonas 93 94 Supplementary Figure S7. Frequency of detection for most abundant microbial genera 95 found in floral nectar and identified by MALDI, plotted by time of the flowering season, 96 defined by Julian date, where ‘early’<100, 100≤‘mid’<150, and ‘late’≥150. Blue bars 97 indicate that taxa was more frequently detected than expected by chance and red bars 98 indicate that taxa was less frequently detected than expected by chance. Bar thickness 99 is proportional to the number of times this species was detected. P-value indicates 100 significance based on an independence model for a contingency table. 101 B early late mid Pearson residuals: Acinetobacter 3.2 Bacillus Candida 2.0 Erwinia Lactobacillus A 0.0 Metschnikowia Micrococcus Pantoea -2.0 -2.7 Pseudomonas p-value = 1.3043e-06 Rosenbergiella 102 103 Supplementary Table S1. Microbial strains used in dispersal assays, closest BLAST 104 match for species and genus, host species sources, and identifying sequences. Primer 105 sequences used to amplify yeasts were NL1/NL4 or ITS1f/ITS4; bacteria were amplified 106 using 27F/1492R. 107 Strain ID Kingdom species_Genus Host plant or sequence insect source EC52 F Metschnikowia_reukaufii Epilobium_canu CCTTCGGGAATTGTATTTTGAAGGT GGGTTTGGTTAGGAAAAGTTACTTT m AAGTCCATTGGAAAATGGCGCCATG GAGGGTGATAGCCCCGTAAAAGTAT CCCTTTTCCTTTTATCCATTCCCTCC AAAGAGTCGAGTTGTTTGGGAATGC AGCTCTAAGTGGGTGGTAAATTCCA TCTAAAGCTAAATATTGGCGAGAGA CCGATAGCGAACAAGTACAGTGATG GAAAGATGAAAAGCACTTTGAAAAG AGAGTGAAAAAGTACGTGAAATTGT TGAAAGGGAAGGGCTTGCAAGCAG ACACAACCTCGGTTGGGCCAGCATC GGAGTGGGGGGAGACAAAAAAGAA AAGGAATGTAACTCATTGAGTATTAT AGCCTTTTTCTCATATCTCCACCCCC TTCCG EC124 B Rosenbergiella_nectarea Epilobium_canu TCTTTTGCAACCCACTCCCATGGTG TGACGGGCGGTGTGTACAAGGCCC m GGGAACGTATTCACCGTAACATTCT GATTTACGATTACTAGCGATTCCGA CTTCATGGAGTCGAGTTGCAGACTC CAATCCGGACTACGACGCACTTTAT GAGGTCCGCTTGCTCTCGCGAGGT CGCTTCTCTTTGTATGCGCCATTGTA GCACGTGTGTAGCCCTACTCGTAAG GGCCATGATGACTTGACGTCATCCC CACCTTCCTCCGGTTTATCACCGGC AGTCTCCTTTGAGTTCCCACCATTAC GTGCTGGCAACAAAGGATAAGGGTT GCGCTCGTTGCGGGACTTAACCCAA CATTTCACAACACGAGCTGACGACA GCCATGCAGCACCTGTCTCAGAGTT CCCGAAGGCACTAAAGCATCTCTGC TAAATTCTCTGGATGTCAAGAGTAG GTAAGGTTCTTCGCGTTGCATCGAA TTAAACCACATGCTCCACCGCTTGT GCGGGCCCCCGTCAATTCATTTGAG TTTTAACCTTGCGGCCGTACTCCCC A FO-03 B Rosenbergiella_sp Frankliniella_occi CTTGCTACTTTGCTGACGAGTGGCG GACGGGTGAGTAATGTCTGGGGAT dentalis CTGCCTGATGGAGGGGGATAACTAC TGGAAACGGTAGCTAATACCGCATA ATGTCGCAAGACCAAAGCGGGGGA CTTTCGGGCCTCGCACCATCAGATG AACCCAGATGGGATTAGCTAGTAGG TAAGGTAATGGCTTACCTAGGCGAC GATCCCTAGCTGGTCTGAGAGGATG ACCAGCCACACTGGAACTGAGACAC GGTCCAGACTCCTACGGGAGGCAG CAGTGGGGAATATTGCACAATGGGC GCAAGCCTGATGCAGCCATGCCGC GTGTATGAAGAAGGCCTTCGGGTTG TAAAGTACTTTCAGTCAGGAGGAAG GGTGTGAAATTAATACTTTCATGCAT TGACGTTACTGACAGAAGAAGCACC GGCTAACTCCGTGCCAKC SCC477 B Acinetobacter_pollinis Scrophularia_cali TACTAGCGATTCCGACTTCATGGAG TCGAGTTGCAGACTCCAATCCGGAC fornica TACGATCGGCTTTTTGAGATTAGCAT CACATCGCTGTGTAGCAACCCTCTG TACCGACCATTGTAGCACGTGTGTA GCCCTGGCCGTAAGGGCCATGATG ACTTGACGTCGTCCCCGCCTTCCTC CAGTTTGTCACTGGCAGTATCCTTA AAGTTCCCATCCGAAATGCTGGCAA GTAAGGAAAAGGGTTGCGCTCGTTG CGGGACTTAACCCAACATCTCACGA CACGAGCTGACGACAGCCATGCAG CACCTGTATCTAAGTTCCCGAAGGC ACCAATCTATCTCTAGAAAGTTCTTA GTATGTCAAGGCCAGGTAAGGTTCT TCGCGTTGCATCGAATTAAACCACA TGCTCCACCGCTTGTGCGGGCCCC CGTCAATTCATTTGAGTTTTAGTCTT GCGACCGTACTCCCCAGGCGGTCT ACTTATCGCGTTAGCTGCGCCACTA AGTCCTCAAAGGACCCAACGGCT FO-01 B Pantoea_agglomerans Frankliniella_occi TCTTTTGCAACCCACTCCCATGGTG TGACGGGCGGTGTGTACAAGGCCC dentalis GGGAACGTATTCACCGTGGCATTCT GAKCCAYGATYAMAMTCGYWTCCG ACTTCACGGAGTCGAGTTGCAGACT CCGATCCGGACTACGACGCACTTTG TGAGGTCCGCTTGCTCTCGCGAGGT CGCTTCTCTTTGTATGCGCCATTGTA GCACGTGTGTAGCCCTACTCGTAAG GGCCATGATGACTTGACGTCATCCC CACCTTCCTCCGGTTTATCACCGGC AGTCTCCTTTGAGTTCCCGACCGAA TCGCTGGCAACAAAGGATAAGGGTT GCGCTCGTTGCGGGACTTAACCCAA CATTTCACAACACGAGCTGACGACA GCCATGCAGCACCTGTCTCASSGTT CCCGAAGGCACYAAAGCATCTCTGC Q1F2 F Starmerella_bombi Bombus_vosnes AACTCGCATTCGGAGAGTCATGCCG TACTCGTCGTCCGCTTAGTGTGATA enskii TAGCGACTGAAGCTATAATACTCCG AGGAGTTACATTCTTCAGCTTTTATC TTCCCCCCAAACACGACTCTACGTG GTTAGCGGCCTACCCTTCCATTTCA ACAATTTCACGTACTTTTTCACTCTC TTTTCAAAGTTCTTTTCATCTTTCCTT CACAGTACTTGTTCGCTATCGGTCT CTCGCAGATATTTAGCTTTAGATGG AGCATACCACCCATTTGAGCTGCAT TCCCAAACAACTCGACTCCATGCCA AGGTCCTACAGTGGGGTCAATGTCG TACGGGGCTATCACCCTCCATGGCG CTCCTTTCCAGAAGACTTAGACATC GGTTTCCCAGGACCAAGGCTTCAGA ATACAATGCCCCGAAAGGCTTTCAA ATCTGAGCTCTTGCCTGTTCACTCG CCGCTACTAAGGCAATCCCTGTTGG TTTCTTTTCCACCGCTTTTGATATGC AAA EC102 F Aureobasidium_pullulans Epilobium_canu CCCGACCTCCAACCCTTTGTTGTTA AAACTACCTTGTTGCTTTGGCGGGA m CCGCTCGGTCTCGAGCCGCTGGGG ATTCGTCCCAGGCGAGCGCCCGCC AGAGTTAAACCAAACTCTTGTTATTT AACCGGTCGTCTGAGTTAAAATTTT GAATAAATCAAAACTTTCAACAACGG ATCTCTTGGTTCTCGCATCGATGAA GAACGCAGCGAAATGCGATAAGTAA TGTGAATTGCAGAATTCAGTGAATC ATCGAATCTTTGAACGCACATTGCG CCCCTTGGTATTCCGAGGGGCATGC CTGTTCGAGCGTCATTACACCACTC AAGCTATGCTTGGTATTGGGCGTCG TCCTTAGTTGGGCGCGCCTTAAAGA CCTCGGCGAGGCCACTCCGGCTTT AGGCGTAGTAGAATTTATTCGAACG TCTGTCAAAGGAGAGGAACTCTGCC GACTGAAACCTTTATATTTTTCTAGG TTGACCTCGGATCAGGTAGGGATAC CCGCTGAACTT EC69 F Metschnikowia_koreensi Epilobium_canu GCTCAAATTTGAAATCCTCCGGGAA TTGTAATTTGAAGGTGGGGTTGAAT s m AGGTCTAGATACTTTAAGTCCATTG GAAAATGGCGCCMTGGAGGGTGAT AGCCCCGTAAAAGTATTCAAACCTT CTTTTCTTCCCCTCCTAAGARTCGA GTTGTTTGGGAATGCAGCTCTAGTG GGTGGTAAATTCCATCTAAAGCTAA ATATTGGCGAGAGACCGATAGCGAA CAAGTACAGTGATGGAAAGATGAAA AGCACTTTGAAAAGARAGTGAAAAA GTACGTGAAATTGTTGAAAGGGAAG GGCTTGCAAGCAGACACAACCTCG GTTGGGCCAGCATCGGARTGGGGG GARACAAAAAAGGTTA SCC187 B Pantoea_agglomerans Calystegia_occid TCTTTTGCAACCCACTCCCATGGTG TGACGGGCGGTGTGTACAAGGCCC entalis GGGAACGTATTCACCGTGGCATTCT