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Supplementary material: Bee diversity and pollination services improve with revegetation effort

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Supplementary material: Bee diversity and pollination services improve

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Craigburn Cleared Arctotheca calendula High Div Acacia myrtifolia

Arctotheca calendula Dodonea viscosa Echium plantagineum Eucalyptus leucoxylon Goodenia amplexans Grevillia lavandacea Grevillia sp._1 Grevillia sp._2 Olearia ramulosa Low Div Acacia acinacea

Acacia paradoxa Arctotheca calendula Dodonea viscosa Echium plantagineum Remnant Acacia paradoxa

Arthropodium strictum Burchardia umbellata Caesia calliantha Fumaria capreolata Hibbertia sp. _3 Hibbertia sp._1 Millbrook Cleared Hypochairis sp.

High Div Acacia acinacea Acacia myrtifolia Arctotheca calendula Arthropodium strictum Billardiera cymosa Bulbine bulbosa

Chrysocephalum apiculatum Chrysocephalum semipapposum Dianella revoluta

Grevillia lavandacea Hakea carinata Hakea rostrata

Hardenbergia violacea Hibbertia sp._2 Hypochaeris sp.

Kennidia prostrata

Leptorhynchos squamatus Leptospermum myrsinoides Linum marginale

Scaevola albida Tricoryne elatior Vittadinia cuneata Wahlenbergia sp

Xerochrysum bractiatum Remnant Arthropodium strictum

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Billardiera cymosa Brunonia australis Dillwinia hispida Diuris sp.

Drosera sp.

Goodenia blackiana Hibbertia sp. _3 Hibbertia sp._1 Hibbertia sp._2 Hypochaeris sp.

Kennidia prostrata Leptospermum sp.

Pultenaea largiflorens Calytrix sp.

Thelymitra antennifera Onkaparinga Cleared Arctotheca calendula

Trifolium sp.

Low Div Acacia paradoxa Acacia verticilata Sinapis sp.

Remnant Acacia myrtifolia Acacia verticilata Anagallis arvensis Arthropodium strictum

Boronia coerulescens s sp.coerulescens Burchardia umbellata

Caesia calliantha Calytrix tetragona Chamaescilla corymbosa Daviesia ulicifolia Dillwinia hispida Diuris sp.

Dodonea viscosa Eucalyptus sp.

Glossodia major

Helichrysum scorpioides Hibbertia sp._1

Thysanotus patersonia Wahlenbergia sp.

Table S2: Bee species list at each survey site Site Treatment Bee species

Clarendon High Div Amegilla (Zonamegilla) murrayensis Homalictus sp. BIN:AAM1019 Lasioglossum (Chilalictus) clelandi Lasioglossum (Chilalictus) expansifrons

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Lasioglossum (Chilalictus) lanarium Lasioglossum (Chilalictus) repraesentans Lasioglossum (Chilalictus) sp.

Lasioglossum (Chilalictus) willsi

Lasioglossum (Parasphecodes) hiltacus Lasioglossum sp.

Leioproctus (Euryglossidia) sp. BIN:AEC5195 Leioproctus (Leioproctus) platycephalus Leioproctus (Leioproctus) sp._b

Lipotriches (Austronomia) australica Trichocolletes venustus

Remnant Callohesma calliopsella Euhesma (Euhesma) bronsis Euriglossina sp. BIN:AEC4857 Euryglossa capitata

Euryglossina (Euryglossina) cf. hypochroma Euryglossina (Euryglossina) hypochroma Euryglossina (Euryglossina) stygica Exoneura (Brevineura) sp._b Exoneura (Exoneura) sp._a Exoneura sp. BIN:AAY4584 Exoneura sp. BIN:ABX9558 Exoneura sp. BIN:AEC5987

Homalictus (Homalictus) punctatus

Homalictus (Homalictus) sp. BIN:AAM1019 Homalictus sp. BOLD:AEC2529

Homalictus sp. BIN:AAM1019 Hylaeus (Euprosopis) elegans Hylaeus (Euprosopis) honestus

Hylaeus (Prosopisteron) sp. BIN:AAX2611 Hylaeus (Prosopisteron) sp. BIN:AEC3080 Hylaeus (Prosopisteron) sp. BIN:AEC3194 Hylaeus (Prosopisteron) sp. BIN:AEC4499 Hylaeus (Rhodohylaeus) proxima

Hylaeus sp. BOLD:AEC1767 Hylaeus sp. BOLD:AEC2227 Lasioglossum (Chilalictus) clelandi Lasioglossum (Chilalictus) erythrurum

Lasioglossum (Chilalictus) erythrurum BOLD:AEC6142 Lasioglossum (Chilalictus) imitans

Lasioglossum (Chilalictus) littleri Lasioglossum (Chilalictus) mundulum Lasioglossum (Chilalictus) sp.

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Lasioglossum (Chilalictus) sp._b Lasioglossum (Chilalictus) speculatum Lasioglossum (Chilalictus) vitripenne Lasioglossum (Chilalictus) willsi Lasioglossum (Ctenonomia) sp._c Lasioglossum (Parasphecodes) sulthica Lasioglossum(Chilalictus) vitripenne

Leioproctus (Euryglossidia) sp. BIN:AEC5195 Leioproctus (Leioproctus) cupreus

Leioproctus (Leioproctus) maculatus Leioproctus (Leioproctus) plumosus Leioproctus (platycephalus group) Leioproctus cupreus

Leioproctus sp. BIN:AEC3994 Lipotriches (Austronomia) australica Lipotriches (Austronomia) flavoviridis Lipotriches (Austronomia) moerens Craigburn Low Div Callohesma calliopsella

Euryglossa adelaidae Euryglossa sp.

Homalictus (Homalictus) punctatus Hylaeus (Prosopisteron) sp. BIN:AEC4499 Hyphesma atromicans

Megachile (Hackeriapis) tosticauda High Div Lasioglossum (Chilalictus) lanarium

Lipotriches (Austronomia) flavoviridis Megachile (Eutricharaea) obtusa Megachile (Eutricharaea) sp.

Megachile (Hackeriapis) oblonga Megachile (Torridapis) apicata Megachile (Unplaced) ordinaria Megachile sp.

Megachilidae sp.

Remnant Callohesma calliopsella Euryglossa adelaidae

Homalictus (Homalictus) punctatus Hylaeus (Rhodohylaeus) proxima Lasioglossum (Chilalictus) bicingulatum Lasioglossum (Chilalictus) brunnesetum Lasioglossum (Chilalictus) clelandi Lasioglossum (Chilalictus) speculatum Leioproctus (Leioproctus) maculatus Leioproctus (Leioproctus) sp._b

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Leioproctus (unplaced) sp. BIN:AEC1806 Lipotriches (Austronomia) flavoviridis Megachile sp.

Pachyprosopis (Pachyprosopis) haematosoma Millbrook High Div Lasioglossum (Chilalictus) clelandi

Lasioglossum (Chilalictus) lanarium Lasioglossum (Chilalictus) plorator Lasioglossum (Chilalictus) repraesentans Lasioglossum (Chilalictus) sp. BOLD:AEJ4129 Leioproctus (Leioproctus) maculatus

Leioproctus (Leioproctus) platycephalus

Leioproctus (unplaced or irroratus group) sp. BIN:AEC399 Lipotriches (Austronomia) australica

Lipotriches (Austronomia) flavoviridis Trichocolletes venustus

Remnant Lasioglossum (Chilalictus) lanarium Leioproctus (Leioproctus) maculatus Leioproctus (Leioproctus) sp._b Trichocolletes venustus

Onkaparinga Remnant Euryglossinae sp.

Exoneura sp.

Homalictus (Homalictus) punctatus Hylaeus (Euprosopis) honestus

Hylaeus (Prosopisteron) sp. BIN:AEC4499 Hylaeus (Prosopisteron) sp. near BIN:AEC2886 Hylaeus (Prosopsteron) sp. BIN:AEC4499 Hylaeus (Rhodohylaeus) proxima Hylaeus sp.

Lasioglossum (Chilalictus) brunnesetum Lasioglossum (Chilalictus) sp. BIN:AEC5169 Lasioglossum (Chilalictus) sp._a

Lipotriches (Austronomia) australica Trichocolletes venustus

Xanthesma (Xanthesma) argosomata

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Figure S1. Phylogenetic tree providing an overview of bee diversity across the study. Built using a list of unique species from the study with phyloT v2, based on NCBI taxonomy. Note:

this figure represents a very abbreviated list of species, since many species were identified to morphotypes, and many species were not available in NCBI at the time of building (June 2023).

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Table S3: Tukey’s pairwise comparison statistics for means of Shannon’s diversity indices for flower and bee surveys in four vegetation treatments. LCL and UCL are lower and upper 95% confidence limits.

Flower diversity Bee diversity

contrast Est. LCL UCL P val. Est. LCL UCL P val.

Low Div - Cleared 0.168 -0.712 1.05 0.94 0.570 -1.069 2.208 0.72 High Div - Cleared 1.65 0.766 2.53 <0.001 2.207 0.568 3.845 <0.01 Remnant - Cleared 1.27 0.456 2.09 <0.005 2.381 0.864 3.898 <0.005 High Div - Low Div 1.48 0.537 2.42 <0.005 1.64 -0.115 3.389 0.069 Remnant - Low Div 1.1 0.223 1.98 <0.05 1.811 0.173 3.450 <0.05 Remnant - High Div -0.375 -1.25 0.505 0.58 0.174 -1.464 1.813 0.99

Table S4: Tukey’s pairwise comparison statistics for means of species richness for flower and bee surveys, plus extrapolated bee species richness in four vegetation treatments.

Table S5. The estimated regression parameters, standard errors, z-values and significance of the difference in fruit set of Cullen australasicum and Arthropodium strictum with the cleared site (GLMM), assuming a binomial model. Bold indicates significance > 0.05.

Fixed effect (Treatment) Cullen australasicum Arthropodium strictum Est. S.E. z P val. Est. S.E. z P val.

(Intercept) -3.25 0.674 -4.83 1.38E-6 -2.44 0.451 -5.4 6.58E-08 Low Div 1.15 0.984 1.17 0.243 -0.235 0.662 -0.356 0.722 High Div 0.586 0.954 0.615 0.539 0.359 0.515 0.696 0.486 Remnant 1.72 0.924 1.86 0.0635 1.33 0.51 2.62 <0.01

Flower richness Bee richness Extrapolated bee richness contrast Est. S.E. Stat. adj. P val. Est. LCL UCL P val. Est. LCL UCL P val.

Low Div -

Cleared 0.88 0.56 1.57 0.37 0.69 -1.06 2.45 0.64 0.75 -1.34 2.84 0.70 High Div -

Cleared 2.57 0.47 5.47 <5e-07 2.52 0.76 4.28 <0.01 3.09 0.99 5.18 <0.01 Remnant -

Cleared 2.60 0.46 5.60 <1e-07 2.75 1.12 4.38 <0.005 3.75 1.81 5.69 <0.001 High Div - Low

Div 1.69 0.36 4.67 <5e-05 1.83 -0.05 3.7 0.057 2.34 0.10 4.57 <0.05 Remnant -

Low Div 1.72 0.36 4.84 <1e-05 2.06 0.30 3.81 <0.05 3.0 0.90 5.09 <0.01 Remnant -

High Div 0.03 0.19 0.13 1.0 0.23 -1.53 1.99 0.977 0.66 -1.43 2.75 0.771

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Table S6. Comparison between treatments of fruit set in Arthropodium strictum.

Columns are estimate of the difference between treatment means (Est.) using Tukey’s correction a family of 4 estimates, with standard errors (S.E.). Values with a

significance < 0.05 are indicated in bold. Results are on the log odds ratio scale. P value adjustment: calculated using the emmeans package in RStudio.

Pairwise comparison bet. treatments Est. S.E. z P val.

Cleared - Low Div 0.235 0.662 0.356 0.985

Cleared - High Div -0.359 0.515 -0.696 0.90

Cleared - Remnant -1.335 0.51 -2.618 <0.05

Low Div - High Div -0.594 0.549 -1.081 0.701

Low Div - Remnant -1.57 0.548 -2.866 <0.05

High Div - Remnant -0.976 0.349 -2.794 <0.05

Plaintext

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CHAPTER 3: A novel approach for pollen identification and quantification using hybrid capture-based DNA metabarcoding

Statement of Authorship

Title of Paper A novel approach for pollen identification and quantification using hybrid capture- based DNA metabarcoding

Publication Status

Publication Details Submitted to Molecular Ecology Resources, May 2023

Principal Author

Name of Principal Author (Candidate)

Contribution to the Paper Designed the study, undertook the practical work, analysed the data, and wrote the manuscript.

Overall percentage (%) 70%

Certification: This paper reports on original research I conducted during the period of my Higher Degree by Research candidature and is not subject to any obligations or contractual agreements with a third party that would constrain its inclusion in this thesis. I am the primary author of this paper.

Signature Date 5/6/23

Co-Author Contributions

By signing the Statement of Authorship, each author certifies that:

i. the candidate’s stated contribution to the publication is accurate (as detailed above);

ii. permission is granted for the candidate in include the publication in the thesis; and

iii. the sum of all co-author contributions is equal to 100% less the candidate’s stated contribution.

69 Plaintext

Name of Co-Author Karen Bell

Contribution to the Paper Study design, manuscript revision

Signature Date 6th June 2023

Name of Co-Author Katja Hogendoorn

Contribution to the Paper Study design, manuscript revision, project supervision

Signature Date 6/6/2023

Name of Co-Author Andrew Lowe

Contribution to the Paper Study design, manuscript revision, project supervision

Signature Date 6th June 2023

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A novel approach for pollen identification and quantification using