Multilevel interactions between microbes, insects and plants: … · 2020. 10. 13. · 17 Matteo Brunetti PhD students "Annual report day" - 25 September 2019 Perspectives • Extract

Multilevel interactions between microbes, insects and plants:

ecological and evolutionary constraints underlying interactions

PhD student:

Matteo Brunetti

Tutor: Franco Faoro

Co-tutor: Matteo Montagna

Insect-plant coevolutionary arm race

Futuyma & Agrawal 2009, Proc. Natl. Acad. Sci., 106(43), 18054-18061

Host plantsPhytophagous

insects

Erlich & Raven, 1964, Evolution, 586-608; Bernays & Graham, 1988, Ecology, 69(4), 886-892; Mitter et al., 1988, Am Nat,

107-128; Farrell et al., 1992, BioScience, 34-42; Karban & Agrawal, 2002, Annu. Rev. Ecol. Syst., 641-664; Futuyma &

Agrawal, 2009, Proc. Natl. Acad. Sci., 106(43), 18054-18061; Janz, 2011, Annu. Rev. Ecol. Evol. Syst., 42(1), 71; Ali &

Agrawal, 2012, Trends Plant Sci., 17(5), 293-302; Forister et al., 2012, Ecology, 93(5), 981-991

Matteo

BrunettiPhD students "Annual report day" - 25 September 2019

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mod. from Hansen & Moran (2014). Molecular ecology, 23(6), 1473-1496.

Moran et al., 2008, Annu. Rev. Genet., 42, 165-190

Clark et al., 2010, Protoplasma, 244(1-4), 25-51

Feldhaar, 2011, Ecol. Entomol., 36(5), 533-543

Frago et al., 2012, Trends ecol. evol., 27(12), 705-711

Guerrero et al., 2013, Int. Microbiol., 16(3), 133-143

Chung et al., 2013, Proc Natl A Sci, 110(39), 15728-15733

Zhu et al., 2014, New Phytologist, 204(2), 315-321

Hansen & Moran, 2014, Mol ecol, 23(6), 1473-1496

Douglas, 2015, Annu rev entomol, 60, 17-34

Mason et al., 2018, Plant Cell Environ, 0

Jones et al., 2019, eLife, 9(1), 2792

Traits mediated by microbial symbionts

in phytophagous insects

• Provide essential nutrients lacking in

insect diet (e.g. Buchnera

aphidicola)

• Digestion of plant macromolecules

(e.g. Termites fermentation

chambers)

• Detoxification (e.g. Dendroctonus

ponderosae)

• Suppress plant defences (e.g.

Liberibacter psyllaurous)

Matteo


Microbiota of phytophagous insects

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Multilevel interactions

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~ 37,000 species (worldwide)

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Leaf beetles (Coleoptera: Chrysomelidae)

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Bacterial communities associated to insects feeding on the same host

plant share common features, both in terms of taxonomic composition

and dominant metabolic pathways

Bacterial communities associated to insects feeding on plants with

common defense strategies (e.g. same toxic compounds) share common

features, both in terms of taxonomic composition and dominant

metabolic pathways

Bacterial communities hosted by polyphagous insect species are more

complex than those hosted by monophagous species.

[H1]

[H2]

[H3]

Matteo


Hypothesis

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7

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Characterize the bacterial communities associated to ~200 species of leaf

beetles (Coleoptera: Chrysomelidae) of the Euro-Mediterranean region

and investigate possible correlation with the phylogeny and the ecology of

the insect hosts

Characterize the metabolic potential of the identified microbiotas and

correlate it with characteristics of the host plants

Extend the framework of this project using already published microbiotas

data (online databases) and correlate features of the microbial

communities with characteristics of the insect host (e.g., taxonomy, diet,

habitat, life stage)

[A1]

[A2]

[A3]

Aims

• DNA extraction, libraries preparation

and 16S (V1-V2 & V4 regions) high-

throughput sequencing (Ion Torrent)

• Cluster analysis and taxonomic

identification (QIIME2 platform) [A1]

• Functional characterization of bacterial

communities (PICRUSt2) [A2]

• Published microbiota data mining (NCBI

SRA tool kit) [A3]

• Data analysis (QIIME2, R software)

[A1-A3]

Matteo


Materials and methods

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Bibliographic research

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Best methodology for 16S metabarcoding

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Preliminary results: whole bacterial communities

Monophagous Oligophagous Poliphagous

NO clear pattern

NO statistically

significant result

Primary symbionts could

play a confounding role

Exclude known primary

symbionts (e.g., Wolbachia)

from subsequent analyses

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Preliminary results: alpha diversity

Kruskal-Wallis test H p-value

Monophagous-Oligophagous 1.31 0.25

Monophagous-Poliphagous 6.22 0.013

Oligophagous-Poliphagous 2.14 0.14

All groups 6.77 0.049

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All-groups 11.54 0.32

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All-groups 11.54 0.32

p-value < 0.05

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Preliminary results: Beta diversity

Pairwise-permanova pseudo-F p-value





Unweighted Unifrac Distance (D):

➢ D = 0 →

➢ D ~ 0.5 →

➢ D = 1 →Monophagous Oligophagous Poliphagous

identical

communities

related

communities

unrelated

communities

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Matteo


Preliminary results: Beta diversity

Pairwise-permanova pseudo-F p-value





Unweighted Unifrac Distance (D):

➢ D = 0 →

➢ D ~ 0.5 →

➢ D = 1 →Monophagous Oligophagous Poliphagous

identical

communities

related

communities

unrelated

communities

17

Matteo


Perspectives

• Extract DNA from about 200 species of leaf beetles from Euro-

Mediterranean region and sequence the V1-V2 and V4 regions

of the bacterial 16S rRNA gene.

• Development of the dataset made of already published data on

microbiota associated with insects.

• Characterize the metabolic potential of the obtained microbiotas

(PICRUSt2) and correlate it with the secondary metabolites

present in the host plant tissues.

• Analyze (QIIME2, R software) the obtained datasets in the light

of information on characteristics of the insect hosts (e.g.,

taxonomy, host plant, habitat, life stage)

• Prof. Franco Faoro

• Dr. Matteo Montagna

• Dr. Giulia Magoga

THANK YOU FOR YOUR ATTENTION!

WORKING GROUP:

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Multilevel interactions between microbes, insects and plants: … · 2020. 10. 13. · 17 Matteo Brunetti PhD students "Annual report day" - 25 September 2019 Perspectives • Extract