Plants and Arthropods Friends or Foes

What are arthropods?
Arthropods are an immense, diverse group of invertebrates
Some are terrestrial and others aquatic
Some mites and insects are major herbivores that can cause extensive damage to living plants

Arthropods cause crop yield losses of ~10 - 30% every year
Photo by Scott Bauer USDA
How do arthropods damage plants?
How do plants defend themselves from herbivores?
How do herbivores cope with plant defenses?
How do plants establish mutualistic interactions with arthropods including pollinators and their herbivores' natural enemies?

Lecture outline
1. 400 million years of living together
2. Basic conflict – herbivory
Plant defense and herbivore counter-measures
Constitutive defenses
Induced defenses
Secondary metabolites in defense
3. Alliance #1 – Carnivorous and / or parasitoid arthropods
Herbivore-induced volatiles guide foraging carnivores to prey
Domatia and extrafloral nectar can accommodate carnivores
4. Alliance #2 – Pollinators
Physiological compatibility between plants and pollinators
Controlling pollen and nectar theft

A long-term, complicated relationship
M. sexta eating N. attenuata
The basic conflict: plants are food for herbivorous arthropods
Nicotiana attenuata pollinated by Manduca sexta moth
Alliance #2: Most angiosperms rely upon arthropods for successful reproduction
Alliance #1: Predatory or parasitic arthropods protect plants from herbivore damage
Photo credits: Danny Kessler; R.J. Reynolds Tobacco Company Slide Set, Bugwood.org; Wu, J., Hettenhausen, C., Meldau, S. and Baldwin, I.T. (2007). Herbivory rapidly activates MAPK signaling in attacked and unattacked leaf regions but not between leaves of Nicotiana attenuata. Plant Cell. 19: 1096-1122.
M. sexta after attack by predator

The evolution of herbivory
Photo credit: Fossil mall

400 million years ago the world was very different
Map copyright Ron Blakey and Colorado Plateau Geosystems, Inc.
The land was mostly barren, continents were far from their present locations, and the atmosphere, flora and fauna were vastly different than today

400 million year-old fossils show evidence of herbivory
Image credit: Miguasha National ParkImage credit: Miguasha National Park; Jeram, A.J., Selden, P.A. and Edwards, D. (1990). Land Animals in the Silurian: Arachnids and Myriapods from Shropshire, England. Science. 250: 658-661 reprinted with permission from AAAS. Edwards, D., Selden, P.A., Richardson, J.B. and Axe, L. (1995). Reprinted by permission from Macmillan Publishers Ltd: Coprolites as evidence for plant-animal interaction in Siluro-Devonian terrestrial ecosystems. Nature. 377: 329-331.
Fossils of early plants
Fossils of early terrestrial arthropods
Fossilized feces (coprolites) showing ingested plant material

Plants and arthropods share 400 million years of evolution
Image derived from : L. Shyamal Image derived from : L. Shyamal based on work by Bruce Tiffney

The ongoing conflict: Herbivory
Phloem-sucking aphids
Mesophyll-sucking mites
Mesophyll-grazing leaf miners
Root-vascular cylinder sucking nematodes
Leaf-chewing larvae
Plants produce energy from photosynthesis; animals are hungry and eat plants (or they eat plant-eaters)
Photo credits: Sate Al AbbasiPhoto credits: Sate Al Abbasi; David CappaertPhoto credits: Sate Al Abbasi; David Cappaert, Michigan State University, Bugwood.org; University of MissouriPhoto credits: Sate Al Abbasi; David Cappaert, Michigan State University, Bugwood.org; University of Missouri. Published by MU Extension,Photo credits: Sate Al Abbasi; David Cappaert, Michigan State University, Bugwood.org; University of Missouri. Published by MU Extension, all rights reserved. William WergiPhoto credits: Sate Al Abbasi; David Cappaert, Michigan State University, Bugwood.org; University of Missouri. Published by MU Extension, all rights reserved. William Wergi; John R. MeyerPhoto credits: Sate Al Abbasi; David Cappaert, Michigan State University, Bugwood.org; University of Missouri. Published by MU Extension, all rights reserved. William Wergi; John R. Meyer, North Carolina State University; Scott Bauer, USDA

Plants respond differently to different types of feeding behaviour
Chewing insects cause extensive wounding, and some partially digest their food outside their bodies by regurgitation
Piercing arthropods have needle-like stylets that pierce tissues and then suck nutrients out
Photo credits: Sate Al AbbasiPhoto credits: Sate Al Abbasi; John R. Meyer, North Carolina State University;

Piercing-sucking and chewing may have evolved more than once
Chewers and suckers are found among both carnivores and herbivores
Photo credits: Jan van Arkel (IBED; University of Amsterdam); R.J. Reynolds Tobacco Company Slide SetPhoto credits: Jan van Arkel (IBED; University of Amsterdam); R.J. Reynolds Tobacco Company Slide Set, Bugwood.org; Scott Bauer, USDA Agricultural Research Service, Bugwood.org

Some arthropods form galls
Clemson University Clemson University - USDA Cooperative Extension Slide Series; Milan ZubrikClemson University - USDA Cooperative Extension Slide Series; Milan Zubrik, Forest Research Institute, Slovakia; Gyorgy CsokaClemson University - USDA Cooperative Extension Slide Series; Milan Zubrik, Forest Research Institute, Slovakia; Gyorgy Csoka, Hungary Forest Research Institute; Milan ZubrikClemson University - USDA Cooperative Extension Slide Series; Milan Zubrik, Forest Research Institute, Slovakia; Gyorgy Csoka, Hungary Forest Research Institute; Milan Zubrik, Forest Research Institute, Slovakia; Sturgis McKeever, Georgia Southern University, Bugwood.org
Gall on oak caused by cynipid gall wasp; opened gall showing adults (A) and larvae (L)
Galls are localized tissue proliferations induced by arthropods, which lay their eggs in them, or pathogens. Insects and some pathogens form galls by manipulating plant hormones. Galls are often more nutritious and less defended than other tissues, but it is not yet understood how this occurs
L
A
A
L

Constitutive plant defenses and herbivore countermeasures
Photo scopyright Chris DarlingPhoto scopyright Chris Darling; Dussourd, D., and Eisner, T. (1987). Vein-cutting behavior: insect counterploy to the latex defense of plants. Science 237: 898-901 reprinted with permission of AAAS; .
Labidomera clivicollis cutting a trench through the leaf tissue to avoid ingesting sticky, toxic latex

Plants and plant tissues vary in their degree of defensiveness
Nutritional value also matters – nutrient-rich tissues including seeds are often heavily defended
Seeds of apricots and related fruits contain toxins such as amygdalin
Photo credit Stephen Ausmus
Herbivores preferentially consume nutrient-rich plant material

Plants have evolved many ways to defend against herbivory
Physical
Ranger, C.M., and Hower, A.A. (2001). Glandular morphology from a perennial alfalfa clone resistant to the potato leafhopper. Crop Sci. 41: 1427-1434.
Nicotine

Physical defenses against herbivory
Thorns
Trichomes (hairs)
Wax and wall
Latex and resin
Waxy cuticle
Cell wall
Cardoso, M.Z. (2008). Herbivore handling of a Plant’s trichome: the case of Heliconius charithonia (L.) (Lepidoptera: Nymphalidae) and Passiflora lobata (Killip) Hutch. (Passifloraceae). Neotropical Entomology 37: 247-252.

Eisner, T., Eisner, M. and Hoebeke, E.R. (1998). When defense backfires: Detrimental effect of a plant’s protective trichomes on an insect beneficial to the plant. Proc. Natl. Acad. Sci. USA 95: 4410-4414, copyright National Academy of Sciences, USA.
Mentzelia pumila trichomes and trapped insects
From an arthropod’s perspective trichomes can be lethal

Some arthropods avoid sticky trichomes or push past them
Voigt, D. and Gorb, S. (2010). Locomotion in a sticky terrain. Arthropod-Plant Interactions. 4: 69-79Voigt, D. and Gorb, S. (2010). Locomotion in a sticky terrain. Arthropod-Plant Interactions. 4: 69-79; Russ Ottens, University of Georgia, Bugwood.org.
Long legs help rise above sticky materials
Force and non-stick coatings help push past sticky materials
Claws grab onto trichome stalks to generate force for movement

Some arthropods cover trichomes with silk or bite off the hooked ends
Photo copyright Dale Clark; Cardoso, M.Z. (2008). Herbivore handling of a Plant’s trichome: the case of Heliconius charithonia (L.) (Lepidoptera: Nymphalidae) and Passiflora lobata (Killip) Hutch. (Passifloraceae). Neotropical Entomology 37: 247-252.
The hooked trichomes of a passionflower have been bitten off and covered with silk
Passion flower
Zebra Longwing larvae

Trichomes can release chemical deterrents to arthropods
Reprinted by permission from Macmillan Publishers Ltd. from Gibson, R.W., and Pickett, J.A. (1983). Wild potato repels aphids by release of aphid alarm pheromone. Nature 302: 608-609.
Tomato (Solanum lycopersicum)
Tomato trichomes produce chemicals repellent to whiteflies
Wild-potato trichomes produce aphid alarm pheromones
Many trichomes produce sticky substances

Sometimes freshly hatched caterpillars “lick” the sugars off trichomes …
… but this gives them a “smell” that betrays them to their enemies
Manduca licks the trichomes of Nicotiana to eat the acyl-sugars these secrete
The ant Pogonomyrmex rugosus forages using the smell of those same volatile acyclic fatty acids
Suggestion for extra slide
Photo credits: Alex Weinhold and Ian BaldwinPhoto credits: Alex Weinhold and Ian Baldwin; Weinhold, A. and Baldwin, I.T. (2011). Trichome-derived O-acyl sugars are a first meal for caterpillars that tags them for predation. Proc. Natl. Acad. Sci. 108: 7855-7859Photo credits: Alex Weinhold and Ian Baldwin; Weinhold, A. and Baldwin, I.T. (2011). Trichome-derived O-acyl sugars are a first meal for caterpillars that tags them for predation. Proc. Natl. Acad. Sci. 108: 7855-7859; Louisa Howard, Dartmouth University
False color image of Nicotiana trichomes

Latex can be avoided through vein biting or trenching
Dussourd, D., and Eisner, T. (1987). Vein-cutting behavior: insect counterploy to the latex defense of plants. Science 237: 898-901 reprinted with permission of AAAS.
Labidomera clivicollis cutting veins of Asdcepias syraca prior to consuming distal tissues
Latex is sticky and often toxic. Herbivores can cut the veins to drain out latex and render the tissue edible

Induced defenses and herbivore countermeasures

Perception of herbivory
Biting and wounding damage
Piercing and saliva
Oral secretions and regurgitant
Egg laying damage and secretions
Phillip RobertsPhillip Roberts, USDA Forest ServicePhillip Roberts, USDA Forest Service University of Georgia, Bugwood.org; Mithöfer, A. and Boland, W. (2008). Recognition of herbivory-associated molecular patterns. Plant Physiology. 146: 825-831; .

Volicitin and inceptin are herbivory-specific compounds
Volicitin is produced from the conjugation of a plant-derived fatty acid conjugated to glutamine in the herbivore midgut, then regurgitated onto plant tissues
Inceptin is a peptide derived from proteolytic cleavage by the insect of a chloroplastic ATP synthase from the plant

Wu, J., Hettenhausen, C., Schuman, M.C. and Baldwin, I.T. (2008). A comparison of two Nicotiana attenuata accessions reveals large differences in signaling induced by oral secretions of the specialist herbivore Manduca sexta. Plant Physiology. 146: 927-939.
The fatty-acid conjugates (FACs) and other herbivore signals induce expression of secondary metabolite biosynthetic genes and other defense & repair genes
?
Herbivory-specific compounds induce plant defenses

Phillip RobertsPhillip Roberts, USDA Forest Service University of Georgia, Bugwood.org
Jasmonate
(JA)
Salicylate
(SA)
Ethylene
(ET)
Direct responses (e.g. toxins)
Indirect responses (e.g. predator attraction)
Hormone dependent signaling
Many induced defense responses are mediated by hormones
Hormone-independent signaling
eggs

Infestation induces expression of defense-associated genes
Plants respond to the type of herbivory – different herbivores induce different subsets of genes. In general, infested plants induce synthesis or accumulation of toxins, anti-nutritives, and damage repair compounds.
Photo credits: Sate Al AbbasiPhoto credits: Sate Al Abbasi; John R. MeyerPhoto credits: Sate Al Abbasi; John R. Meyer, North Carolina State University; Mites Photo credits: Sate Al Abbasi; John R. Meyer, North Carolina State University; Mites CopyrightPhoto credits: Sate Al Abbasi; John R. Meyer, North Carolina State University; Mites Copyright 1993 to 2011 University of Missouri. Published by MU Extension, all rights reserved.

Herbivory induces direct and indirect defenses locally and systemically
Local signals and hormone accumulation
Long distance signals and systemic responses
Induction of volatiles that attract natural enemies of herbivores (indirect defenses)
Wu, J., Hettenhausen, C., Meldau, S., and Baldwin, I.T. (2007). Herbivory rapidly activates MAPK signaling in attacked and unattacked leaf regions but not between leaves of Nicotiana attenuata. Plant Cell 19: 1096-1122.
Induction of direct defenses
(e.g. alkaloids and other toxins, proteinase inhibitors)
Induction of direct defenses
(e.g. alkaloids and other toxins, proteinase inhibitors) albeit later and often less strong

Some herbivores can suppress induced plant defense responses
T. urticae induces tomato defenses and its fitness on tomato is moderate
T. evansi suppresses induced defenses in tomato and is a serious pest on it
The fitness of T. urticae increases in the presence of T. evansi
Tetranychus evansi
T. urticae and T. evansi together
Tetranychus urticae
Some, but not all, spider mite species suppress or delay plant defenses
Photo: Jan van Arkel (IBED; University of Amsterdam)

Systemic signals are molecules that can move through vascular tissues and are related to jasmonate and salicylate. Also volatile hormones (ethylene) and possibly derivatives (methyl jasmonate and methyl salicylate) contribute to the systemic response
Green, T.R., and Ryan, C.A. (1972). Wound-induced proteinase inhibitor in plant leaves: A possible defense mechanism against insects. Science 175: 776-777.
Summary: defenses are induced locally and some also systemically

Defensive chemicals and herbivore countermeasures
Plants produce >100.000 compounds
80% of the known natural compounds have a botanical origin
Many of these chemicals function in defense
The raison d’être of secondary plant substances “is to repel and attract insects” – Fraenkel 1959

Some defense compounds are “secondary metabolites”
Primary metabolites: amino acids, sugars, nucleotides, lipids →
found throughout the plant kingdom
Secondary metabolites: defense and attractant functions, phylogenetically restricted
Redrawn from Hartmann, T. (1996). Diversity and variability of plant secondary metabolism: a mechanistic view. Entomologia Experimentalis et Applicata 80: 177-188.

Some secondary compounds attract pollinators, predators or parasitoids
Aroma
Color
Nectar
Odor
pollinator
predator
parasitoid
herbivore
Photo source: Klaus Bolte, Natural Resources Canada Ottawa, Ontario, Canada

Other compounds are toxic, or can be converted to toxins, or are anti-nutritive
Nicotine, a toxin found in tobacco and its relatives
Cyanide
A cyanogenic glycoside that releases toxic cyanide
Anti-nutritives interfere with herbivores’ digestion or assimilation of nutrients, impairing their growth, development and reproduction

Some herbivores have evolved tolerance to plant toxins
In some cases the target enzyme has been modified to now be unaffected by the toxin
Herbivores can tolerate plant toxins through degradation, excretion and sequestration (through chemical modification and storage in specialized glands).

Redrawn from Hartmann, T. (1996). Diversity and variability of plant secondary metabolism: a mechanistic view. Entomologia Experimentalis et Applicata 80: 177-188.
Defensive secondary metabolites can be roughly divided in three groups
Terpenoids:
e.g. Limonoids
Saponins
Pinene
Phenolic: e.g. Flavonoids; Salicylic acid; Lignins etc
Nitrogen containing:
e.g. Alkaloids
Glucosinolates

Phenolics and terpenes include medicines, insecticides and irritants
And at least 10,000 more, many uncharacterized…
Taxol
From Pacific yew tree
Pyrethrin
From Chrysanthemum
Urushiol
From poison ivy
Photo credit: Dave Powell, USDA Forest Service, Bugwood.org

Parasitic plants also sense volatile terpenoids produced by hosts…
Runyon, J.B., Mescher, M.C. and De Moraes, C.M. (2006). Volatile chemical cues guide host location and host selection by parasitic plants. Science. 313: 1964-1967 reprinted by permission from AAAS.
Cuscuta pentagona (dodder) grows towards a tomato plant, but shows the same response to volatiles collected from the tomato

Opium Poppy
Coffee
Cacao
Alkaloids contain nitrogen and include stimulants and narcotics
Coca
caffeine
Nicotine
Cocaine
Morphine
Theobromine

Glucosinolates are typical for the Cabbage (Brassicaceae) Family

Myrosinases and glucosinolates are stored in separate plant cells…

Glucosinolate accumulation pattern
Helicoverpa armigera (the cotton bollworm) avoids tissues with high concentrations of glucosinolates
Shroff, R., Vergara, F., Muck, A., Svatoš, A. and Gershenzon, J. (2008). Nonuniform distribution of glucosinolates in Arabidopsis thaliana leaves has important consequences for plant defense. Proc. Natl. Acad. Sci. USA 105: 6196-6201.
Glucosinolates accumulate in midveins and the outer parts of the leaf blade. Bollworms selectively eat the middle of the blade on normal plants but not on mutant plants that that do not accumulate glucosinolates.
Less herbivory of high-glucosinolate tissue

Most herbivores avoid Brassicaceae but some can eat it
Image credits (all Bugwood.org): Whitney CranshawImage credits (all Bugwood.org): Whitney Cranshaw, Colorado State University; David CappaertImage credits (all Bugwood.org): Whitney Cranshaw, Colorado State University; David Cappaert, Michigan State University; Keith NaylorImage credits (all Bugwood.org): Whitney Cranshaw, Colorado State University; David Cappaert, Michigan State University; Keith Naylor; David CappaertImage credits (all Bugwood.org): Whitney Cranshaw, Colorado State University; David Cappaert, Michigan State University; Keith Naylor; David Cappaert, Michigan State University; David JonesImage credits (all Bugwood.org): Whitney Cranshaw, Colorado State University; David Cappaert, Michigan State University; Keith Naylor; David Cappaert, Michigan State University; David Jones, University of Georgia; David Riley, University of Georgia.

The diamondback moth has an enzyme that eliminates glucosinolates
Ratzka, A., Vogel, H., Kliebenstein, D.J., Mitchell-Olds, T. and Kroymann, J. (2002). Disarming the mustard oil bomb. Proc. Natl. Acad. Sci. USA. 99: 11223-11228Ratzka, A., Vogel, H., Kliebenstein, D.J., Mitchell-Olds, T. and Kroymann, J. (2002). Disarming the mustard oil bomb. Proc. Natl. Acad. Sci. USA. 99: 11223-11228; Russ Ottens, University of Georgia, Bugwood.org
The diamondback moth sulfatase (DBM GSS) enzyme removes the glucosinolate’s sulfate group such that myrosinase does not recognize the glucosinolate
anymore
Plutella xylostella
Glucosinolate
Toxins

Cabbage white butterfly larvae convert glucosinolates into less-toxic products
It sequesters the glucosinolates as protection against predators
Wittstock, U., Agerbirk, N., Stauber, E.J., Olsen, C.E., Hippler, M., Mitchell-Olds, T., Gershenzon, J., and Vogel, H. (2004). Successful herbivore attack due to metabolic diversion of a plant chemical defense. Proc. Natl. Acad. Sci. USA 101: 4859-4864Wittstock, U., Agerbirk, N., Stauber, E.J., Olsen, C.E., Hippler, M., Mitchell-Olds, T., Gershenzon, J., and Vogel, H. (2004). Successful herbivore attack due to metabolic diversion of a plant chemical defense. Proc. Natl. Acad. Sci. USA 101: 4859-4864; David Cappaert, Michigan State University, Bugwood.org
Adult form

Glucosinolate feeding has been lost in some lineages (blue bars). Black dots indicates NSP, white dots indicates absence of NSP
Wheat, C.W., Vogel, H., Wittstock, U., Braby, M.F., Underwood, D., and Mitchell-Olds, T. (2007). The genetic basis of a plant–insect coevolutionary key innovation. Proceedings of the National Academy of Sciences 104: 20427-20431.
NSP production has been lost in some related butterflies

Summary: ingested secondary defense compounds can be toxic but sometimes herbivores have adapted
Rapid degradation or conversion
Rapid excretion
Sequestration
Health problems

Case study: Milkweeds and monarch butterflies
Photo credits: Kenneth Dwain HarrelsonPhoto credits: Kenneth Dwain Harrelson, Steven Katovich, USDA Forest Service, Bugwood.org
Milkweed (Asclepias syriaca) produces latex that contains a toxic alkaloid ouabain
Ouabain binds to the Na+, K+ ATPase and interferes with its function

The Na+, K+ ATPase from monarch butterfly larvae is insensitive to ouabain
Reprinted from Vaughan, G.L., and Jungreis, A.M. (1977). Insensitivity of lepidopteran tissues to ouabain: Physiological mechanisms for protection from cardiac glycosides. J. Insect Physiol. 23: 585-589, with permission from Elsevier.
Silkmoth Hyalophora cecropia, Tobacco hornworm Manduca sexta (sensitive)
Monarch Danaus plexippus (insensitive)
HIS
ASN
An amino acid substitution in the Na+, K+ ATPase confers ouabain insensitivity to monarchs

Monarch butterflies avoid predation through ouabain accumulation

Some butterflies mimic the monarch’s colors to avoid predation
Viceroy butterfly – nontoxic but protected by mimicry

Some predators have become tolerant of the monarch’s ouabain

Case study: Heliconian butterflies and passion flowers
South American Heliconian butterflies were collected and studied as early as the 17th century, and provided the basis for ideas about mimicry
Maria Sybilla Merian 1705; William M. CieslaMaria Sybilla Merian 1705; William M. Ciesla, Patricia M. Ciesla, Forest Health Management International, Bugwood.org

Some Heliconius butterflies can detoxify a cyanogenic glucoside
Reprinted with permission from Macmillan Publishers Ltd: Engler, H.S., Spencer, K.C., and Gilbert, L.E. (2000) Preventing cyanide release from leaves. Nature 406: 144–145Reprinted with permission from Macmillan Publishers Ltd: Engler, H.S., Spencer, K.C., and Gilbert, L.E. (2000) Preventing cyanide release from leaves. Nature 406: 144–145; Photo credit Dale Clark.
The larvae of more than 60 species of Heliconius butterflies are specialized feeders of Passiflora and are tolerant of their various secondary metabolites
Toxic cyanide

Passionflower plants make structures that resemble butterfly eggs
Female butterflies prefer to lay their eggs on an unoccupied leaf to protect their young from cannibals and hence avoid leaves with ‘fake’ eggs
Photo copyright Missouri Botanic GaPhoto copyright Missouri Botanic Garden; Williams, K.S. and Gilbert, L.E. (1981). Insects as selective agents on plant vegetative morphology: Egg mimicry reduces egg laying by butterflies. Science. 212: 467-469Photo copyright Missouri Botanic Garden; Williams, K.S. and Gilbert, L.E. (1981). Insects as selective agents on plant vegetative morphology: Egg mimicry reduces egg laying by butterflies. Science. 212: 467-469 reprinted with permission from AAAS; Jerry A. Payne, USDA Agricultural Research Service, Bugwood.org
Heliconius sara

Case study: Aphids and whiteflies, phloem-feeding insects
Uzest, M., Gargani, D., Drucker, M., Hébrard, E., Garzo, E., Candresse, T., Fereres, A., and Blanc, S. (2007). A protein key to plant virus transmission at the tip of the insect vector stylet. Proceedings of the National Academy of Sciences 104: 17959-17964.
Phloem-feeding insects are major agricultural pests. They insert stylets into phloem, depleting the host plant of nutrients and spreading disease.
Downy mildew growing on honey dew
Whitefly
Whitefly juveniles (crawlers)
Salivary channel
Common duct
Food channel
Aphid stylet tip

Aphid saliva interferes with the normal phloem-sealing wound response
Aphids insert a thin stylet into the phloem
Reprinted from Hogenhout, S.A., and Bos, J.I.B. (2011) Effector proteins that modulate plant–insect interactions. Curr. Opin. Plant Biol. 14: 422-428 with permission from Elsevier.

The green pea aphid genome has been sequenced
The International Aphid Genomics Consortium 2010 Genome Sequence of the Pea Aphid Acyrthosiphon pisum. PLoS Biol 8(2): e1000313.
Acyrthosiphon pisum
Aphids can reproduce clonally as well as sexually. Shown here is a female giving birth to live female clones

Plants and herbivores - summary
Constitutive defenses like poisonous or sticky trichomes deter most herbivores
Scott BauerScott Bauer, USDA Agricultural Research Service, Bugwood.org; Mike Speed, University of Liverpool
Synthesis of anti-nutritives and toxins
Wound healing responses
Herbivory elicits the induction of additional defense responses
Some herbivores have evolved counter-measures to deal with plant defenses

Alliance #1 – Plants and Carnivores or Parasitoids
Parasitoid larvae
Photo: T. Bukovinszky / www.bugsinthepicture.comwww.bugsinthepicture.com; Thailand IPM; R.J. Reynolds Tobacco Company Slide SetR.J. Reynolds Tobacco Company Slide Set , Bradley HigbeeR.J. Reynolds Tobacco Company Slide Set , Bradley Higbee, Paramount Farming; Bugworld.org; Scott Bauer, USDA
Lady beetle devours a pea aphid
Assassin bug

Plants betray herbivores to their natural enemies via volatile signals
http://pb.pharmazie.uni-halle.de/anschrift/forschung/2012131_2199270/?lang=en
Photo credit: Ted Turlings

Tritrophic interactions involve three food levels
Reprinted from Dicke, M., and Baldwin, I.T. (2010) The evolutionary context for herbivore-induced plant volatiles: beyond the ‘cry for help’. Trends Plant Sci. 15: 167-175, with permission from Elsevier.
1st trophic level
2nd trophic level
3rd trophic level

Chemical information moves between and amongst trophic levels
Herbivore-induced plant volatiles can attract or repel carnivores and other herbivores. Other plants may also perceive this information
Reprinted from Dicke, M., and Baldwin, I.T. (2010) The evolutionary context for herbivore-induced plant volatiles: beyond the ‘cry for help’. Trends Plant Sci. 15: 167-175, with permission from Elsevier.

Most natural enemies of arthropods use a combination of visual and olfactory cues to track down their prey or host
When plants facilitate such prey-finding by natural enemies, for example via the release of herbivore-induced odors, we call this induced indirect defense
Volatiles contribute to induced indirect defenses

Set-up for collecting volatiles from whole plants
Induced organic volatiles contribute to indirect defense responses
Plants produce many volatile compounds – some of these are herbivore-induced plant volatiles that contribute to the establishment of indirect defenses.
The arbitrary air volume surrounding a plant we call the plant’s “head space”.
Absorbent to collect volatiles
Airflow with volatiles from plants
Airflow without volatiles
Photo: Merijn R. Kant

Gas chromatography mass spectrometry (GC-MS)
Induced volatiles: qualifying and quantifying plant volatiles
Sampled volatiles (in solvent) injected into the GC
Separation in the GC
Identification via the MS ionization spectra
Photo: Merijn R. Kant

Infested plant head-space chromatogram
Y-axis: Number of ions counted by the MS after ionization
X-axis: Time it took each molecule to reach the end of the GC
Number of ions counted by the MS
All different ions counted at 308 sec
All different ions counted at 410 sec
All different ions counted at 215 sec
Image: Merijn R. Kant

Kant, M.R., Bleeker, P.M., Van Wijk, M., Schuurink., R.C., Haring, M.A. (2009). Plant volatiles in defence. Adv. Bot. Res. 51: 613-666.
Herbivore-induced plant volatiles
Terpenes
Phenolics

Green leaf volatiles are rapidly released from wounded tissue
Reprinted from Matsui, K. (2006). Green leaf volatiles: hydroperoxide lyase pathway of oxylipin metabolism. Curr. Opin. Plant Biol. 9: 274-280, with permission from Elsevier.
Green leaf volatiles (GLV) are released from wounded tissues (they are the “cut grass” smell) and convey information.

Terpenoids are common plant compounds: when induced they often attract predatory arthropods
Herbivore induced terpenes
Photo credit: Ted Turlings

Spider mite populations grow rapidly and destroy plants: their blind natural enemy uses plant odors to find them
Spider mite (0.5 mm)
Tetranychus urticae attacked by its natural enemy, the predatory mite Phytoseiulus persimilis
Photo credits: Bill Bumgarner

Quantifying volatile effects on arthropod foraging behaviour – the olfactometer
As an example, predatory mites chose volatiles collected from leaves invested with herbivorous mites 3x as often as from control leaves
Dicke, M., van Loon, J.J.A. and Soler, R. (2009). Chemical complexity of volatiles from plants induced by multiple attack. Nat Chem Biol. 5: 317-324Dicke, M., van Loon, J.J.A. and Soler, R. (2009). Chemical complexity of volatiles from plants induced by multiple attack. Nat Chem Biol. 5: 317-324; Shimoda, T. and Dicke, M. (2000). Attraction of a predator to chemical information related to nonprey: when can it be adaptive? Behavioral Ecology. 11: 606-613, by permission of Oxford University Press; Photo credit : Merijn R. Kant.
Odor from control plant
Odor from infested plant
Hungry predator that has to choose
?
Infested with T. urticae
control
Using a Y-shaped tube, the arthropod is given a choice between two volatile samples, and the frequency that each is chosen is determined
Phytoseiulus persimilis is blind and uses odors to find plants with T. urticae

Plants can be engineered to produce predator-attracting volatiles
From Kappers, I.F., Aharoni, A., van Herpen, T.W.J.M., Luckerhoff, L.L.P., Dicke, M. and Bouwmeester, H.J. (2005). Genetic engineering of terpenoid metabolism attracts bodyguards to Arabidopsis. Science. 309: 2070-2072, reprinted with permission from AAAS.
Predatory mites were attracted to plants producing the terpenoid nerolidol, even in the absence of herbivores

Herbivore-induced plant volatiles can also attract parasitoid arthropods
Parasitoid wasps lay their eggs in other arthropods. When the larvae hatch they eat the host……
Photo: T. Bukovinszky / www.bugsinthepicture.com

Glucosinolate hydrolysis results in plant volatiles that attract parasitoids
Glucosinolate hydrolysis due to P. rapae feeding
The parasitic wasp Cotesia rubecula is attracted to plants that produce nitriles while Pieris rapae butterflies avoid ovipositing on these plants when it can choose. The P. rapae larvae however do not mind these nitriles.
Photo credit: Hans van Pelt

Some plants form longer-term alliances with resident “bodyguards”
Acacias provide ants with shelter and food from extrafloral nectaries (EFN) and food bodies (FB), also known as Beltian bodies
EFN
Ants protect acacias from other plants and other arthropods.
Photos courtesy of Dan Janzen, University of Pennsylvania

Myrmecophyte nectar is optimized for its ant partner
Most nectar contains sucrose and most ants produce invertase that hydrolyzes sucrose to monosaccharides
The Pseudomyrmex ants that live on acacia do not produce invertase, and the nectar of myrmecophytes contains fructose and glucose but not sucrose
Heil, M., Rattke, J., and Boland, W. (2005). Postsecretory Hydrolysis of Nectar Sucrose and Specialization in Ant/Plant Mutualism. Science 308: 560-563.
There are > 100 ant-mutualist plants called myrmecophytes

Other plants have other kinds of domatia that shelter predatory arthropods
Photo courtesy Jim Conrad: Matos, C.H.C., Pallini, A., Chaves, F.F. and Galbiati, C. (2004). Do coffee domatia benefit the predatory mite Iphiseiodes zuluagai Denmark & Muma (Acari: Phytoseiidae)? Neotropical Entomology. 33: 57-63.
Domatia can be simple or elaborate, but by providing a dwelling for predatory arthropods plants can protect themselves from herbivory

Plant alliances with other arthropods against herbivores
Plant volatiles operate as direct defenses when they are toxic or when they repel herbivores
The indirect defense mechanism of plants involves:
the attraction of predatory and parasitoid arthropods via induced plant volatiles
arrestment and longer-term associations with these beneficial organisms via alternative food and shelter
Photo: T. Bukovinszky / www.bugsinthepicture.com

Summary
Many of the compounds that contribute to plant direct and indirect defenses have been identified, opening up the possibility to engineer plants with enhanced defenses
Reprinted from Dicke, M., and Baldwin, I.T. (2010) The evolutionary context for herbivore-induced plant volatiles: beyond the ‘cry for help’. Trends Plant Sci. 15: 167-175, with permission from Elsevier.

Towards herbivore-resistant crops
Defense compounds are often toxic to humans as well and can cost the plant quite some energy to produce. Hence many such defenses were lost, deliberately or not, during breeding, making crops vulnerable to pests
Enhancing plants’ inducible defenses may make them more herbivore-resistant
Human alliances with herbivore-predators can enhance food production

Many plants distribute their pollen via mobile arthropods
Image source: Market wallpapers

Alliance #2 – Plants and Pollinators
Christian Konrad Sprengel (1793) “Das entdeckte Geheimnis der Natur im Bau und in der Befruchtung der Blumen”

Pollination is the transfer of pollen from the anther to the stigma
The pollen contains the sperm which travels through the pistil to fertilize the egg
Only a small number of plants regularly self-pollinate; most require cross-pollination

Pollination by arthropods is mutually beneficial
Approximately 84% of commercial crops depend on pollinators, mostly insects and mostly honeybees
Pollinators are rewarded by eating nectar or protein-rich pollen
Photo courtesy of Jeff Pettis, ARS.

What attracts pollinators?
Visual cues – color, shape, patterns
Olfactory cues – fragrances, other volatile compounds
Nectar
Surface characteristics of petals

Flowers and pollinators evolved physiological compatibilities
Photos by Jack DykingaPhotos by Jack Dykinga; Rob Flynn, USDA-ARSPhotos by Jack Dykinga; Rob Flynn, USDA-ARS; Hans Hillewaert

Bee vision color spectrum is shifted as compared to human
Bee photoreceptors are most sensitive to UV, blue and green
Human photoreceptors are most sensitive to blue, green and red
Bee spectral sensitivity adapted from Arnold, S., Savolainen, V. and Chittka, L. (2009). Flower colours along an alpine altitude gradient, seen through the eyes of fly and bee pollinators. Arthropod-Plant Interactions. 3: 27-43.

Flower pigments also reflect or absorb UV-light which is visible to bees
Visible light
Simulated bee color vision
Images (c) Dr Klaus Schmitt, Weinheim, www.uvir.eu; Benitez-Vieyra, S., de Ibarra, N.H., Wertlen, A.M. and Cocucci, A.A. (2007). How to look like a mallow: evidence of floral mimicry between Turneraceae and Malvaceae. Proc. Roy. Soc. B. 274: 2239-2248.

Flowers vary in their aroma, and aroma production is developmentally regulated
Guterman, I., et al. (2002). Rose scent: Genomics approach to discovering novel floral fragrance– related genes. Plant Cell. 14: 2325-2338.

Many fragrance elements are conserved but the blend is unique
Dudareva, N. and Pichersky, E. (2000). Biochemical and molecular genetic aspects of floral scents. Plant Physiology. 122: 627-634.

Floral nectar is an attractor and sweet reward for pollinators
Nectar was an early innovation of flowers and is an important contributor to the success of angiosperms
Ren, D. (1998). Flower-associated brachycera flies as fossil evidence for jurassic angiosperm origins. Science 280: 85-88., reprinted with permission from AAAS; Image by artist Joseph ScheerRen, D. (1998). Flower-associated brachycera flies as fossil evidence for jurassic angiosperm origins. Science 280: 85-88., reprinted with permission from AAAS; Image by artist Joseph Scheer. David Cappaert, Michigan State University, Bugwood.org
150 million year old insect with nectar-feeding mouthparts
Many pollinators have tongues or other mouthparts specialized for nectar sipping

Nectaries are structurally and positionally diverse
Wist, T.J., and Davis, A.R. (2006). Floral Nectar Production and Nectary Anatomy and Ultrastructure of Echinacea purpurea (Asteraceae). Ann. Botany 97: 177-193Wist, T.J., and Davis, A.R. (2006). Floral Nectar Production and Nectary Anatomy and Ultrastructure of Echinacea purpurea (Asteraceae). Ann. Botany 97: 177-193, by permission of Oxford University Press; Heil, M. (2011). Nectar: generation, regulation and ecological functions. Trends Plant Sci 16: 191-200.
Nectaries can be found at the base of the ovary, filament or petal, often as a ring of tissue. Nectar can be secreted through trichomes, epidermal cells or non-functioning guard cells

Antirrhinum and bumblebee
Its genetic resources make Antirrhinum an excellent experimental organism for studies of plant – pollinator coevolution
These studies indicate that the shape, color, pattern, scent, and arrangement of flowers on the inflorescence are optimized for pollination by heavy, short-tongued, bumblebees

Rough petals help bees get a grip
Reprinted from Whitney, H.M., Chittka, L., Bruce, T.J.A., and Glover, B.J. (2009). Conical epidermal cells allow bees to grip flowers and increase foraging efficiency. Curr.Biol. 19: 948-953 with permission from Elsevier.
Wild-type conical cells
mixta flat cells
Movie S6. Flat Landing 3.
Bees need a rough petal surface to grip onto – the smooth cells of the mixta mutant cause increased aborted landings as bees slip and slide

Which floral cues are most important?
Hoballah, M.E., Gübitz, T., Stuurman, J., Broger, L., Barone, M., Mandel, T., Dell'Olivo, A., Arnold, M., and Kuhlemeier, C. (2007). Single gene-mediated shift in pollinator attraction in petunia. Plant Cell 19: 779-790Hoballah, M.E., Gübitz, T., Stuurman, J., Broger, L., Barone, M., Mandel, T., Dell'Olivo, A., Arnold, M., and Kuhlemeier, C. (2007). Single gene-mediated shift in pollinator attraction in petunia. Plant Cell 19: 779-790; Reprinted from Klahre et al., (2011) Pollinator choice in petunia depends on two major genetic loci for floral scent production, Curr. Biol. 21: 730-739 with permission from Elsevier.
Petunia axillaris
Petunia integrifolia
Petunia exserta
Three closely related petunias are pollinated by moths, bees and hummingbirds

Which cues are most important?
Petunia axillaris
Petunia integrifolia
Petunia exserta
Hoballah, M.E., Gübitz, T., Stuurman, J., Broger, L., Barone, M., Mandel, T., Dell'Olivo, A., Arnold, M., and Kuhlemeier, C. (2007). Single gene-mediated shift in pollinator attraction in petunia. Plant Cell 19: 779-790Hoballah, M.E., Gübitz, T., Stuurman, J., Broger, L., Barone, M., Mandel, T., Dell'Olivo, A., Arnold, M., and Kuhlemeier, C. (2007). Single gene-mediated shift in pollinator attraction in petunia. Plant Cell 19: 779-790; Reprinted from Klahre et al., (2011) Pollinator choice in petunia depends on two major genetic loci for floral scent production, Curr. Biol. 21: 730-739 with permission from Elsevier.

Mixing and matching traits in Petunia
Petunia axillaris
Petunia integrifolia
Petunia exserta
When presented with mixed cues (red scented vs white non-scented), moths get confused and select at random.
Hoballah, M.E., Gübitz, T., Stuurman, J., Broger, L., Barone, M., Mandel, T., Dell'Olivo, A., Arnold, M., and Kuhlemeier, C. (2007). Single gene-mediated shift in pollinator attraction in petunia. Plant Cell 19: 779-790Hoballah, M.E., Gübitz, T., Stuurman, J., Broger, L., Barone, M., Mandel, T., Dell'Olivo, A., Arnold, M., and Kuhlemeier, C. (2007). Single gene-mediated shift in pollinator attraction in petunia. Plant Cell 19: 779-790; Reprinted from Klahre et al., (2011) Pollinator choice in petunia depends on two major genetic loci for floral scent production, Curr. Biol. 21: 730-739 with permission from Elsevier.

Plants are picky about which pollinators they choose as allies
Photo courtesy of David CappaertPhoto courtesy of David Cappaert, Michigan State University, Bugwood.org; Hoballah, M.E., Gübitz, T., Stuurman, J., Broger, L., Barone, M., Mandel, T., Dell'Olivo, A., Arnold, M., and Kuhlemeier, C. (2007). Single gene-mediated shift in pollinator attraction in petunia. Plant Cell 19: 779-790;

Benzyl acetone
Some flower components are attractive
Pollinators
Nutrients – sugars and animo acids
Pollinators
Predators, parasitoids
Redrawn from Heil, M. (2011). Nectar: generation, regulation and ecological functions. Trends Plant Sci 16: 191-200 with permission from Elsevier.

Some flower components e.g. from nectar are “repellent”
Catalpol
Nectar robbers
Nectarins
Microorganisms (i.e. yeast)
Nectar robbers
Pollinators
Gelsemine
H2O2
Hydrogen peroxide accumulation at nectary opening
Nicotine
H2O2 prevents microbial growth in sugar-rich nectar
Redrawn from Heil, M. (2011). Nectar: generation, regulation and ecological functions. Trends Plant Sci 16: 191-200Redrawn from Heil, M. (2011). Nectar: generation, regulation and ecological functions. Trends Plant Sci 16: 191-200 with permission from Elsevier; Carter, C., Healy, R., O'Tool, N.M., Naqvi, S.M.S., Ren, G., Park, S., Beattie, G.A., Horner, H.T., and Thornburg, R.W. (2007). Tobacco nectaries express a novel NADPH xxidase implicated in the defense of floral reproductive tissues against microorganisms. Plant Physiology 143: 389-399..

Fragrance can also be deceptive or repellent
γ-decalactone
Howard, D.F., Blum, M.S., and Fales, H.M. (1983). Defense in thrips: Forbidding fruitiness of a lactone. Science 220: 335-336 with permission from AAAS; Ômura, H., Honda, K., and Hayashi, N. (2000). Floral scent of Osmanthus fragrans discourages foraging behavior of cabbage butterfly, Pieris rapae. J. Chem. Ecol. 26: 655-666. J. Chem. Ecol. 26: 655-666; Reprinted by permission from Macmillan Publishers Ltd. Ledford, H. (2007) Plant biology: The flower of seduction. Nature 445: 816-817.
Osmanthus fragrans produces a pollination deterrent that is also a defense compound produced by thrips

Nectar location forces the pollinator to interact with reproductive tissues
Dave Powell, USDA Forest Service, Bugwood.org
The length of the spur correlates with the length of the pollinators tongue

Figs and fig wasps need each other
Photo credits: David KarpPhoto credits: David Karp, Forest & Kim Starr, Starr Environmental, Bugwood.org
Figs and fig wasps are mutually interdependent for reproduction. Most figs are pollinated by a single species of wasp

Figs have an enclosed inflorescence that the pollinator must enter
Adapted from Cook, J.M., and Rasplus, J.-Y. (2003). Mutualists with attitude: coevolving fig wasps and figs. Trends Ecol. Evol. 18: 241-248.
1. Adult female crawls into immature “fruit” and deposits eggs into female flowers
2. Fruit matures, wasps hatch, pick up pollen as leave “fruit” (some flowers make seeds)
3. Pollen-laden wasps enter immature “fruit”, fertilize female flowers and deposit eggs (REPEAT....)
Female flowers inside inflorescence

Nicotiana attenuata and Manduca sexta
Manduca is a specialist herbivore that feeds on Nicotiana and can sequester and secrete nicotine. Manduca is also the main pollinator of this plant, so it is both a “friend” and “foe”.
How does the plant balance its two conflicting relationships with Manduca??

Severe caterpillar herbivory shifts flowers to opening in the morning
The adult moths are nocturnal and normally the flowers open at night
Kessler, D., Diezel, C., and Baldwin, I.T. (2010). Changing pollinators as a means of escaping herbivores. Curr. Biol. 20: 237-242, reprinted by permission of Elsevier.
Caterpillar infestation leads to more flowers opening in the morning, when the moths are not active

The morning-opening flowers don’t attract moths
Benzyl acetone
As a consequence, adult moths don’t pollinate and don’t lay eggs, reducing caterpillar numbers
DUSK
DAWN
The moth-attractant benzyl acetone (BA) is emitted at night, when open in the morning the flowers are not producing the attractant BA.
Kessler, D., Diezel, C., and Baldwin, I.T. (2010). Changing pollinators as a means of escaping herbivores. Curr. Biol. 20: 237-242, reprinted by permission of Elsevier.

The morning-opening flowers don’t attract moths
Benzyl acetone
DUSK
DAWN
The moth-attractant benzyl acetone (BA) is emitted at night, when open in the morning the flowers are not producing the attractant BA.
Opportunistic hummingbirds take nectar from and pollinate the morning-open flowers
The plant switches pollinators to escape herbivory!
Kessler, D., Diezel, C., and Baldwin, I.T. (2010). Changing pollinators as a means of escaping herbivores. Curr. Biol. 20: 237-242, reprinted by permission of Elsevier.

Nicotine gets plants more pollination service for less nectar production
Kessler, D., Gase, K. and Baldwin, I.T. (2008). Field experiments with transformed plants reveal the sense of floral scents. Science. 321: 1200-1202 reprinted by permission of AAAS.
Plants producing nicotine get more and shorter visits from pollinators: shorter because the pollinator can handle only small amounts of nicotine; more since the pollinator needs nectar

Plants and pollinators - summary
Millions of years of evolution underlie plant-pollinator mutualism, and the ability of plants to guard against nectar and pollen thieves

Natural selection maximizes reproductive success and dishonesty can do just that
But do organisms that cheat, steal and deceive succeed on the long run?
Deceiver – tricks other species into providing services under false pretences
Cheater – A freeloader who abuses the honesty of its own kind
Cheaters, Thieves and Deceivers

Thieves: Nectar robbers take nectar without loading pollen
Photo credit: Debbie Roos, North Carolina Agricultural Extension Agent

Shiojiri, K., Ozawa, R., Kugimiya, S., Uefune, M., van Wijk, M., Sabelis, M.W. and Takabayashi, J. (2010). Herbivore-specific, density-dependent induction of plant volatiles: Honest or “Cry wolf” signals? PLoS ONE. 5: e12161.
Cheaters: Some cabbages excessively produce volatiles (they cry wolf)
HELP!
?
HELP!
Plants that cry wolf or produce large amounts of volatiles under low herbivory pressure pressure gain in the short term but undermine the alliance between plant and carnivore……

Deceivers: Why does this flower look like a female bee?
Many orchids have evolved similarity to female arthropods, to entice visits by males. Some also emit the female’s chemical attractants. By the time the males realize their mistake, they’ve already delivered or picked up pollen.
Final score: Plant 1, Pollinator 0.
Image courtesy Hans Hillewaert

Some plants have moved up to the third tropic level: deceivers?
Photo credits: Tom DonaldPhoto credits: Tom Donald; Sturgis McKeeverPhoto credits: Tom Donald; Sturgis McKeever, Georgia Southern University; Rebekah D. Wallace, University of Georgia, Bugwood.org
Carnivorous plants use trap and sticky trichomes to catch their prey

Leaf cutter bee: Friend or Foe?
Leaf cutter bees are efficient pollinators but also damage plants: friendly foes?
Photo courtesy Joseph BergerPhoto courtesy Joseph Berger, Bugwood.org; Jim Bennett

Alliances are tenuous
Organisms act in their own self-interest and are the product of selection for maximal fitness
When species use each other, opportunities arise for stealing, cheating and / or deception
As honesty becomes rarer, the advantage of dishonesty decreases
Many arthropods can learn; this will select against dishonesty in plants
Cheating and deception may emerge and disappear through time