Transcript Document

2nd Quiz, Name, date
• 1 Pick one of the following two:
– A)What are microsatellites (SSR) and what are
they used for?
– B)Is it true plants are always susceptible to exotic
microbes?
• 2 Pick one of the following two:
– A) What are the infection pathways of Sudden Oak
Death?
– B)- What are the disease characteristics of
Sudden Oak Death?
Summary of 2nd class
• Host-Pathogen interactions.
Quantitative vs. qualitative resistance
• Plant recognition system recognizes
multiple signals, including sometimes
exotic pathogens
• Upon recognition plant produces
secondary metabolites that act as
antibiotics
Summary of 2nd class
• Introduced diseases are characterized
by simplified genetic structure
• Simplified means less allelic diversity
and/or less divergence among alleles
• DNA allows to compare individuals
based on differences of nucleotides
• DNA sequences need to be aligned
before they can be compared
Summary of 2nd class
• One locus may be useful or not, but we must be careful
because a single marker can be deceptive (gene acquired
from other species)
• Multiple unlinked markers boost confidence and allow to
identify individual. Following “individual” important to
understand pathways
• Need to compare sequence with dataset and unknown must
cluster with known species. To be significant, each species
must have its own statistically supported clade
Summary of 2nd class
• With multiple markers we can also see
whether there is linkage disequilibrium
or not, which is indicative of sex
• Sex important to predict adaptation to
new environment, but also important
because often exotic diseases have
only one mating type (hence no sex)
where they have been introduced
Summary of 2nd class
• Emergent disease: changes in host,
pathogen or environment
• Cypress canker a disease in the Med
because introduced (low genetic
diversity,no sex, all individuals
descending from a founder in Central
Italy which in turn comes from Northern
California)
• In CA, more diversity, possibly sex,
disaese because cypress planted away
from coast, or because a new species
was created for ornamental purposes
Why a disease in CA?
• If pathogen is native to California, why is it
causing such a serious disease?
• We observed that disease incidence is
variable with:
– cypress species,
– location,
Range of susceptibility
• Leyland cypress, Italian, monterey are
listed as most susceptible
• Arizona and McKnob are regarded as
more resistant
Range of susceptibility
• 90% of Leyland are heavily infected
• 10% of monterey
• LEYLAND CYPRESS IS AN ORNAMENTAL
CROSS, NOT NATIVE
Range of susceptibility
• Monterey is more susceptible in inland areas
where it is NOT NATIVE: we believe that
colder temperatures cause more wounds
that lead to infection
CONCLUSIONS
• Cypress canker is a serious disease in
Europe because pathogen was introduced
• Cypress canker is a serious disease in
California because hosts were introduced
either through planting off range (Monterey
cypress) or because host is artificial creation
(Leyland cypress); extinction of LEYLAND is
most likely
“Emergent diseases”:
2: environmental changes
• Forestry and intensive forest use:
timber production
tree felling and creation of stumps
fire exclusion and increase in density
oversimplified forest composition
changes in forest composition
changes in forest structure
Heterobasidion root disease
• Heterobasidion (a bracket or shelf
mushroom) infects trees through
wounds and stumps, then it spreads
through the roots to neighboring trees
• With tree felling,stumps and wounds are
created, suddenly exponentially
increasing infection levels
Heterobasidion shelf fruit-body
Use of molecular genetics:
• Differentiate Heterobasidion on fir/sequoias
(H. occidentalis) from that on pine/junipers
(H.irregularis)
• Show that airborne meiospores are
responsible for most infection of
Heterobasidion
• Show that in pines most infections start on
stumps and that in true firs most infections on
wounds
True firs
Pines
Each spore is a genetically different individual:
In pines we found the same genetic individual in stumps and
adjacent trees indicating direct contagion between the two
In true firs and true firs/sequoias we find same individual
in adjacent standing trees indicating infection not linked
to stumps but to wounds on standing trees
CONCLUSIONS:
• Logging activities increase Heterobasidion
infection because of stump creation in pines
and because of wounding in true firs
sequoias
• We have shown that in pine stumps H.
irregularis and H. occidentalis can both be
present and create a new hybrid entity
• We have shown that in the past these
hybridization events have lead to sharing of
genes among these two species (Horizontal
gene transfers)
Armillaria root diseases
• Armillaria, the honey mushroom, normally infects the
roots of trees. It can be a saprobe and a pathogen
and is common amongst oaks
• If woodland composition shifts to pine/oak, pines
become the target of attacks and gaps in canopy
enlarge over time. Stress (e.g. flooding) exacerbates
susceptibility
Clusters of Armillaria
How Does it Infect?
OAK
or
PINE
Two means of dispersal
to other trees:
DEAD
OAK
1.Mycelium can grow
through direct root contacts
and grafts with uninfected
trees.
2.Rhizomorphs can grow
through soil to contact
uninfected trees.
SOURCE: http://www.forestpathology.org/dis_arm.htm
l
What are Rhizomorphs?
• …“conglomerations of
differentiated parallel
hyphae with a protective
melanized black rind
on the outside.”
SOURCE: http://www.nifg.org.uk/armillaria.htm
• Rhizomorphs are able to transport food and
nutrients long distances which allows the fungus to
grow through nutrient poor areas located between
large food sources such as stumps.
SOURCE: http://botit.botany.wisc.edu/toms_fungi/apr2002.html
Humongous Fungus
It’s One of
U-HAUL’s
“Bizarre
Roadside
Attractions”
http://botit.botany.wisc.edu/toms_fungi/apr2002.html
CONCLUSIONS
Human activities shifting from oak
woodlands to mixed oak-pine lead
to large mortality gaps in pines
around oaks if honey mushroom is
present
CHANGING SPECIES
COMPOSITION LEADS TO
SEVERE DISEASE
Many gaps with very little regeneration and have not closed in
Change in gap area 19721999
Year
Area in gaps
(m2)
Percent in gaps
1972
6125
3.5
1999
53,981
31
“Emergent diseases”:
3: exotic pathogens
• 99% of times human responsible for their
introduction
Like the conquistadores brought
diseases that were lethal to
those who had never been
exposed to them, so do exotic
diseases cause true devastation
in plant communities because of
lack of coevolution between
hosts and microbes
California invaded: 1849 A.D.
Port Orford Cedar Root Disease
1950s
Sudden Oak Death
1990s
Canker-stain of
Sycamores 1980’s
Pitch canker disease
1980s
New hybrid root pathogen
1990s
Manzanita/madrone
die-back
White pine blister rust
1930s
Dutch Elm Disease
1960s
Oak root canker
2000
How can people transport
pathogens
• By transporting plants and plant parts
– Crops, and seeds
– Raw food
– Ornamental plants
Untreated lumber
Soil
Insects vectoring fungi
Military activity
The Irish Potato Famine
• From 1845 to 1850
• Phytophthora
infestans
• Resulted in the
death of 750,000
• Emigration of over 2
million, mainly to the
United States.
What favors invasion of exotic
fungi ?
– Density of host increases severity of disease
– Corridors linking natural habitats
– Synchronicity between host susceptibility and
pathogen life cycle
– Ecological and environmental conditions
Girdling aerial ‘cankers’
removed from roots
Big Sur
2006
K. Frangioso
% Mortality of Tanoak by Stem Size Class
% Mortality
45
40
35
30
25
20
15
10
5
0
35.8
P. ramorum absent
Non-infec
ted plots
P. ramorum
present
Infec
ted plots
10.7 11.8
28.5
12.4
4.1
1-<5
34.1
5-<10
4.8
10-<20
>20
Tanoak st em diamet er size c lass ( c m)
Wickland et al., unpublished
P. ramorum growing in a Petri dish
Organism new to science
•
•
•
•
Origin unknown
Biology unknown
Symptoms caused unknown
Immediately though highly regulated
Rhododendron:
In EU mostly a nursery
issue, but also present in
nurseries in US and Canada
Stem canker
Leaf necrosis
Phytophthora
ramorum
Sporangia
Chlamydospores
Is it exotic?
• Our studies have indicated that California
population is extremely simplified, basically
two strains reproducing clonally as expected
of an introduced organism
• Many hosts appear to have no resistance at
all
• Limited geographic distribution
Where does it come from?
• It is unknown where pathogen originally
comes from, but previous studies have shown
that California forest population is derived
from a relatively genetically diversified US
nursery population, indicating ornamental
nurseries were the most likely avenue for
pathogen introduction
Let’s look at its genetic
structure
• Need a number of independent and
neutral DNA markers
• Used AFLP, a technique that scans the
entire nuclear genome
• Are our isolates the same as the
European ones?
• Is the genetic structure suggestive of an
introduced or native species?
•US forest isolates
clearly distinct from EU
nursery isolates, also
have different mating
type
•Isolates from nurseries
in WA, OR, & BC both
of the US and EU types
•Potential for XXX sex
and recombination in
US nurseries
•US forest population
is genetically very
homogeneous,
trademark of an
introduced species
The entire genome was sequenced in
less than 3 years since discovery of organism
* 12 SSR loci (di- and tri- repeats identified)
* Loci selected to be polymorphic both between
and within continental populations
* 500+ representative isolates analyzed
CCGAAATCGGACCTTGAGTGCGGAGAGAGAGAGAGACTGTACGAGCCCGAGTCTCGCAT
Mating
Type
Growth
Rate
A1
Fast
A2
Slow
A2
Fast
Terminology
Genotype Lineage
Population
Results of 1st microsatellite
study
• There actually three distinct (genotypically
and phenotypically) lineages of P. ramorum
• Very low diversity in US forests (microsats
cannot discriminate among individuals,
clonality confirmed), only one lineage
• Several genotypes but only one lineage in EU
nurseries
• Three lineages in US nurseries
Was the pathogen first in US
forests or in US nurseries?
Slide 12
Was the pathogen first in US
forests or in US nurseries?
Slide 12
nurserie
forests
Where was it introduced?
• First reports mid 90’s
• Pathogen identified in
2000
• By then, the pathogen
was widespread
• CLUES: severity of
symptoms and
anedoctal stories
Positive isolation
P. ramorum
We found
same
genotypes in
nurseries and
forests proving
origin of wild
outbreak
Introduction phase
1- Escape of pathogen from
Infected nursery plants at two
locations: Mount Tamalpais
(Marin County), and Scott’s
Valley (Santa Cruz County)
2- Nurseries and two sites
have identical strain
composition, but distance
between sites is impossible
for natural spread of
organism
nurseries
What favors invasion of exotic
fungi ?
– Density of host increases severity of disease
– Corridors linking natural habitats
– Synchronicity between host susceptibility and
pathogen life cycle
– Ecological and environmental conditions
Bay/Oak association
Bay
Coast Live Oak (no sporulation)
Canker margin in phloem
Bleeding canker
Sporangia
Site
Mantel test among all individuals.
[Moran’s I vs ln (geographic
distance)]
ID
Correlation P-value
coeff. (r)
(1000,000
perm)
ALL
-0.2153
<0.000001
0.5
0.4
Moran's I
0.3
0.2
0.1
0
-0.1
-0.2
10
100
1000
Mean Geographic Distance (m)
10000
100000
Synchrony pathogen-host
Susceptibility of oaks
(lesion size)
Average lesion (mm_)
80
60
40
Wetness > 12 h
20
0
0
6
12
18
24
30
36
42
48
54
Time (h)
2
Lesion area (mm )
50
40
30
Temp >19 C
20
10
0
15
17
19
21
23
25
Temperature (¡C)
27
29
Bay Laurel / Tanoak SOD Spore Survey
35
Temp (C)
30
Rain (mm)
25
20
15
10
5
0
Date
How to control emergent
exotic diseases
• PREVENT THEIR INTRODUCTION
• LIMIT THE HUMAN-SPREAD OF
PATHOGENS (infected plants, plant
parts, dirty tools)
• EMPLOY HOST RESISTANCE
• CHEMICAL AND OTHER MITIGATION
STRATEGIES
Forest pathogens can never be
eradicated
PREVENT: Diagnose
Symptoms relatively generic, very
variable, and pathogen not always
culturable
DNA TESTS
LAB CULTURES
AgriFos and
PentraBark
Topical
Application
+
Agrifos vs. Azomite Treatments
(efficacy 1 - 24 months)
a
14
a
Canker Size (mm)
12
10
8
6
b
4
2
0
Azom ite
Pos itive Control
Agr ifos
Why emphasis on molecular
analyses?
• As a way to identify and quantify
microbes in the environment
• As a way to understand microbial
biology: how do microbes reproduce
and infect hosts
• As a way to determine epidemiology:
follow the movement of a strain
Why emphasis on molecular
analyses?
• As a way to determine potential for
spread: use genes as markers for
individuals
• As a way to determine whether
population of microbes is exotic or
native
• As a way to identify source of a
pathogen and migration patterns
Why emphasis on molecular
analyses?
• As a way to determine the size of the
gene pool of a pathogen, Important to
scale management options
• As a way to determine rapid
evolutionary changes linked to an
introduction
• As a way to determine epigenetic
effects