Xylella fastidiosa

Invasive Potential of Xylella fastidiosa?
Alexander Purcell
Dept. Environmental Science, Policy, and Management
University of California, Berkeley
Does Europe/Asia have the requirements
for Xylella to survive from year to year?
To cause and sustain crop/forest diseases?
Host plants are widespread and common in Europe:
Known propagative hosts
Known and potential vectors
Historical and current importations of live host plants
So why no permanently established Xylella
fastidiosa in Europe/Asia?
the players
Photo UC IPM
Layers of complexity:






environment (e.g. temperature)
vector ecology
pathogen ecology
host plant ecology
outcome of various interactions
disease management
Xylella fastidiosa
• Xylem-limited bacterium
• Colonizes a wide range of host plants, usually
without causing disease
• Present throughout the Americas
– Causing disease in grape at least since the 1880s
– Major crops affected
• Grape, alfalfa, peach, etc
• Xylem sap-feeding insects are only vectors
• Pierce’s Disease in grapes,
• Alfalfa dwarf,
• Almond leaf scorch
• Phony peach disease, Plum leaf scald,
• Citrus variegated chlorosis
• Elm, oak, sycamore leaf scorch
• Oleander leaf scorch
* and a very large list of new emerging
diseases of crops and ornamentals
Pierce’s disease (PD)
Photo UC IPM
Environmental conditions determine the
spatial and temporal spread of Xylellacaused diseases
Pierce’s Disease in coastal California
(ex.: Napa Valley)
Photo J. Clark
Main vector = blue-green sharpshooter
Breeds in vegetation along streams (riparian);
Adults enter vineyards in spring
* Disease pattern resembles spring pattern of vector
Pierce’s Disease in central California
Usually occurs near irrigated
crops or canals with grass weeds
Main vectors = green and red-headed
Sharpshooters; breed in grasses near vineyards
Pierce’s Disease in Napa Valley)
Photo J. Clark
Incidence highest at edge of vineyard
Incidence decreases away from edge
grape
alfalfa
Guadalupe Valley, Baja California
Bermuda grass sources for redheaded sharpshooter
(bare field lost to Pierce’s disease)
Vector is a rare visitor to grape
Redheaded
sharpshooter
Bermuda grass sources for
red-headed sharpshooter
Pierce’s Disease in Napa Valley)
Photo J. Clark
What causes this pattern?
Pierce’s disease:
Effects of date of inoculation on recovery
100
Percent recovery
80
Oakville 1998
Davis 1998
Davis 1999
Parlier 1999
Parlier 2000
60
40
20
0
80
100
April
120
140
160
Day of year
Feil, et al. 2003 Phytopathology 93:244+
180
200
220
August
240
Infectious adults (can fly)
needed for April-May
infections
Seasonality component…
Hill & Hashim 2006
1. Seasonal acquisition efficiency
-low Xylella populations
early in the year
-threshold population for
sharpshooter acquisition
-minimal transmission early on?
Hill and Purcell 1997
2. Vine recovery
-vines lose infection over winter
-mechanism not well understood
(pruning, infection level)
-recovery rate highly dependent on
date of original infection
Feil & Purcell 2003
Does the lack of overwintering adult vectors
prevent the establishment of chronic disease
in perennial plants in Europe?
Meadow spittle bug (Philaenus spumarius)
is abundant and widespread in Europe, as are
other potential vectors
BUT they overwinter as eggs, NOT ADULTS
Winter severity (sub-freezing temperatures)
determine the limits of Pierce’s disease?
Seasonal and climatic effects
“severe” = commercial
viticulture is impossible
Isotherms (black lines) = average
minimum January temperature
Texas – PD Risk Ratings
Reasons for disappearance of Xylella
during winter (winter recovery)?
Unknown – thus cannot use climatic
criteria for predictions
Points to Remember:
* Infection alone is not disease
(Xylella is NOT a virus)
Most infected plants are symptomless
* Plants can recover from Xylella infection
* There are numerous ways to create and
sustain PD epidemics. The proper combinations
of vectors, plant communities, climate are
especially important.
Invasions by new vectors
can trigger epidemics
BGSS: native vector
GWSS: new vector
GWSS as a vector in Temecula Valley, CA
-3 years after first symptomatic plant-
Photo A.H. Purcell
Glassy-winged sharpshooter
(GWSS)
Photo R. Krugner
General impact of GWSS introduction into California
GWSS large populations
More X. fastidiosa-vector encounters
?
More successful infections
New vector-pathogen associations?
or
New Xylella strains?
Higher disease incidence
oleander leaf scorch
Strains of Xylella behave differently
in different plant hosts
Example:
Almond group strains (A) do not cause
Pierce’s disease (PD+) in grape
Grape strains (G) cause disease in both
almond (ALS+) and grape (PD+)
Hendson et al. 2001
Grape group
PD3+
PD+
ALS+
Almond group 1
PD3PD-
Almond group 2
PD3+
Grape (G) and almond (A)
strains have different media
requirements in vitro
Almeida and Purcell 2003
Appl. Env. Microbiol. 69:7447
Analysis of strain differences indicate that
Xylella evolves by recombination rather than
mutation.
Refs:
Schuentzel et al. 2005. Appl. Environ Microbiol. 71:3832+
Scally et al. 2005. Appl. Environ Microbiol. 71:8491+
Almeida et al. 2008. Appl. Environ Microbiol. 74:3690+
This implies an ability to rapidly evolve
adaptations to new hosts, as well as virulence
in established hosts.
Southern hemisphere strains of Xf
(citrus ci and coffee co) compared
to northern hemisphere strains
(grape G, almond A, oleander Ol,
Oak)
Almeida et al. AEM 2008
Citrus Variegated Chlorosis (CVC)
www.CNR.Berkeley.EDU/xylella/
Photo by Steve Lindow
Small fruits cause
most damage
o by Steve Lindow
CVC
Citrus variegated chlorosis (CVC) spread
is from infected orchards to nearby orchards
Movements of trees from commercial nurseries
accelerated the spread of CVC throughout Brazil
A specific background of natural vegetation
or weeds does not seem to be a requirement
to sustain CVC epidemics. Citrus alone is
sufficient if vectors are present in or near the
crop.
Numerous vector species occur in Brazil.
Many different mixes of vector species seem
to facilitate spread of CVC
Movement of Xylella within plants and the
role of cell signaling in its biology
Karyn Newman Steve Lindow
UC Berkeley
Intravessel movement
Lateral movement
passive
active
Steve Lindow
X. fastidiosa cells are most commonly
found in modest sized micro-colonies
within xylem vessels
Steve Lindow
Most X. fastidiosa communities are relatively small,
even in symptomatic leaves
Asymptomatic
Symptomatic
Does X. fastidiosa really benefit from crowing into vessels?
Steve Lindow
X. fastidiosa cells that cannot escape crowded vessels die
Vessel 1
Vessel 2
Vessel 3
Cells in top of vessel 2 are escaping into vessel
1 and are green (live), cells in bottom of vessel
2 are not escaping and are red (dead).
Steve Lindow
Plant colonization phase
Extensive vessel colonization
Low cell numbers in most vessels
Disease symptoms may not be present
Insect acquisition phase
Some vessels have high cell numbers
Disease symptoms may be present
Further multiplication in crowded vessels slows
DSF Abundance
Expression of adhesins
Stickiness to Surfaces
Type IV pili
Twitching Motility
Pgl and Eng expression
Pit Membrane Degradation
Gum Production
Ref.: Annu. Rev. Phytopath. 46: 243-271
Xanthomonas campestris pv. campestris
rpfA
rpfB
rpfF
76%
rpfC
70%
rpfA
rpfH
66%
orf orf rpfB
rpfG
58%
//
rpfF
lysS prfB
rpfD orf orf orf
rpfI
recJ
76%
rpfC
rpfE
greA
55%
rpfG
lysS prfB recJ
rpfE
greA
Xylella fastidiosa
rpfB,F
Synthesis of “diffusible signal factor” (DSF)
rpfG,C
DSF perception and signal transduction leading
to virulence trait expression
Wang et al. Molec. Microbiol. 51:903-910 (2004)
X. fastidiosa Decreases Virulence by Coordinating
Virulence Genes in Cell Density-Dependent Fashion
Wild type
rpfF mutant
RpfF- mutants of Xylella fastidiosa are MUCH more virulent
to grape than wild-type strains
Loss of leaves in needle inoculated grapevines
16
Number of lost leaves
14
12
buffer
rpfB- (pKLN 57)
rpfF- (pKLN 61)
rpfF- (pKLN 62)
Temecula
10
8
6
4
2
0
105
114
Days post-inoculation
.
Over-expression of RpfF in Xylella fastidiosa reduces the movement
of the pathogen in the plant and limits disease to site of inoculation
Identification of DSF-producing and DSF-interfering
bacterial strains
Strain
Genus
Origin
Mechanism of DSF Interference
A
B
C
D
E
G
H
J
L
I
U
V
W
X
Y
Z
Paenibacillus
Paenibacillus
Pseudomonas
Staphylococcus
Bacillus
Pseudomonas
Pseudomonas
Pseudomonas
Staphylococcus
Xanthomonas
Xanthomonas
Xanthomonas
Xanthomonas
Xanthomonas
Xanthomonas
Xanthomonas
Grape
Grape
Cabbage
Grape
broccoli
Cabbage
Cabbage
Tomato
Grape
Tomato
Broccoli
Broccoli
Broccoli
Broccoli
Tomato
Grape
Unknown inhibition
Unknown inhibition
Enzymatic digestion
Unknown inhibition
Enzymatic digestion
Enzymatic digestion
Enzymatic digestion
Enzymatic digestion
Unknown inhibition
DSF production
DSF production
DSF production
DSF production
DSF production
DSF production
DSF production
Proportion of grapevines infected
Control of Pierce’s disease of grape by co-inoculating
DFS-producing and DSF-degrading strains into
grape with Xylella fastidiosa
1.0
0.9
0.8
Pathogen plus DSF Inhibitors
Pathogen plus DSF Producers
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0.0
Xf U W X Y V
I
E G C H K
Treatment
J
F
Blockage of DSF production in Xanthomonas suppresses its ability
to inhibit disease in grape when co-inoculated with X. fastidiosa
Over-expression of RpfF in Xylella fastidiosa reduces the
movement of the pathogen in the plant and limits disease to site
of inoculation
Co-inoculating DFS-producing or DSF-degrading strains
into grape with Xylella fastidiosa reduce symptoms of
Pierce’s disease of grape
Blocking DSF production in Xanthomonas suppresses its ability
to inhibit disease in grape when co-inoculated with X. fastidiosa
Other bacteria can interfere with multiplication
and movement of Xylella fastidiosa in grape
through effects on signaling
Could this explain the difficulties of Xylella
establishing in some new regions?
hypopharynx
Temecula
pfB-
epipharynx
rpfF-
Temecula
rpfB-
Signaling (DSF) from Xylella is required for
vector transmission and biofilm formation
in vector food canal (Ref Newman et al. PNAS
rpfF
The density of Xylella within plants determines
the density of diffusible signaling factor (DSF).
The concentration of DSF profoundly affects
the expression levels of genes required for
multiplication, attachment to surfaces,
movement, vector transmission, and virulence.
Nabil Killiny
Rodrigo Almeida
CCGPVE
Almeida Purcell 2006 AESA
Newman et al. 2003 AEM
Xylella cells during early
colonization of insect food canal
Nabil Killiny
Nabil Killiny
Nabil Killiny
Hypothesis for insect colonization
Nabil Killiny
Xylella fastidiosa under in flowing medium in vitro
Notice flick motility and “pseudosocial” behavior
time
Videos from
Harvey Hoch
Cornell Univ.
Why congregate?
Mass attack principle?
Cooperate in bulding biofilm?
(both in plants and insects)
Xylella may have
to survive in
insects for
prolonged periods
Lawn of Xylella cells
Nabil Killiny
Attachment Via CBPs
and HxfB
HxfA
Exopolysaccharides protect
the biofilm
Transition from attached cells
from laterally to polarly attached
Cell binary division allow double
layer biofilm formation
 Persistent in adults
Insect maintains Xf overwinter
Another environmental cue might exist in the insect foregut
to maintain the adhesive (transmissible) state of X. fastidiosa
Xylella eats chitin very efficiently
Nabil Killiny
Steve Lindow
What do we NOT know about the invasive
potential of Xylella fastidiosa?
A lot!!
What governs the survival of Xylella in
natural vegetation?
How important is this for diseases in
crops?
Are new strains of Xylella due to rapid
evolution or an increase in incidence from
“hidden” (rare) to noticeable caused by
changes in environmental conditions?
Do other microbes (including phage) play
a role in the ecology of Xylella?
Implications for quarantine
Expand thinking to include symptomless
hosts
New vectors can rapidly change the
ecology of Xylella.
Proactive approaches are probably
warranted because of the huge number of
vector species.
Acknowledgements
Steve Lindow, UC Berkeley
Subhadeep Chaterjee
Karyn Newman
Clelia Bacarri
Rodrigo Almeida, UC Berkeley
Nabil Killiny
Joao Spotti Lopes
Purcell Lab, UC Berkeley
Rodrigo Almeida
Helene Feil
Barry Hill
Stuart Saunders
Tina Wistrom
Harvey Hoch, Cornell University