Plant Pathology
PhD Symposium
Hunt Library |Duke Energy Hall
April 24, 2015
8:00 am - 12:00 noon
Alumnus Enrichment Speaker
Dr. Cheryl Kuske
SPEAKERS:
Jack Bittner
Torey Gonzalez
Kestrel McCorkle
Anna Thomas
1
2
Schedule of Events
MORNING COFFEE
8:00 AM
SESSION II
WELCOME & INTRODUCTIONS
DR. MARC CUBETA
8:25 AM - 8:30 AM
SESSION I
JACK BITTNER, A.L. MILA,
D.M. BENSON, F.J. LOUWS, &
L.R. FISHER
" Evaluation of oxathiapiprolin for control of the
black shank pathogen, Phytophthora nicotianae"
8:30 AM - 9:00 AM
TOREY GONZALEZ, E.L. DAVIS,
J.M. ALONSO, G.A. PAYNE, &
M. ROJAS-PIERCE
“Multiple RNAi Strategies Targeting the Secreted
Chorismate Mutase Reveal its Role in
Suppression of Upstream Defense Signaling in the
Beet Cyct Nematode - Arabidopsis System”
9:00 AM – 9:30 AM
BREAK
KESTREL MCCORKLE, H.D. SHEW,
R.S. LEWIS, M.A. CUBETA, &
I. CARBONE
"Adaptation to quantitative host resistance
by Phytophthora nicotianae"
10:00 AM – 10:30 AM
ANNA THOMAS, P.S. OJIAMBO,
I. CARBONE, H.D. SHEW, &
L.M. QUESADA
“Population biology of Pseudoperonospora
cubensis, the causal agent of cucurbit
downy mildew”
10:30 AM – 11:00 AM
DR. CHERYL KUSKE
“Metagenomics of soil fungi and bacteria in
temperate biomes reveals taxonomic and
functional responses to environmental change”
11:00 AM – 12:00 NOON
Table of Contents
JACK BITTNER, A.L. MILA, D.M. BENSON, F.J. LOUWS, & L.R. FISHER
“Evaluation of oxathiapiprolin for control of the black shank pathogen, Phytophthora nicotianae”
- Page 1 -
TOREY GONZALEZ, E.L. DAVIS, J.M. ALONSO, G.A. PAYNE, & M. ROJAS-PIERCE
“Multiple RNAi Strategies Targeting the Secreted Chorismate Mutase Reveal its Role in
Suppression of Upstream Defense Signaling in the Beet Cyct Nematode – Arabidopsis System”
- Page 2 -
KESTREL MCCORKLE, H.D. SHEW, R.S. LEWIS, M.A. CUBETA, & I. CARBONE
“Adaptation to quantitative host resistance by Phytophthora nicotianae”
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ANNA THOMAS, P.S. OJIAMBO, I. CARBONE, H.D. SHEW & L.M. QUESADA
“Population biology of Pseudoperonospora cubensis, the causal agent of cucurbit downy mildew”
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Evaluation of Oxathiapiprolin for Control of the Black Shank Pathogen, Phytophthora
nicotianae
J. BITTNER1, A.L. Mila1, D.M. Benson1, F.J. Louws1 and L.R. Fisher2
1
2
Department of Plant Pathology, North Carolina State University, Raleigh, NC 27695
Department of Crop Science, North Carolina State University, Raleigh, NC 27695
Black shank, caused by Phytophthora nicotianae, is one of the most destructive soilborne diseases of tobacco
(Nicotiana tabacum L.). Chemical control is an effective management strategy when combined with other
practices. Oxathiapiprolin is a new fungicide with activity against oomycetes and in-vitro assays were
conducted to determine its effect on P. nicotianae isolates from tobacco in North Carolina. The efficacy of
oxathiapiprolin on mycelial growth, sporangia production, zoospore motility, and zoospore germination was
assessed. Isolates were highly sensitive to oxathiapiprolin. EC50 values ranged from 0.0039 to 0.0049 µg
a.i./ml for mycelial growth, 0.00052 to 0.00081 µg a.i./ml for sporangia production, 0.0035 to 0.0051 µg
a.i./ml for encysted zoospore germination, and 0.0055 to 0.0166 µg a.i./ml for zoospore motility. In addition,
66 isolates from tobacco and ornamental plants, were examined for their sensitivity to oxathiapiprolin at 1 µg
a.i./ml and mycelial growth was observed in one tobacco isolate. The efficacy of oxathiapiprolin was also
investigated in studies conducted in 2012 and 2013 in three naturally infested fields in North Carolina.
Treatments were applied to the soil on the day of transplanting tobacco to the field, 2 to 4 weeks after
transplant, and 7 to 9 weeks after transplant. Treatments with three applications of oxathiapiprolin at a rate of
9.6 fl oz/a resulted in from 55 to 83% control compared to the untreated checks, whereas three applications of
mefenoxam at 16 fl oz/a resulted in 3 to 55% control. These results demonstrate that oxathiapiprolin is
effective in the control of black shank.
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Multiple RNAi Strategies Targeting the Secreted Chorismate Mutase Reveal its Role in
Suppression of Upstream Defense Signaling in the Beet Cyst Nematode - Arabidopsis System
T.M. GONZALEZ1, E.L. Davis1, J.M. Alonso2, G.A. Payne1, M. Roja-Pierce2, R. Strauch3 and Sirius Li3
1
Department of Plant Pathology, North Carolina State University, Raleigh, NC 27695
Department of Plant & Microbial Biology, North Carolina State University, Raleigh, NC 27695
3
Plants for Human Health Institute, North Carolina State University, Kannapolis, NC 28081
2
Plant-parasitic nematodes are microscopic round-worms living in the soil and they use a stylet to inject
effector proteins into living plant cells, resulting in suppression of defenses and formation of a feeding site.
While most effectors act through protein-protein interactions, the secreted chorismate mutase (CM) appears to
act by ‘metabolic manipulation’. Interestingly, plant pathogens from nearly every kingdom appear to use
some form of secreted CM, suggesting that different pathogens have converged upon a common point of
vulnerability in plants. A better understanding of this mechanism could help produce engineered plants
resistant to this effector. The established model is that CM is able to compete with the plastid for substrate,
and so prevent Isochorismate Synthase1 (ICS1) from converting chorismate to salicylic acid, a major plant
defense hormone. However, we explore an alternative model where metabolic changes caused by CM are
translated into suppression of plant defense signaling. Since ICS1 expression is induced by upstream signals
such as ROS and calcium, interference with defense signaling could make ICS1 rather than chorismate the
limiting factor for SA production. Quantitative PCR was used to measure pathogenesis related genes as SA
markers, and were found to be inversely correlated with CM1 expression in both Wt and RNAi lines,
consistent with a role in defense suppression. ICS1 was also inversely correlated with CM1 expression,
suggesting a role in upstream defense suppression. This data, along with the wide-spread presence of secreted
chorismate mutases suggests an uncharacterized connection between metabolism and defense signaling
apparently vulnerable to manipulation.
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Adaptation to Quantitative Host Resistance by Phytophthora nicotianae
K. McCORKLE1, H.D. Shew1, R. Lewis2, M. Cubeta1,3, and I. Carbone1,3
1
Department of Plant Pathology, North Carolina State University, Raleigh, NC 27695
Department of Crop Science, North Carolina State University, Raleigh, NC 27695
3
Center for Integrated Fungal Research, North Carolina State University, Raleigh, NC 27695
2
Black shank, caused by Phytophthora nicotianae, is an important disease of tobacco worldwide. Host
resistance is the easiest and most cost effective tool for disease control. Since resistance genes exert selection
pressure on the pathogen, phenotypic and genotypic shifts in the population may be observed. The objectives
of this study were to determine genetic variability of the pathogen across tobacco growing regions in the
United States and how isolates adapt to varying sources and levels of partial resistance. To determine
pathogen variability, five genes were sequenced in isolates collected from KY, VA, and GA and used to find
single nucleotide polymorphisms (SNPs) among individuals. The highest numbers of SNPs were found in the
COXII region in isolates from VA, and in the EF region in isolates from KY and GA. Genotypic variability
was high, with multiple genotypes being recovered in each state. The level of detected genetic variation was
highest in GA followed by VA and KY. For adaptation experiments, greenhouse plants with resistance from
Florida 301, Beinhart 1000, or the Wz gene were inoculated with race 0 and race 1 isolates for six continuous
generations. Adapted isolates were compared across generations for each variety and pathogen race
combination. An increase in pathogen aggressiveness was observed over generations for both races on
multiple varieties. More aggressive isolates had decreased incubation periods and increased root rot.
Understanding how P. nicotianae utilizes genotypic variability to adapt to host resistance can result in better
recommendations for variety rotation to effective disease management.
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Population Biology of Pseudoperonospora cubensis, the Causal Agent of Cucurbit Downy
Mildew
A. THOMAS1,2, P.S. Ojiambo1,2, I. Carbone1,2, H.D. Shew1 and L.M. Quesada-Ocampo1.
1
2
Department of Plant Pathology, North Carolina State University, Raleigh, NC 27695
Center for Integrated Fungal Research, North Carolina State University, Raleigh, NC 27695
Cucurbit downy mildew, considered as the most economically important disease of cucurbits, resurged in the
United States in 2004 overcoming host resistance that had been effective for more than 40 years. Introduction
of a new lineage, a new pathotype or new genetic recombinants of the causal pathogen, Pseudoperonospora
cubensis, have been suggested as the potential reasons for the resurgence of cucurbit downy mildew. The
potential for sexual reproduction in P. cubensis, the current pathotype structure of the pathogen in the United
States and the genetic basis for host specialization in P. cubensis were investigated. An isolate-pairing assay
using known mating type testers from Israel revealed, for the first time, the presence of A1 and A2 mating
types of P. cubensis in the United States. Inoculation experiments using a total of nineteen isolates on nine
host differentials; Cucumis spp., Citrullus lanatus, Cucurbita spp., Luffa cylindrica, Lagenaria siceraria,
Trichosanthes cucumerina, revealed the presence of new pathotypes in the United States, namely III and VI,
that were previously known to exist only in Asia. Comparative genomic analysis of a set of nine isolates of P.
cubensis collected from diverse set of cucurbit hosts revealed the presence of two distinct evolutionary host
specific lineages. The association of these host specific lineages with mating types and pathotypes was also
investigated and the results from this association analysis may further facilitate our understanding of the
resurgence of cucurbit downy mildew in the United States.
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