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The purpose of the proposed research is to define and control the relationship
between economically important plants and three diverse fungal groups that produce
mycotoxins on corn and forage grasses. One group includes fungi belonging to the
Liseola section of the Fusarium, represented by Fusarium verticillioides (synonym=F.
moniliforme), the second includes the black-spored Aspergillus species, and the third
group consists of obligate grass endophytes of the family Clavicipitaceae, species of
Neotyphodium Glenn, Bacon & Hanlin. There are four objectives: <ol> <li> To decrease the
fumonisin mycotoxin content in corn using biocontrol strategies based on the
endophytic bacterium Bacillus mojavensis and soil fungi such as Trichoderma koningii;
<li> Define the importance of the fumonisins to corn development and plant or fungus
survival, as well as its absorption and translocation in corn; <li> To develop the use
of novel Neotyphodium fungal endophytes in tall fescue for specific reduction of
cattle toxins, and enhanced forage persistence measured by nematode resistance; and
<li> To develop an integrated analysis to determine the extent of the mycotoxin
ochratoxin A production by isolates of the lesser known but widely prevalent
temperate to tropical species Aspergillus niger and A. carbonarius (the Nigri
section) that are endophytic to corn kernels, and to determine any co-endophytic
interactions with F. verticillioides and/or the fumonisins produced during corn
storage.</ol>
Approach: <BR> Objective 1: Pre-harvest control of fumonisin production will be conducted under
controlled conditions of soil moisture, soil inoculum content, over time using corn
seedlings and plants of three cultivars that will be inoculated with and without our
patented bacterial endophyte, Bacillus mojavensis or Trichoderma koningii, each co-
inoculated with strains of Fusarium moniliforme. Fumonisin and fusaric acid, and
plant performance will be determined under the above conditions. <P>Objective 2: To
address the importance of fumonisin on corn development and virulence factor as well
as its translocation by plants, several commercial and experimental corn lines will
be screened for tolerance or sensitivity to fumonisin, and identified tolerance and
sensitive corn lines will be tested for their ability to translocate fumonisin. All
plants will be analyzed for fumonisin content using standard chemical procedures and
fungal biomass determined by real-time PCR. The mechanism of corn tolerance will be
determined with suppression subtraction hybridization of cDNAs and microarray
analyses of sensitive and tolerant lines of corn seedlings. Additionally, a specific
approach will target corn genes involved in sphingolipid biosynthesis, such as ASC1.
<P>Objective 3: Sterile nematode assays, using two or more nematode species, will be
used to determine toxic fractions from the tall fescue endophyte-grass association.
The nematode assay will be used both with wild-type and novel endophyte-infected tall
fescues. Chemical extracts from infected grasses will be used in the bioassay, and
all positive extracts will be separated into specific chemical fractions for chemical
identification of the toxin responsible for nematode toxicity. <P>Objective 4: Survey
both egg and broiler houses (feed, and litter) for the black-spored Aspergillus niger
complex, determine if any of these can produce ochratoxin A on one of several media,
including corn, and identify positive isolates to species using molecular techniques.
Infect corn seedlings with positive A. niger isolates and determine host-parasite
interactions, as well as the effects on toxin accumulation in planta, especially when
co-infected with fumonisin producing and non-producing strains of F. verticillioides.