Submitted to: Physiological and Molecular Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/27/2016
Publication Date: 4/28/2016
Citation: Mellon, J.E., Mattison, C.P., Grimm, C.C. 2016. Identification of hydrolytic activities expressed by Aspergillus flavus grown on cotton carpel tissue. Physiological and Molecular Plant Pathology. 94:67-74.
Interpretive Summary: Aflatoxin is a very potent carcinogen and toxin that is produced by the fungus Aspergillus flavus. When this fungus infects target crops (corn, cotton, peanuts, tree nuts), the developing seed can be contaminated with this toxin, rendering the product unfit for additional agricultural uses. Xyloglucans, xylans and pectins are plant polysaccharides that are important structural components of plant cell walls. Cottonseed contains high concentrations of xylans and is particularly susceptible to aflatoxin contamination. The fungus is capable of producing a number of hydrolytic enzymes, including xylanase and pectinase that are crucial to breaking down xylans and pectins. Cotton fruits (bolls) are divided into sections by a tough fibrous defensive tissue called carpellary tissue. Carpel tissue inhibits the spread of potential microbial pathogens such as A. flavus from spreading between sections of the cotton fruit. These hydrolase activities help the fungus breach plant cell walls and gain access to potential nutrient sources in the seed that include free sugars, polysaccharides, lipids, and storage proteins. In order to better understand the roles of these enzymes in fungal virulence, A. flavus was grown in a medium containing cotton carpel tissue as a carbon substrate. The fungus secreted at least two different xylanase activities into the growth medium, as well as copious amounts of pectinase. These A. flavus-produced hydrolases assist the fungus in maceration of plant cell walls. This research will benefit oilseed breeders, producers and pathologists, and will aid in the formulation of methods to prevent aflatoxin contamination of target crops.
Technical Abstract: Aspergillus flavus secreted at least two endoxylanase activities, two esterase activities and a pectolytic activity when grown on a medium containing cotton carpel tissue as a sole carbon source. A concentrated sample of A. flavus growth medium (6-day) was subjected to gel filtration chromatography on a BioGel P-30 column. A major endoxylanase activity was separated from the other fungal secreted proteins. In a subsequent step, remaining fungal secreted proteins were partially resolved by gel filtration chromatography on a BioGel P-60 column. A secondary endoxylanase activity was partially resolved from a major esterase activity. Denatured gel electrophoresis indicated this esterase activity was associated with a protein in the 24 kD range and may represent an acetylxylan esterase. The pectolytic activity appears to be associated with a protein in the 35 to 40 kD range. In addition, multiple additional proteins with molecular weights in the 40 to 70 kD range were present in the harvested fungal growth medium. These enzymes probably assist A. flavus in the maceration of plant cell walls, allowing for pathogenic entry and accession of host nutrient resources. Both pectolytic and xylanolytic hydrolases appear to be important A. flavus virulence factors.