A histological study of Aspergillus flavus colonization of wound inoculated maize kernels of resistant and susceptible maize hybrids in the field

Authors: Windham, Gary; Williams, William; Mylroie, John; REID, CEDRIC - Mississippi State University; Womack, Erika

Submitted to: Frontiers in Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/25/2017
Publication Date: 2/15/2018
Citation: Windham, G.L., Williams, W.P., Mylroie, J.E., Reid, C., Womack, E.D. 2018. A histological study of Aspergillus flavus colonization of wound inoculated maize kernels of resistant and susceptible maize hybrids in the field. Frontiers in Microbiology. 9:799. https://doi.org/10.3389/fmicb.2018.00799.
DOI: https://doi.org/10.3389/fmicb.2018.00799

Interpretive Summary: Aspergillus flavus colonization in developing kernels of corn single-cross hybrids resistant and susceptible to aflatoxin accumulation was determined in the field over three growing seasons. Plants were hand pollinated, and individual kernels were inoculated with a needle dipped in a suspension of A. flavus spores 21 days after pollination. Kernels were harvested at 1-2 day intervals from 1 to 21 days after inoculation (DAI). Histological slides are made of thin sections of the kernel tissues and stained with Toluidine blue. Kernels were also collected for aflatoxin analyses. At 2 DAI, A. flavus hyphae were observed among endosperm cells in the susceptible hybrid, but colonization of the endosperm in the resistant hybrid was limited to the wound site of the resistant hybrid. Sections of the scutellum of the susceptible hybrid was colonized by A. flavus by 5 DAI. Fungal growth was slower in the resistant hybrid compared to the susceptible hybrid. By 10 DAI, A. flavus had colonized a large section of the embryo in the susceptible hybrid; whereas in the resistant hybrid, approximately half of the endosperm had been colonized and very few cells in the embryo were colonized. Fungal colonization in some of the kernels of the resistant hybrid was slowed in the aleurone or at the endosperm-scutellum interface. In wounded kernels with an intact aleurone, the fungus spread around the kernel between the pericarp and aleurone layer with minimal colonization of the endosperm. Aflatoxin was first detected in susceptible kernel tissues 8 DAI in 2013 and 2014. Resistant kernels had lower aflatoxin accumulation both years. Our study found differential A. flavus colonization of susceptible and resistant kernel tissues, and that the aleurone and the outer layer of the scutellum slowed the rate of colonization by A. flavus. Future studies should provide information on the basis of resistance in these corn tissues.

Technical Abstract: Aspergillus flavus colonization in developing kernels of maize single-cross hybrids resistant (Mp313E x Mp717) and susceptible (GA209 x T173) to aflatoxin accumulation was determined in the field over three growing seasons (2012-2014). Plants were hand pollinated, and individual kernels were inoculated with a needle dipped in a suspension of A. flavus conidia 21 days after pollination. Kernels were harvested at 1-2 day intervals from 1 to 21 days after inoculation (DAI). Kernels were fixed in FAA, dehydrated, embedded in paraffin, sectioned, and stained with Toluidine blue. Kernels were also collected for aflatoxin analyses in 2013 and 2014. At 2 DAI, A. flavus hyphae were observed among endosperm cells in the susceptible hybrid, but colonization of the endosperm in the resistant hybrid was limited to the wound site of the resistant hybrid. Sections of the scutellum of the susceptible hybrid was colonized by A. flavus by 5 DAI. Fungal growth was slower in the resistant hybrid compared to the susceptible hybrid. By 10 DAI, A. flavus had colonized a large section of the embryo in the susceptible hybrid; whereas in the resistant hybrid, approximately half of the endosperm had been colonized and very few cells in the embryo were colonized. Fungal colonization in some of the kernels of the resistant hybrid was slowed in the aleurone layer or at the endosperm-scutellum interface. In wounded kernels with intact aleurone layers, the fungus spread around the kernel between the pericarp and aleurone layer with minimal colonization of the endosperm. Aflatoxin was first detected in susceptible kernel tissues 8 DAI in 2013 and 2014. Resistant kernels had lower aflatoxin accumulation both years. Our study found differential A. flavus colonization of susceptible and resistant kernel tissues, and that the aleurone and the outer layer of the scutellum slowed the rate of colonization by A. flavus.