Molecular detection of pathogenic Fusarium species in roots and stalks of maize plants with or without transgenic resistance to corn rootworm

Fusarium species are among most common fungal pathogens of maize, causing root rot, stalk rot, stalk lodging and ear rot. Fusarium verticillioides and F. graminearum are two of the most prevalent stalk rot fungi in Nebraska and Iowa. Corn root worms (CRW) are the most costly pests of maize plants, causing damage to both above- and below-ground parts of the plant. CRW feeding on roots creates good invading sites for different fungal species. The goal of this research was to determine whether use of transgenic CRW-resistant maize hybrids will have reduced levels of Fusarium colonization of roots and stalks compared to their near-isogenic susceptible hybrids. Experiments were conducted at two locations each in 2007 and 2008 in fields with high populations of Western corn root worms, (Diabrotica virgifera LeConte): Bruner farm near Ames, IA (2007), ISU Southeast Research Farm near Crawfordsville, IA (2007-2008), and Agricultural Research and Development Center, Univ. of NE near Mead, Nebraska (2007-2008). Commercially available hybrids with CRW-resistance transgenes along with their near-isogenic CRW-susceptible hybrids were planted. The CRW events include DAS-59122 (Herculexy Xtra), MON863 (Yieldgard Plusy), MON88107 (Yieldgard VT Tripley) and MIR604 (Agrisurey RW/CB/LL). In 2008, insecticidal seed treatment also was tested for its effect on CRW feeding and subsequent Fusarium colonization. Sampling was done on roots in mid-July and stalks in mid-September in 2007 and in 2008 mid-September roots were also sampled. Root colonization was assessed by dilution plating of dried ground tissue, and by real-time polymerase chain reaction (PCR). v Primers and fluorogenic probes, specific to mycotoxin biosynthesis genes in F. verticillioides and F. graminearum were used to quantify root and stalk colonization. Standard calibration curves of both species showed linear correlation (r2=0.99) between the fungal genomic DNA and the threshold cycle values. There were highly significant differences in colonization between resistant and near isogenic susceptible hybrids for events MON 88017 and MIR 604 in Nebraska July roots in 2007. F. verticillioides was at high levels in Nebraska whereas F. graminearum was more common in Iowa. Stalks from both locations were highly colonized with F. graminearum. The PCR results were compared to a microbiological dilution plating method. In 2007 the results of the two methods correlated well but in 2008 they did not follow the same pattern. In dilution plating results we could consistently see significantly lower Fusarium CFU/g tissue in hybrids with CRW resistance than in near-isogenic CRW susceptible hybrids. Insecticidal seed treatment did not have a significant effect on level of colonization of fungal species measured by PCR or dilution plating results. CRW feeding on maize roots sometimes enhanced the infection of roots by Fusarium species and transgenic CRW resistant hybrids suffered less colonization than susceptible hybrids in CRW infested fields. However, the interaction between Fusarium spp and CRW varied in their degree among Fusarium species. Known maize pathogens F. verticillioides and F. graminearum were not consistently affected by CRW resistance as much as the total Fusarium population. The impact of this interaction on stalk rots is not clearly demonstrated