RESUMEN
Fusarium oxysporum f. sp. spinaciae, causal agent of spinach Fusarium wilt, is an important soilborne pathogen in many areas of the world where spinach is grown. The pathogen is persistent in acid soils of maritime western Oregon and Washington, the only region of the United States suitable for commercial spinach seed production. A TaqMan real-time polymerase chain reaction (PCR) assay was developed for rapid identification and quantification of the pathogen, based on sequencing the intergenic spacer (IGS) region of rDNA of isolates of the pathogen. A guanine single-nucleotide polymorphism (G SNP) was detected in the IGS sequences of 36 geographically diverse isolates of F. oxysporum f. sp. spinaciae but not in the sequences of 64 isolates representing other formae speciales and 33 isolates representing other fungal species or genera. The SNP was used to develop a probe for a real-time PCR assay. The real-time PCR assay detected F. oxysporum f. sp. spinaciae at 3-14,056 CFU/g of soil in 82 soil samples collected over 3 years from naturally infested spinach seed production sites in western Washington, although a reliable detection limit of the assay was determined to be 11 CFU/g of soil. A significant (P < 0.05), positive correlation between enumeration of F. oxysporum on Komada's agar and quantification of the pathogen using the TaqMan assay was observed in a comparison of 82 soil samples. Correlations between pathogen DNA levels, Fusarium wilt severity ratings, and spinach biomass were significantly positive for one set of naturally infested soils but not between pathogen DNA levels, wilt incidence ratings, and spinach biomass for other soil samples, suggesting that soilborne pathogen population is not the sole determinant of spinach Fusarium wilt incidence or severity. The presence of the G SNP detected in one isolate of each of F. oxysporum ff. spp. lageneriae, lilii, melongenae, and raphani and reaction of the real-time PCR assay with 16 of 22 nonpathogenic isolates of F. oxysporum associated with spinach plants or soil in which spinach had been grown potentially limits the application of this assay. Nonetheless, because all isolates of F. oxysporum f. sp. spinaciae tested positive with the real-time PCR assay, the assay may provide a valuable means of screening for resistance to Fusarium wilt by quantifying development of the pathogen in spinach plants inoculated with the pathogen.
RESUMEN
The maize stalk rot complex is comprised of several fungal pathogens, including Gibberella zeae, Colletotrichum graminicola, Stenocarpella maydis, and several members of the genus Fusarium. The European corn borer (ECB) (Ostrinia nubilalis) can contribute to stalk rot development by creating entry wounds and by serving as a vector of some stalk rot pathogens, particularly Fusarium verticillioides. Transgenic insect protection of maize hybrids with insecticidal proteins derived from Bacillus thuringiensis greatly reduces ECB injury and may therefore alter the species composition and diversity of the stalk rot complex. Field experiments were conducted in 1998, 1999, and 2000 to compare the species composition and diversity of fungi infecting stalks of Bt and non-Bt maize hybrids. Hybrids representing five Bt types (or "events") and their near-isogenic non-Bt counterparts were subjected to manual and natural infestations with ECB larvae. Stalk tissue samples were cultured to determine fungal species composition. At least one species was isolated from nearly every stalk and from both diseased and symptomless tissues. G. zeae was the most common species in 1998 and 1999, but C. graminicola was most common in 2000. The mean proportions of stalks infected with F. verticillioides, F. proliferatum, and F. subglutinans were significantly lower in Bt hybrids than in non-Bt hybrids in 2 of the 3 years. Conversely, the mean proportion of stalks infected with G. zeae was higher in some Bt hybrids than their non-Bt counterparts in two of the three years. F. verticillioides was more likely to be isolated from ECB-injured tissue, whereas G. zeae and C. graminicola were more likely to be isolated from tissue not associated with ECB injury. The overall species diversity of the stalk rot complex was lower in some Bt hybrids compared with their non-Bt counterparts in 1998 and 1999. ECB activity appeared to alter fungal species composition in stalks, reflecting the association between ECB injury and specific stalk rot pathogens, particularly F. verticillioides. The species composition of fungi infecting stalks of Bt hybrids differed from that of non-Bt hybrids, but the implications of this result are not yet clear.
RESUMEN
Stalk rots, caused by a complex of fungal species, are among the most widespread and destructive diseases of maize. Larvae of the European corn borer (ECB) (Ostrinia nubilalis) promote stalk rot development by creating entry points for fungi, serving as vectors of pathogens, and causing physiological stress that may predispose plants to stalk decay. Field experiments were conducted in 1998, 1999, and 2000 to determine whether the use of transgenic Bt hybrids expressing insecticidal proteins would influence stalk rot symptoms (pith disintegration, pith discoloration, and lodging). Five hybrids representing different Bt types (or "Bt events") (176, BT11, MON810, DBT418, and CBH351) were paired with their near-isogenic, non-Bt counterparts and subjected to treatments of manual and natural infestation with ECB larvae. Manual infestation resulted in significantly more ECB tunneling than natural infestation in 1998 and 1999 and significantly more lodging in 1998. There were significant linear correlations between ECB injury and stalk rot symptoms in non-Bt hybrids in 1998 and 1999, but not in 2000. A standard foliar insecticide treatment for ECB did not significantly affect stalk rot symptoms. In 1998, Bt hybrids had significantly less ECB tunneling, stalk discoloration, pith disintegration, and lodging compared with non-Bt hybrids, but these effects depended upon the Bt event and the infestation treatment. Similar but less pronounced effects of Bt events were observed in 1999. The 2000 results were more variable; the amount of pith disintegration was significantly lower but discoloration was significantly higher in the BT11 hybrid compared with its non-Bt counterpart, and the amount of lodging was significantly higher in the event 176 hybrid compared with its non-Bt counterpart. The ratio of stalk strength to grain weight did not consistently differ between Bt and non-Bt hybrids. These results indicate that, although specific Bt events in some years may cause reductions in stalk rot, the overall effect of Bt transformation on stalk rot occurrence is highly variable.