B.H. de Haas

Potential of fungal antagonists for biocontrol of Fusarium spp. in wheat and maize through competition in crop debris

Abstract Pathogenic Fusarium spp. cause head blight in wheat or ear rot in maize leading to yield losses and also a reduction in quality due to mycotoxin contamination of the grain. Infected crop residues are the main inoculum source for epidemics. Saprophytic fungi, obtained from cereal tissues or necrotic tissues of other crops, were screened for their ability to colonise wheat straw and maize stalks and to suppress sporulation of pathogenic Fusarium spp. Results of bio-assays conducted under controlled conditions were variable among Fusarium spp. and host substrates for most antagonists tested, such as yeasts, Trichoderma spp. and non-pathogenic Fusarium spp. Isolates of Clonostachys rosea consistently suppressed sporulation of F. culmorum and F. graminearum on wheat straw, and of F. culmorum, F. graminearum, F. proliferatum and F. verticillioides on maize stalks. Isolates of C. rosea, C. cladosporioides and F. equiseti were applied to pieces of maize stalks or flowering ears in preliminary experiments conducted under field conditions. The colonisation of stalk pieces by pathogenic Fusarium spp. was assessed after 9 months. Colonisation of stalk pieces by pathogenic Fusarium spp. was significantly reduced at several sampling dates. However, results obtained with the antagonists were not consistent for all sampling dates and between experiments.

Biological Control of Botrytis cinerea in Cyclamen with Ulocladium atrum and Gliocladium roseum Under Commercial Growing Conditions

ABSTRACT The effect of treatments with conidial suspensions of Ulocladium atrum and Gliocladium roseum on leaf rot of cyclamen caused by Botrytis cinerea was investigated under commercial greenhouse conditions. Spraying U. atrum (1 x 10(6) conidia per ml) or G. roseum (2 x 10(6) conidia per ml and 1 x 10(7) conidia per ml) at intervals of 2 to 3 weeks during the production period and spraying U. atrum (1 x 10(6) conidia per ml) at intervals of 4 to 6 weeks resulted in a significant reduction of natural infections of petioles by B. cinerea. U. atrum or G. roseum (1 x 10(7)conidia per ml) was as effective as the standard fungicide program. B. cinerea colonized senesced leaves within the plant canopy and infected adjacent petioles and leaves later. The antagonists colonized senesced leaves and reduced B. cinerea development on these leaves. Thus, the inoculum potential on petioles adjacent to necrotic leaf tissues was reduced. The fate of U. atrum conidia on surfaces of green cyclamen leaves during a 70-day period after application was studied. The number of conidia per square centimeter of leaf surface remained relatively constant during the entire experiment. Sixty percent of the conidia sampled during the experiments retained the ability to germinate. When green leaves were removed from the plants to induce senescence and subsequently were incubated in a moist chamber, U. atrum colonized the dead leaves. Senesced leaves also were colonized by other naturally occurring fungi including B. cinerea. On leaves treated with U. atrum from all sampling dates, sporulation of B. cinerea was significantly less as compared with the untreated control. Our results indicate that early applications of U. atrum before canopy closure may be sufficient to achieve commercially satisfactory control of Botrytis leaf rot in cyclamen.

Population Dynamics of Fusarium spp. and Microdochium nivale in Crops and Crop Residues of Winter Wheat

ABSTRACT Naturally occurring populations of Fusarium avenaceum, F. culmorum, F. graminearum, F. poae, and Microdochium nivale were studied in two field experiments from anthesis in June 2003 until harvest in crops of winter wheat, and subsequently during 10 months after harvest until June 2004 on their residues exposed on the soil surface under field conditions. The dynamics of the different pathogens were estimated by quantifying the amount of DNA present in wheat tissues using TaqMan-polymerase chain reaction. While colonization of grain by Fusarium spp. and M. nivale was low, high amounts of DNA of F. avenaceum, F. graminearum, and F. culmorum were found in ear residues, internodes, and nodes of the mature crop. Amounts of DNA of pathogens decreased significantly during the following 10 months in residues of internodes and nodes, but not in residues of stem bases. Knowledge on population dynamics of pathogens will help to develop preventive measures aimed at reduction of inoculum sources of head blight pathogens.

Quantification of Mycelium of Botrytis spp. and the Antagonist Ulocladium atrum in Necrotic Leaf Tissue of Cyclamen and Lily by Fluorescence Microscopy and Image Analysis

ABSTRACT A technique was developed to localize and quantify the internal mycelial colonization of necrotic leaf tissue of cyclamen (Cyclamen persicum) or lily (Lilium) by pathogenic Botrytis spp. and the antagonist Ulocladium atrum. This technique allows investigation of competitive substrate colonization by both fungi, which is a key process for biological control of Botrytis spp. by U. atrum. A combination of differential fluorescent labeling and image analysis was applied on cryostat sections of necrotic leaf tissue. Botrytis mycelium was labeled specifically by indirect immunofluorescence using a monoclonal antibody specific for Botrytis spp. And an antimouse fluorescein conjugate. Wheat germ agglutinin conjugated to the fluorochrome TRITC was used to label mycelium of both fungi. Image analysis was used to measure the relative surface area of the cryostat section covered by fluorescing hyphae of Botrytis spp. and by fluorescing hyphae of both fungi. A mathematical conversion was derived and used to calculate the relative mycelial volume of each fungal species in the necrotic tissue based on the measured relative surface areas. Temporal aspects of substrate colonization were studied in a short time series. An analysis of components of variance provided insight into spatial colonization patterns for the fungal species involved and allowed the design of efficient sampling strategies for future experiments.

Comparative analysis of the role of substrate specificity in biological control of Botrytis elliptica in lily and B. cinerea in cyclamen with Ulocladium atrum

Biological control of Botrytis spp. by the fungal antagonist Ulocladium atrum is based on their interaction in plant tissue. U. atrum is effective against B. cinerea in commercial cyclamen crops but not effective against B. elliptica in lily crops. Based on the necrotrophic nature of the Botrytis spp. and the saprophytic nature of U. atrum it is hypothesised, and experimentally confirmed, that the interaction between Botrytis spp. and U. atrum, resulting in a biocontrol effect, only takes place in necrotic plant tissue. The role of necrotic tissue in the epidemiology of B. cinerea in cyclamen and B. elliptica in lily was found to be different. Removal of symptomless senescing leaves resulted in a significant reduction of the area under the disease severity progress curve (AUDPC) for B. cinerea in cyclamen but had no effect on the disease severity in lily. U. atrum applications significantly reduced B. cinerea AUDPC values in cyclamen but were less efficient than the removal of senescing leaves. In lily, disease severity was not affected by applications of U. atrum. It is concluded that necrotic cyclamen tissue, not killed by B. cinerea, plays an important role in the onset of disease. Colonisation of this tissue by U. atrum prevents saprophytic colonisation of those leaves by B. cinerea. In contrast, conidia of B. elliptica directly infect healthy lily leaf tissue. U. atrum applications aimed at blocking the infection pathway from a saprophytic base are therefore not effective against B. elliptica. Control options based on competitive interactions in and around B. elliptica lesions resulted in a reduced production of conidia by B. elliptica but proved ineffective against disease development. The potential of U. atrum as a biocontrol agent against Botrytis spp. and possibly against other necrotrophs appears to be determined by the competitive saprophytic ability of the antagonist in mutual substrates of pathogen and antagonist and by the role of these substrates in disease epidemiology.