Heinz-Wilhelm Dehne

Recent advances in sensing plant diseases for precision crop protection

Near-range and remote sensing techniques have demonstrated a high potential in detecting diseases and in monitoring crop stands for sub-areas with infected plants. The occurrence of plant diseases depends on specific environmental and epidemiological factors; diseases, therefore, often have a patchy distribution in the field. This review outlines recent insights in the use of non-invasive optical sensors for the detection, identification and quantification of plant diseases on different scales. Most promising sensor types are thermography, chlorophyll fluorescence and hyperspectral sensors. For the detection and monitoring of plant disease, imaging systems are preferable to non-imaging systems. Differences and key benefits of these techniques are outlined. To utilise the full potential of these highly sophisticated, innovative technologies and high dimensional, complex data for precision crop protection, a multi-disciplinary approach—including plant pathology, engineering, and informatics—is required. Besides precision crop protection, plant phenotyping for resistance breeding or fungicide screening can be optimized by these innovative technologies.

Hyperspectral imaging for small-scale analysis of symptoms caused by different sugar beet diseases.

Hyperspectral imaging (HSI) offers high potential as a non-invasive diagnostic tool for disease detection. In this paper leaf characteristics and spectral reflectance of sugar beet leaves diseased with Cercospora leaf spot, powdery mildew and leaf rust at different development stages were connected. Light microscopy was used to describe the morphological changes in the host tissue due to pathogen colonisation. Under controlled conditions a hyperspectral imaging line scanning spectrometer (ImSpector V10E) with a spectral resolution of 2.8 nm from 400 to 1000 nm and a spatial resolution of 0.19 mm was used for continuous screening and monitoring of disease symptoms during pathogenesis. A pixel-wise mapping of spectral reflectance in the visible and near-infrared range enabled the detection and detailed description of diseased tissue on the leaf level. Leaf structure was linked to leaf spectral reflectance patterns. Depending on the interaction with the host tissue, the pathogens caused disease-specific spectral signatures. The influence of the pathogens on leaf reflectance was a function of the developmental stage of the disease and of the subarea of the symptoms. Spectral reflectance in combination with Spectral Angle Mapper classification allowed for the differentiation of mature symptoms into zones displaying all ontogenetic stages from young to mature symptoms. Due to a pixel-wise extraction of pure spectral signatures a better understanding of changes in leaf reflectance caused by plant diseases was achieved using HSI. This technology considerably improves the sensitivity and specificity of hyperspectrometry in proximal sensing of plant diseases.

Epidemiology of Fusarium Infection and Deoxynivalenol Content in Winter Wheat in the Rhineland, Germany

Details of our long-term research programme concerning the epidemiology of Fusarium spp. and mycotoxin production are summarized. Evaluation of the occurrence of Fusarium spp., mainly on winter wheat (Triticum aestivum), was carried out by investigating Fusarium infection and mycotoxin contamination. Two to 15% of grains were infested during 1995–1998 at three climatologically differing localities of the Rhineland, Germany. Disease progress was accelerated by rainfall during the flowering season. The species most frequently isolated were Fusarium avenaceum, F. poae, F. culmorum and F. graminearum. The mean deoxynivalenol (DON) content varied from 19μgkg−1 (1995) to 310μgkg−1 (1998) and was not always correlated with disease severity. Organic farming systems showed lower rates of infection with ear blight and lower mycotoxin contamination than conventional farming systems.

Effect of downy mildew development on transpiration of cucumber leaves visualized by digital infrared thermography.

ABSTRACT Disease progress of downy mildew on cucumber leaves, caused by the obligate biotrophic pathogen Pseudoperonospora cubensis, was shown to be associated with various changes in transpiration depending on the stage of pathogenesis. Spatial and temporal changes in the transpiration rate of infected and noninfected cucumber leaves were visualized by digital infrared thermography in combination with measurements of gas exchange as well as microscopic observations of pathogen growth within plant tissue and stomatal aperture during pathogenesis. Transpiration of cucumber leaf tissue was correlated to leaf temperature in a negative linear manner (r = -0.762, P < 0.001, n = 18). Leaf areas colonized by Pseudoperonospora cubensis exhibited a presymptomatic decrease in leaf tem perature up to 0.8 degrees C lower than noninfected tissue due to abnormal stomata opening. The appearance of chlorosis was associated with a cooling effect caused by the loss of integrity of cell membranes leading to a larger amount of apoplastic water in infected tissue. Increased water loss from damaged cells and the inability of infected plant tissue to regulate stomatal opening promoted cell death and desiccation of dying tissue. Ultimately, the lack of natural cooling from necrotic tissue was associated with an increase in leaf temperature. These changes in leaf temperature during downy mildew development resulted in a considerable heterogeneity in temperature distribution of infected leaves. The maximum temperature difference within a thermogram of cucumber leaves allowed the discrimination between healthy and infected leaves before visible symptoms appeared.

Global crop production and the efficacy of crop protection - current situation and future trends

Actual and potential crop losses of eight major food and cash crops have been estimated by evaluating data from literature and field experiments. Total losses were calculated from yield reductions due to pathogens, animal pests and weeds on a regional, continental and global level. Since 1965, worldwide production of most crops has increased considerably. Simultaneously, crop losses in wheat, potatoes, barley and rice increased by 4 to 10 percent, in maize, soybean, cotton and coffee losses remained unchanged or slightly decreased. The efficacy of crop protection practices was calculated as the percentage of potential losses prevented by control. The efficacy is highest in cotton (55 percent), it reaches only 34 to 38 percent in the food crops rice, wheat and maize. The variability among cropping areas is high: In Western Europe, 61 percent of potential crop losses is prevented, in North America and Oceania 44, in all other regions 38 percent. Due to the small share of Western Europe in worldwide production of 8 percent, the efficacy of actual crop protection worldwide is only 40 percent.In view of population growth and rising food demand crop production has to be increased substantially. As potential loss rates often increase with attainable yields high productivity largely depends on effective crop protection management. Scenarios for the production of food crops by the year 2025 in developed and in developing countries are given. Recent and future developments in crop protection can contribute to establish sustainability in agriculture and to preserve natural resources. However, although effective control methods have been developed for most biotic yield constraints, the use of crop protection products is regulated by economic considerations rather than by food demand.

Next Generation Sequencing Based Transcriptome Analysis of Septic-Injury Responsive Genes in the Beetle Tribolium castaneum

Beetles (Coleoptera) are the most diverse animal group on earth and interact with numerous symbiotic or pathogenic microbes in their environments. The red flour beetle Tribolium castaneum is a genetically tractable model beetle species and its whole genome sequence has recently been determined. To advance our understanding of the molecular basis of beetle immunity here we analyzed the whole transcriptome of T. castaneum by high-throughput next generation sequencing technology. Here, we demonstrate that the Illumina/Solexa sequencing approach of cDNA samples from T. castaneum including over 9.7 million reads with 72 base pairs (bp) length (approximately 700 million bp sequence information with about 30× transcriptome coverage) confirms the expression of most predicted genes and enabled subsequent qualitative and quantitative transcriptome analysis. This approach recapitulates our recent quantitative real-time PCR studies of immune-challenged and naïve T. castaneum beetles, validating our approach. Furthermore, this sequencing analysis resulted in the identification of 73 differentially expressed genes upon immune-challenge with statistical significance by comparing expression data to calculated values derived by fitting to generalized linear models. We identified up regulation of diverse immune-related genes (e.g. Toll receptor, serine proteinases, DOPA decarboxylase and thaumatin) and of numerous genes encoding proteins with yet unknown functions. Of note, septic-injury resulted also in the elevated expression of genes encoding heat-shock proteins or cytochrome P450s supporting the view that there is crosstalk between immune and stress responses in T. castaneum. The present study provides a first comprehensive overview of septic-injury responsive genes in T. castaneum beetles. Identified genes advance our understanding of T. castaneum specific gene expression alteration upon immune-challenge in particular and may help to understand beetle immunity in general.

Oils for Increased Efficacy of Metarhizium anisopliae to Control Whiteflies

The efficacy of Metarhizium anisopliae in combination with sublethal concentrations of oils and potassium-oleate for biological control of whiteflies was tested under controlled conditions. Three commercial products (Biola ® , Naturen ® , Neudosan ® ) and five experimental formulations of plant oils were tested. The efficacy of M. anisopliae against Trialeurodes vaporariorum and Bemisia tabaci without additives was about 50%. At 1/20 of their recommended dosages, all compounds tested significantly increased the efficacy of M. anisopliae for the control of T. vaporariorum , with the formulated sunflower oil Biola ® giving the highest synergistic effect, reaching nearly 100% control. Not only was the level of control increased but also the speed of action was improved, resulting in a higher reliability of control. Three of seven additives showed no effects on the viability of conidia on the leaf surface, whereas the formulations of the other oils and oleates reduced the longevity of spores. The synergistic effect of Biola resulted from the more even distribution of M. anisopliae conidia on leaves and insects. Other positive effects of oils on the efficacy of M. anisopliae are discussed in relation to an extended spectrum of environmental conditions and pests to be controlled.

Localized Adhesion of Nongerminated Venturia inaequalis Conidia to Leaves and Artificial Surfaces

Adhesion to the host surface is the first step for successful plant pathogen development and has been reported to be associated with both passive and active processes. For conidia of Venturia inaequalis, which depend on leaf wetness for germination, this process has not yet been described. Conidia of V. inaequalis adhered to wet hydrophobic surfaces immediately after contact to the surface, hours before initiation of germination. Attachment of nongerminated conidia was much better on hydrophobic surfaces, such as apple leaves and polystyrene, than on hydrophilic glass. Conidia released adhesive material localized in a droplet named spore tip glue (STG) at the spore apex which interacted with a contact surface only when water was present. Histochemical investigations indicated the presence of proteins and carbohydrates in STG, lectin labeling the presence of beta-galactose and N-acetylglucosaminyl residues. Transmission electron microscopy revealed two phases in the STG at the tip of dry mature conidia; as STG was present on the outer side of the intact fungal cell wall its formation should be associated with the secretion of glue through pores of the conidial wall. Surface-active substances affected the adhesion of conidia to hydrophobic surfaces stressing the importance of hydrophobic interactions. The use of protein biosynthesis inhibitors did not affect adhesion of conidia indicating that the adhesive material was preformed. It is concluded that the coincidence of STG, contact to a hydrophobic surface, and free water are essential for the adhesion of V. inaequalis conidia.

Effect of Fungal Colonization of Wheat Grains with Fusarium spp. on Food Choice, Weight Gain and Mortality of Meal Beetle Larvae (Tenebrio molitor)

Species of Fusarium have significant agro-economical and human health-related impact by infecting diverse crop plants and synthesizing diverse mycotoxins. Here, we investigated interactions of grain-feeding Tenebrio molitor larvae with four grain-colonizing Fusarium species on wheat kernels. Since numerous metabolites produced by Fusarium spp. are toxic to insects, we tested the hypothesis that the insect senses and avoids Fusarium-colonized grains. We found that only kernels colonized with F. avenaceum or Beauveria bassiana (an insect-pathogenic fungal control) were avoided by the larvae as expected. Kernels colonized with F. proliferatum, F. poae or F. culmorum attracted T. molitor larvae significantly more than control kernels. The avoidance/preference correlated with larval feeding behaviors and weight gain. Interestingly, larvae that had consumed F. proliferatum- or F. poae-colonized kernels had similar survival rates as control. Larvae fed on F. culmorum-, F. avenaceum- or B. bassiana-colonized kernels had elevated mortality rates. HPLC analyses confirmed the following mycotoxins produced by the fungal strains on the kernels: fumonisins, enniatins and beauvericin by F. proliferatum, enniatins and beauvericin by F. poae, enniatins by F. avenaceum, and deoxynivalenol and zearalenone by F. culmorum. Our results indicate that T. molitor larvae have the ability to sense potential survival threats of kernels colonized with F. avenaceum or B. bassiana, but not with F. culmorum. Volatiles potentially along with gustatory cues produced by these fungi may represent survival threat signals for the larvae resulting in their avoidance. Although F. proliferatum or F. poae produced fumonisins, enniatins and beauvericin during kernel colonization, the larvae were able to use those kernels as diet without exhibiting increased mortality. Consumption of F. avenaceum-colonized kernels, however, increased larval mortality; these kernels had higher enniatin levels than F. proliferatum or F. poae-colonized ones suggesting that T. molitor can tolerate or metabolize those toxins.

Interactions of Fusarium species during prepenetration development

Interspecies interactions between Fusarium avenaceum, Fusarium culmorum, Fusarium graminearum, Fusarium poae, and Fusarium tricinctum were studied during early growth stages of isolates on model surfaces. Additionally, germination and germ tube growth of the pathogens were studied on attached and detached wheat leaves at 10 °C and 22 °C. Two-species interactions between Fusarium isolates during germination and germ tube growth were assessed after 8 hours of incubation. All species except F. tricinctum germinated and grew faster at higher than lower temperature. All species were able to germinate with more than one germ tube per conidium cell; and germination and germ tube growth were faster on leaves than on glass surface. Interactions among Fusarium species during germination and germ tube growth were predominantly competitive with macroconidia-producing species being more competitive. It is concluded that the type of conidia as well as environmental factors influence the competitiveness of Fusarium species during early stages of growth.