Reversible and spatiotemporal control of colloidal structure formation.

Tuning colloidal structure formation is a powerful approach to building functional materials, as a wide range of optical and viscoelastic properties can be accessed by the choice of individual building blocks and their interactions. Precise control is achieved by DNA specificity, depletion forces, or geometric constraints and results in a variety of complex structures. Due to the lack of control and reversibility of the interactions, an autonomous oscillating system on a mesoscale without external driving was not feasible until now. Here, we show that tunable DNA reaction circuits controlling linker strand concentrations can drive the dynamic and fully reversible assembly of DNA-functionalized micron-sized particles. The versatility of this approach is demonstrated by programming colloidal interactions in sequential and spatial order to obtain an oscillatory structure formation process on a mesoscopic scale. The experimental results represent an approach for the development of active materials by using DNA reaction networks to scale up the dynamic control of colloidal self-organization.

Transient self-organisation of DNA coated colloids directed by enzymatic reactions.

Dynamic self-organisation far from equilibrium is a key concept towards building autonomously acting materials. Here, we report the coupling of an antagonistic enzymatic reaction of RNA polymerisation and degradation to the aggregation of micron sized DNA coated colloids into fractal structures. A transient colloidal aggregation process is controlled by competing reactions of RNA synthesis of linker strands by a RNA polymerase and their degradation by a ribonuclease. By limiting the energy supply (NTP) of the enzymatic reactions, colloidal clusters form and subsequently disintegrate without the need of external stimuli. Here, the autonomous colloidal aggregation and disintegration can be modulated in terms of lifetime and cluster size. By restricting the enzyme activity locally, a directed spatial propagation of a colloidal aggregation and disintegration front is realised.

The mechanical properties of polymer-colloid hybrid hydrogels.

The incorporation of monodisperse colloidal particles in hydrogels is a promising approach to create hybrid gels with unique structural, mechanical and functional properties. However, the colloidal structure formation within the hydrogels often remains uncontrolled, leaving behind possible mechanically synergetic effects of the polymeric and the colloidal system. Here we show that colloidal structure formation within the hybrid gels has a significant influence on the elasticity and toughness of the hybrid gels. We combine a polyacrylamide hydrogel with DNA coated colloids (DNAcc), where structure formation can be triggered independently at different points in time. Consequently, we are able to create hybrid gels that are composed of the same components, but do differ in explicit colloidal structure. While monodisperse colloids enhance the storage modulus of the gels, the yield strain is simultaneously drastically reduced. The toughness of these brittle hybrid gels is rescued by colloidal structure formation at higher polyacrylamide concentrations. The toughness is increased at lower polyacrylamide concentrations. We show that the toughness of the hydrogels at 10% (w/v) polyacrylamide and 4% (v/v) DNAcc can be increased by a factor of approx. 35, indicating that control over colloidal structure formation yields access to significant synergetic effects in polymer-colloid hybrid gels.

Development of simple sequence repeat (SSR) markers for discrimination among isolates of Fusarium proliferatum.

The plant pathogen Fusarium proliferatum has a wide host range and occurs worldwide. Many isolates of the fungus produce mycotoxins in plant tissues, which, if ingested, can cause harm to animals and humans. In 2008, an outbreak of salmon blotch of onions, caused by F. proliferatum, was detected in southern Israel. The source and distribution of the fungus in Israel were unknown. Inter-simple sequence repeats (ISSR) were used to identify repetitive motifs present in seven isolates of F. proliferatum from Israel, Germany and Austria. ISSR repeat motifs were, used to develop 17 simple sequence repeat (SSR) loci. Six of these SSR markers were polymorphic in and consistently amplified from ten isolates collected in Israel, Germany, Austria and North America, from cucumber, onion, garlic, maize, and asparagus. These six polymorphic SSR alleles included 5 to 12 copies of di-, tri, and pentanucleotide motifs and yielded six to 9 alleles each. Sixteen of the SSR loci were amplified at least one of the seven Fusarium species, F. verticillioides, F. thapsinum, F. subglutinans, F. andiyazi, F. globosum, F. fujikoroi and F. oxysporum. The data demonstrate that these SSRs can be used for characterization of F. proliferatum isolates from diverse hosts and geographic locations and that they are transferable to other species of Fusarium.

IDENTIFICATION OF DIFFERENT FUSARIUM SPP. IN ALLIUM SPP. IN GERMANY.

In 2013 Allium cepa bulbs from different fields in Northern and Southern Germany, seeds and sets from onion breeders were analysed for infestation with Fusarium species. The same investigation was done in 2014 with different edible Allium spp. from local markets. Different Fusarium spp. were isolated and identified by morphological characterisation. 24 different Fusarium spp. were identified. The diversity of Fusarium spp. and the intensity of infestation was higher on edible bulbs compared to the younger sets and seeds. The analysed onions and other edible Allium spp. from local markets showed also high contents of different Fusarium species. The most prevalent identified Fusarium sp. in the analysed Allium spp. in Germany was Fusarium oxysporum which can cause the Fusarium Basal Rot, followed by Fusarium solani. Fusarium proliferatum, which can cause the Fusarium Salmon Blotch in onions, could be detected in about half of the sampled onion fields and in approximately 10% of all analysed onions from fields. Also in the onion sets, on the surface of the seeds and in other edible Allium spp. F. proliferatum could be identified. Besides F. proliferatum, further mycotoxin producing Fusarium spp. like Fusarium equiseti or Fusarium tricinctum were identified. Other Fusarium spp. like Fusarium sporotrichioides and Fusarium poae were first described in Allium sp. in this study. The two most prevalent Fusarium spp. F. oxysporum and F. solani are able to produce mycotoxins like enniatins, fumonisins, moniliformin and T-2 toxins. Fusarium sp. like F. proliferatum, F. equiseti and F. tricinctum are able to produce additional toxins like beauvericins, zearalenone and diacetoscirpenol. This high number of Fusarium spp., which are able to produce a broad spectrum of different mycotoxins, could be a potential health risk for human beings and livestock.

First Report of the Root-Knot Nematode Meloidogyne hapla Parasitizing Roses in Ethiopia.

Meloidogyne hapla is one of the most damaging plant-parasitic nematodes in temperate regions. This nematode has a wide host range with more than 500 plant taxa including roses. In Ethiopia, rose production has developed over the past 10 years to the second most important export market after coffee. Considering the high damage potential of M. hapla, infestation of roses in Ethiopia with this nematode could result in major economic losses. Therefore, awareness of this nematode species is extremely important. During two surveys conducted in August 2011 and April 2012, M. hapla was detected in soil samples from six out of nine rose producing farms located in the districts of Ziway, Holleta, Sebeta, and Menagesha. At infested farms, rose plants appeared stunted and less productive and often showed symptoms of chlorosis and wilting. Identification was based on morphological and morphometrical characters of females, males, and second-stage juveniles, which were all within the range of variability known for this species (4). Shape of juvenile stylet knobs, shape of male head, and perennial pattern of the females with characteristic punctuations between the anus and tail terminus were also typical for M. hapla. The morphological identification was confirmed by sequence analysis of the D2-D3 expansion segment of the 28S rDNA gene following amplification with the primers D2A (5'-ACAAGTACCGTGAGGGAAAGTT-3') and D3B (5'-TCGGAAGGAACCAGCTACTA-3') (1). PCR products were purified and sequenced at the Macrogene sequencing facility service (Amsterdam, The Netherlands). Sequences were deposited in GenBank (KJ645427 to 33). The sequences were compared with previously published sequences in NCBI database and showed 96 to 100% sequence similarity with M. hapla accession nos. GQ130139, DQ328685, KF430798, and DQ145641. Unfortunately, comparison of sequences did not provide further information about the origin of this Ethiopian population, if it is native to Ethiopia or was imported with infected plant material. To the best of our knowledge, this is the first record of M. hapla occurring in Ethiopia. M. hapla is known as a serious pest of roses in colder climate regions. In Africa, it was previously reported from Tanzania (3) and South Africa (2). Thus, it appears that this species has now become also established in Ethiopia at higher altitudes (1,400 to 2,100 m above sea level) within the urban hinterland of Addis Ababa. References: (1) Baldwin et al. Mol. Phy. Evol. 8:248, 1887. (2) J. H. O'Bannon. Institute Agri. Res. 29, 1975. (3) E. Onkendi and L. N. Moleleki. Eur. J. Pl. Pathol. 136:1, 2013. (4) A. G. Whitehead. Trans. Zool. Soc. Lon.31:263, 1968.

Microbiological and SYBR green real-time PCR detection of major Fusarium head blight pathogens on wheat ears.

Fusarium head blight (FHB) caused by several Fusarium species is one of the most serious diseases affecting wheat throughout the world. The efficiency of microbiological assays and real-time PCRto quantify major FHB pathogens in wheat ears after inoculation with F. graminearum, F. culmorum, F. avenaceum and F. poae undergreenhouse and field conditions were evaluated. The frequency of infected kernel, content of fungal biomass, disease severity and kernel weight were determined. To measure the fungal biomass an improved DNA extraction method and a SYBR Green real-time PCR were developed. The SYBR Green real-time PCR proved to be highly specific for individual detection of the species in a matrix including fungal and plant DNA. The effect of Fusarium infection on visible FHB severity, frequency of infected kernels and thousand-kernel mass (TKM) significantly depended on the Fusarium species/isolate. F. graminearum resulted in highest disease level, frequency of infected kernels, content of fungal biomass, and TKM reduction followed by F. culmorum, EF avenaceum and F. poae, respectively. The comparison of frequency and intensity of kernel colonization proved differences in aggressiveness and development of the fungi in the kernels. Only for F. graminearum, the most aggressive isolate, application of microbiological and real-time PCR assays gave similar results. For the other species, the intensity of kernel colonization was lower than expected from the frequency of infection.

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.

Thermal imaging of cucumber leaves affected by downy mildew and environmental conditions.

Pathogenesis of Pseudoperonospora cubensis causing downy mildew of cucumber resulted in changes in the metabolic processes within cucumber leaves including the transpiration rate. Due to the negative correlation between transpiration rate and leaf temperature, digital infrared thermography permitted a non-invasive monitoring and an indirect visualization of downy mildew development. Depending on the stage of pathogenesis and the topology of chloroses and necroses, infection resulted in a typical temperature pattern. Spatial heterogeneity of the leaf temperature could be quantified by the maximum temperature difference (MTD) within a leaf. The MTD increased during pathogenesis with the formation of necrotic tissue and was related to disease severity as described by linear and quadratic regression curves. Under controlled conditions, changes in temperature of infected leaves allowed the discrimination between healthy and infected areas in thermograms, even before visible symptoms of downy mildew appeared. Environmental conditions during thermographic measurement, in particular air temperature and humidity, as well as water content and age of the leaf influenced the temperature of its surface. Conditions enhancing the transpiration rate facilitated the detection of changes in leaf temperature of infected leaves at early stages of infection. As modified by environmental conditions, MTD alone is not suitable for the quantification of downy mildew severity in the field.

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.

Effects of arbuscular mycorrhizal fungi and a non-pathogenic Fusarium oxysporum on Meloidogyne incognita infestation of tomato.

Arbuscular mycorrhizal (AM) fungi and non-pathogenic strains of soil-borne pathogens have been shown to control plant parasitic nematodes. As AM fungi and non-pathogenic fungi improve plant health by different mechanisms, combination of two such partners with complementary mechanisms might increase overall control efficacy and, therefore, provide an environmentally safe alternative to nematicide application. Experiments were conducted to study possible interactions between the AM fungus Glomus coronatum and the non-pathogenic Fusarium oxysporum strain Fo162 in the control of Meloidogyne incognita on tomato. Pre-inoculation of tomato plants with G. coronatum or Fo162 stimulated plant growth and reduced M. incognita infestation. Combined application of the AM fungus and Fo162 enhanced mycorrhization of tomato roots but did not increase overall nematode control or plant growth. A higher number of nematodes per gall was found for mycorrhizal than non-mycorrhizal plants. In synergisms between biocontrol agents, differences in their antagonistic mechanisms seem to be less important than their effects on different growth stages of the pathogen.

The IPM Wheat Model--results of a three-year study in North Rhine-Westphalia, Lower Saxony and Schleswig-Holstein.

Under the primary utilisation of phytosanitary production factors such as selection of variety, crop rotation and N fertilisation according to plant requirements, the IPM Wheat Model comprises the elements diagnosis (qualitative = type of pathogen, quantitative = disease severity), scientifically grounded treatment thresholds which, as critical values in pathogen development, can be applied to define the optimum time of fungicide application, and pathogen-specific effective fungicides and application amounts. This leads to the location and year-specific optimised control of the pathogen and of the associated yield performance. After several years of development in Bavaria (from 1985 on) and Schleswig-Holstein (1993-1999), the model was tested as part of a project involving the Universities of Bonn and Kiel and the plant protection services of the German states of Lower Saxony, North Rhine-Westphalia and Schleswig-Holstein in a three-year study (1999-2001) in interregional locations (usually nine per state) with the winter wheat variety Ritmo (interregional indicator variety) and a further variety of regional importance in different variations (untreated control, three to four times growth stage-oriented variants for the determination of the absolute damage potential, IPM-variant). In exact records (approx. 12 dates per vegetation period), the disease epidemics were recorded weekly. With the genetically uniform indicator variety Ritmo, the results documented substantially differing year- and location-specific disease and yield patterns. Interregionally, a broad wheat pathogen spectrum (Puccinia striiformis, P. recondita, Septoria tritici, Stagonospora (syn. Septoria) nodorum, Blumeria (syn. Erysiphe) graminis, Pseudocercosporella herpotrichoides, Drechslera tritici-repentis) in differing composition, disease severity and damage effect was demonstrated. The heterogeneity of the infection and damage patterns was increased in the case of the second variety, in association with the genetic variability. The epidemiologically-orientated indications (average two, reduced application amounts) according to the IPM Wheat Model in association with time-diverging progressions led, on an interregional basis, with minimum input and in association with the diverging dynamics, to a biologically and economically optimised fungicide application. In the context of economic and ecological performance, the comprehensive results of the project demonstrated the valuable functionality of the IPM Wheat Model.

Impact of strobilurins on physiology and yield formation of wheat.

Strobilurin fungicides have a broad spectrum activity against all major foliar pathogens of wheat. In addition to this extraordinary fungicidal activity side-effects have been reported which result in higher yields of cereals, e.g. the reduction of respiration, delayed leaf senescence, activation of nitrogen metabolism as well as increased tolerance against abiotic stress factors. In the vegetation period 2000/2001 field trials were carried out at three sites in North Rhine-Westphalia to study the effects of three strobilurin fungicides on the yield formation of six winter wheat varieties. The strobilurins were applied two times as the commercial products Stratego (trifloxystrobin + propiconazole), Amistar/Pronto Plus (azoxystrobin/spiroxamine + tebuconazole) and Juwel Top (kresoxim-methyl + epoxiconazole + fenpropimorph. Fungicide-treated plants were kept disease-free by an initial azole-application in GS 31 in order to exclude disease effects on physiological parameters relevant to yield formation. Photosynthetic electron transport of strobilurin-treated wheat, was improved as early as at GS 65 compared to azole-treated plants. Differences often increased with growth stage and were closely related to a delay in leaf senescence. A higher photosynthetic activity of strobilurin-treated plants was confirmed by gas exchange and chlorophyll fluorescence measurements under field conditions. The yield benefit of wheat from strobilurin treatments varied from 2% to 9% depending on an improved photosynthetic capacity due to a higher and/or prolonged activity. Neither yield potential nor disease susceptibility of the cultivar had an effect on the height of the extra yield which, in contrast was modified by location and wheat genotype.

Evaluation of the IPM Wheat Model in the Rhineland 2001.

The results of field trials at nine locations with two wheat cultivars in each case demonstrated the practicability of the action threshold-based fungicide strategy of the IPM Wheat Model under the intensive wheat production conditions in the Rhineland. Action thresholds used and dosage of fungicides applied are outlined. Despite using reduced rates of fungicides the epidemic development of relevant pathogens--mainly Septoria tritici and Puccinia recondita--was controlled in early stages of disease development. In plots treated according to the IPM Wheat Model disease incidence and disease severity were reduced effectively compared to untreated plots with low and high disease incidence, respectively. The effective control of pathogens using pathogen-specific action thresholds resulted in yield levels very similar to those of disease-free plots simultaneously minimising the total amount of fungicides applied. Disease control according to the IPM Wheat Model raised yields by up to 25% with monetary benefits averaging more than [symbol: see text] 100 ha-1 in 2001.

The role of some weeds as hosts of Capsicum viruses in the rift valley parts of Ethiopia.

Hot pepper (Capsicum annuum. L.) is a traditionally and economically important spice crop in Ethiopia. Chronic infection with viruses is reported as major constraint, that often force farmers to ban its production. However, epidemiological studies on these viruses are lacking in the country. Knowledge of the weed flora as virus reservoir is essential to reduce the spread of plant viruses. The potential role of some frequently and abundantly occurring weed species as host plants for Capsicum viruses was therefore investigated. Surveys were conducted during the 2000/2001 cropping- and off-season. Samples of various weed species revealing virus symptoms were collected from different districts. Collection was done from inside and at the border of pepper cropped and fallow fields. It was also performed along road sides and irrigation channels. The identity of the collected isolates was established based on serology, electron microscopy and host range studies. Moreover, seeds were collected from natural populations of the most prevalent weeds, namely Datura stramonium and Nicandra physalodes. They were grown under glass house conditions and tested for their ability to host viruses. To verify their role further, healthy seedlings from these weeds were artificially inoculated with all viruses isolated from weeds and Capsicum leaf samples. Natural infection of different Potyviruses with an average incidence of 33% in the weed samples were the most dominant. In addition, the occurrence of Cucumber mosaic virus, Tomato mosaic virus and Potato virus X in these weeds was demonstrated. However, none of the isolated viruses was found to be seed borne in the tested weed species and pepper crop (var.Mrkofanna) under glass house conditions. Recently, there was an explosion in population size of some solanaceous weeds, particularly Datura and Nicandra species in the area. The presence of infected weeds throughout the year means, that they are an important reservoir and source for secondary spread. It is suggested therefore, that a rigorous weed control is an important step to reduce the incidence of viruses infecting Capsicum in the area.

Biological control of grey mould (Botrytis cinerea) with the antagonist Ulocladium atrum.

The competitive saprophytic fungus Ulocladium atrum was selected on the basis of its antagonistic potential for the control of grey mould caused by the necrotrophic pathogen Botrytis cinerea. Field trials were carried out to asses the efficacy of U. atrum as a biocontrol agent against B. cinerea in grapevine. The results demonstrated that under moderate disease pressure U. atrum had the potential to control grey mould, whereas under high disease pressure the efficacy was not sufficient to substitute the use of fungicides completely.

Variation in the spectrum of Fusarium species on winter wheat.

In 1998-2000 a monitoring of the spectrum of Fusarium species on winter wheat was carried out in the Rhineland. The epidemic spread ofFusarium spp. on wheat plants during growing season was investigated as well as the grain contamination after harvest.F avenaceum was the Fusarium species isolated most frequently followed byF culmorum, F poae andF graminearum. Microdochium nivale occurred considerably only in 1998. Both, susceptibility and plant height of the cultivars were correlated to the incidence of Fusarium species /M nivale on harvested kernels; interactions with cropping intensities were detected. The incidence ofF poae seemed to be independent of the cultivar-specific Fusarium susceptibility. Despite the lack of disease symptoms, between growth stages 45-85 mycelium ofFusarium spp. was detectable in the leaves as well as conidia on the leaf surfaces.

Significance of different inoculum sources for the Fusarium infection of wheat ears.

From 1997 to 1999 the occurrence ofFusarium spp. on wheat grain and its contamination with the mycotoxins deoxynivalenol and invalenol were investigated under organic farming conditions in the Rhineland, Germany. For comparison, some trials were also run under integrated farming conditions. The importance of the seed contamination withFusarium spp. as well as the impact of farming system, previous crop and soil preparation on the inoculum sources ofFusarium spp. in the soil were investigated. The data on the inoculum sources was compared to the Fusarium infection of grains and their content of DON and NIV. The crop residues in the soil were the most important inoculum source for the Fusarium species infecting wheat ears and grains. The amount of potential inoculum in the soil largely depended on the previous crop and the system of soil preparation.

Rheological and breadmaking properties of wheat samples infected withFusarium spp.

Rheological and breadmaking properties of untreated and suboptimally stored wheat samples (grain moisture: 20%, temperature: 20°C) and also of wheat which was inoculated withFusarium spp. were investigated. The deoxynivalenol (DON) content of the stored and inoculated wheat samples ranged between 820-12,000 µg/kg. Gluten proteins were isolated with different extraction solutions and the fractions obtained were analysed by means of RP-HPLC. Microextension tests and micro-baking tests were used for the determination of dough properties (maximum resistance (MR) and extensibility (EX)) and bread volume, respectively. In spite of the extremely high DON concentrations of some wheat samples contaminated withFusarium spp. they showed only a slight decrease of the amount of gluten proteins. Extension tests of dough led to a slight decrease of MR, bread volumes stayed almost the same compared with the non-contaminated grain. The contamination of wheat withAspergillus andPenicillium led to a high decrease of gluten proteins, which resulted in an extremely decreased MR of the dough and a very low bread volume.

Impact of growth conditions on the occurrence ofFusarium spp. and the mycotoxin content of wheat.

In 1997-99 the occurrence ofFusarium spp. on winter wheat and the contamination with mycotoxins was investigated at three locations in the Rhineland, Germany. All cultivation methods investigated had an effect on the level ofFusarium infection, however, rainfall during flowering was the most important factor. The choice of cultivar and soil cultivation proved to be the most promising tools to reduce head scab severity and mycotoxin contamination.