Epidemiology of cercospora leaf spot on sugar beet: modeling disease dynamics within and between individual plants

ABSTRACT Disease dynamics of Cercospora leaf spot (CLS) of sugar beet was analyzed at two hierarchical scales: as vertical profiles within individual plants and in relation to disease on neighboring plants. The relative contribution of different leaf layers to increase in CLS was analyzed using a simple continuous-time model. The model was fitted to data from two field trials in the Netherlands: one in an area with a long history of CLS, the other in an area where CLS has only recently established; in each case these were unsprayed and twice-sprayed treatments. There were differences in the relative contribution of different leaf layers to disease increase on the target leaf layer according to the CLS history and whether the plants were sprayed or unsprayed. In both field trials, parameter estimates giving the relative contribution of the target leaf layer to disease increase at that leaf layer were higher than those for the lower leaf layer. On only a few occasions the contribution of an upper leaf layer to disease increase at the target leaf layer was significant. Thus, CLS increase at the target leaf layer was determined mainly by disease severity at that leaf layer and to a lesser extent by disease at the lower leaf layer. Our continuous-time model was also used to analyze CLS increase on an individual sugar beet plant in relation to its own and its neighbor's level of disease in field trials at five locations in the two CLS areas over two years. In all field trials, the contribution of the target plant itself to disease increase (auto-infection) was larger than that of its neighboring plants (allo-infection). The overall analysis in the two CLS areas also indicated a larger contribution of the target plant to its disease increase than of neighboring plants, and this pattern was also apparent in a pooled analysis across all sites. Thus, CLS increase on a sugar beet plant was mainly determined by the disease severity on that plant and to a lesser extent by its within-row neighboring plants.     

Impact of climate change on the temporal and regional occurrence of Cercospora leaf spot in Lower Saxony

Journal of Plant Diseases and Protection - The effect of climate change on the temporal and regional occurrence of Cercospora leaf spot (CLS) of sugar beet in Lower Saxony is analysed using the...     

Root infection of sugar beet by <Emphasis Type="Italic">Cercospora beticola</Emphasis> in a climate chamber and in the field

Sugar beet root infection by Cercospora beticola, the causal agent of Cercospora leaf spot (CLS), was studied in a climate chamber and in the field. In the climate chamber, root incubation of susceptible seedlings with a conidial suspension resulted in disease incidences that were significantly different for two sugar beet cultivars (Auris: 0.8 ± 0.14 and A00170: 0.5 ± 0.18; P < 0.05) with regard to the control treatment 35 days after root incubation in a standard potting soil-fine river sand mixture. In a field trial with susceptible cv. Savannah with soil-incorporated CLS-infested leaf material, disease developed four weeks earlier in the infested plots than in the control plots. The probability that disease develops in the field was significantly higher for the infested than for the control plots (P < 0.05). Symptomless plants from infested field plots transferred to the glasshouse to induce leaf spot symptoms showed a significantly higher probability to induce symptom development (0.4 ± 0.08), than plants from control plots (0.02 ± 0.02) (P <0.05) 14 days after transfer. This probability was significantly higher than for plants that remained in three of the infested field plots (0.2 ± 0.04; 0.2 ± 0.05 and 0.2 ± 0.04 respectively), except for one infested field plot (0.4 ± 0.05) on July 5. We conclude that C. beticola is able to infect sugar beet seedlings through their roots and that latent CLS infections in sugar beet lead to symptom development at high temperatures (> 20 °C) and high relative humidity (> 95) in our climate chamber or after canopy closure in the field. Quantification of root infection and long term survival in soil is necessary to assess its contribution to the epidemiology and life cycle of Cercospora beticola. Cultural methods such as a wider crop rotation, management of crop debris and ploughing systems may provide control strategies alternative to or reducing fungicide input.     

Possible Root Infection of Cercospora beticola in Sugar Beet

A potential primary infection site of the foliar pathogen Cercospora beticola in sugar beet is described. Sugar beet seedlings of the susceptible cv. Auris were grown in a standard soil for 14 days. A monoconidial culture of a C. beticola isolate was grown to produce conidia. In experiment 1, roots were immersed in a conidial suspension of isolate code IRS 00-4, or in tap water (control), for 2 days. After incubation seedlings were potted in a peat – fine river sand mixture and placed at low relative humidity (RH) (<80%) or high RH (100%). Twelve days after infection, seedlings at high RH showed more disease incidence (90%) than seedlings grown at low RH (disease incidence = 25%), whereas no disease symptoms developed in the control seedlings. Cercospora leaf spots (CLSs) developed on the cotyledons, leaves, petioles and stems of the seedlings. In experiment 2, roots were immersed in a conidial suspension of isolate code IRS 00-2 for 5 h. Thirty-four days after infection at high RH, 100% disease incidence was observed in the treated seedlings and one CLS in the control treatment. First indications of leaf spot development were observed as reddish purple discolouration of individual parenchymatic cells. Because splash dispersal and symptoms due to infested soil were excluded, we showed that it is possible to obtain CLS symptoms in sugar beet seedlings when their roots were immersed in conidial suspensions of C. beticola, thus demonstrating that roots can be a primary infection site.     

The use of digital image analysis and real‐time PCR fine‐tunes bioassays for quantification of Cercospora leaf spot disease in sugar beet breeding

Cercospora leaf spot, caused by the fungus Cercospora beticola, is a major fungal sugar beet disease worldwide and the cause of significant yield losses. The disease is most successfully countered by the introduction of genetic tolerance into elite sugar beet hybrids. To this end, breeding programmes require high quality biological assays allowing discrimination of minor differences between plants within a segregating population. This study describes the successful implementation of image analysis software in the bioassays for quantification of necrotic lesions at different stages of C. beticola infection, allowing selection on minor phenotypic differences during the sugar beet breeding process for C. beticola resistance. In addition, a real‐time PCR assay was developed for the quantification of C. beticola pathogen biomass in infected beet canopy. The use of both techniques, even in an early stage of infection, fine‐tunes current bioassays, allowing more accurate and efficient selection of resistant breeding material.     

Genetic diversity and pathogenicity of Pseudomonas syringae pv. aptata isolated from sugar beet

Pseudomonas syringae pv. aptata is the causal agent of bacterial leaf spot disease of sugar beet (Beta vulgaris). During 2013, 250 samples were collected from leaf lesions with typical symptoms of bacterial leaf spot in commercial fields of sugar beet in Serbia, and 104 isolates of P. syringae pv. aptata were obtained. Identification and characterization was performed using biochemical, molecular and pathogenicity tests. Identification included LOPAT tests and positive reactions using primers Papt2F and Papt1R specific for P. syringae pv. aptata. Repetitive (rep) sequence‐based PCR typing with ERIC, REP and BOX primers revealed high genetic variability among isolates and distinguished 25 groups of different fingerprinting profiles. Pulse‐field gel electrophoresis (PFGE) and multilocus sequence analysis (MLSA) of representative isolates showed higher genetic variability than in rep‐PCR analysis and distinguished three and four major genetic clusters, respectively. A pathogenicity test performed with 25 representative isolates on four cultivars of sugar beet confirmed the occurrence of leaf spot disease and showed correlation between the most aggressive isolates and the genetic clusters obtained in MLSA. All these findings point to the existence of several lines of P. syringae pv. aptata infection in Serbia that are genetically and pathologically different.     

Early detection of sugar beet pathogen Ramularia beticola in leaf and air samples using qPCR

A quantitative PCR method (qPCR) was developed for the detection and quantification of Ramularia beticola causing Ramularia leaf spot in sugar beet. R. beticola specific primers were designed based on the internal transcribed spacer region 2 (ITS2). The assay was applied on DNA extracted from spores trapped on tape from Burkard spore traps placed in an artificially inoculated sugar beet field trial and in two sugar beet fields with natural infections. R. beticola DNA was detected at variable amounts in the air samples 14 to 16 days prior to first visible symptoms. R. beticola DNA was detected in air samples from fields with natural infection at significant and increasing levels from development of the first symptoms, indicating that spore production within the crop plays a major role in the epidemic development of the disease. Sugar beet leaves sampled from the inoculated field trial were also tested with the qPCR assay. It was possible to detect the presence of R. beticola in the leaves pre-symptomatic at least 10 days before the occurrence of the visible symptoms of Ramularia leaf spot. This is the first report of a molecular assay, which allows screening for the presence of R. beticola in plant material and in air samples prior to the appearance of visible symptoms. An early detection has potential as a tool, which can be part of a warning system predicting the onset of the disease in the sugar beet crop and helping to optimise fungicide application.     

Optimierte Strategien zur Krankheitsbekämpfung in der Gerste unter Berücksichtigung neuer Erkenntnisse zur Epidemiologie des Blattfleckenkomplexes

The leaf spot complex constitutes an increasing challenge for integrated pest control in barley. The complex consists of biotic and abiotic factors causing early ripening after a rapid loss of green leaf area, in particular in upper, exposed leaves. Severe symptoms appear late in the growing season after heading. The experience from recent years shows that the control of the leaf spot complex improves yield. Best control is achieved by new, effective fungicides. Efficiency is improved by late fungicide application and is dose-dependent. In contrast, late treatments provide poor control of early appearing diseases. For a successful disease control in barley good efficiency in the control of the leaf spot complex appears based on a sufficient control of the other diseases. For optimization of disease control, we require improved knowledge of the epidemics of the leaf spot complex and of the contribution of individual biotic and abiotic factors. Besides the weather, the fungal pathogen Ramularia collo-cygni and its photodynamic toxins play a mayor role in symptom development. By combination of conventional and molecular diagnostics we aim at a better understanding of the complex as a basis for early and reliable prediction of epidemics. Because non fungicidal factors like the physiological plant age play a significant role in the occurrence of the leaf spot complex, they should be taken into account for integrated control.     

Foliar spray of a cell wall protein fraction from the biocontrol agent <Emphasis Type="Italic">Pythium oligandrum</Emphasis> induces defence-related genes and increases resistance against Cercospora leaf spot in sugar beet

After a cell wall protein fraction (CWP) of Pythium oligandrum was sprayed on sugar beet leaves, we screened leaves for induced expression of defence-related genes and for resistance against Cercospora leaf spot. In a western blot analysis, the CWP was primarily retained on the surface of leaves without degradation for at least 48 h after spraying. In northern blot analyses, four defence-related genes (β-1, 3-glucanase, acidic class III chitinase, 5-enol-pyruvylshikimate-phosphate synthase and oxalate oxidase-like germin) were expressed more rapidly in CWP-treated leaves compared to control leaves treated with distilled water (DW). When CWP was applied to a suspension of cultured cells of sugar beet, an oxidative burst was observed that did not occur after the DW treatment. In growth chamber trials after inoculation with Cercospora beticola, the severity of Cercospora leaf spot was significantly reduced in CWP-treated plants compared to the DW-treated controls. In a field experiment, CWP treatment was also effective against the disease. CWP did not reduce growth rate of the pathogen in plate tests. The results together suggest that the CWP from P. oligandrum can be retained on the leaf surface and induce expression of disease resistance genes, thereby reducing Cercospora leaf spot on sugar beet.     

Selectivity of foramsulfuron + thiencarbazone‐methyl and classic herbicides in sensitive and non‐sensitive sugar beet genotypes

A new herbicide for sugar beet cultivation using the ALS‐inhibiting active ingredients foramsulfuron and thiencarbazone‐methyl is under approval in the EU member states. Sugar beet genotypes that are non‐sensitive to this herbicide are currently under development. Selectivity of the ALS‐inhibiting herbicide and yield response of the non‐sensitive genotypes might be relevant to meet the requirements for variety registration. To evaluate these issues, six field trials were conducted in Germany in 2013 and 2014. Classic herbicides and the ALS‐inhibitor herbicide were applied in dosages of up to fourfold the authorised (or applied for) application rates. The ALS‐inhibitor herbicide did not cause any significant phytotoxicity and had no effect on leaf area index at a single, double or fourfold dosage. By contrast, classic herbicides had significant negative effects at the single dosage. At fourfold dosage, they caused 41% phytotoxicity and reduced leaf area index by 35%. The relative yield difference between ALS‐inhibitor and classic herbicide treatments was 8.6% and 17.4% of white sugar yield at double and fourfold dosage respectively. The ALS‐inhibitor herbicide thus showed higher selectivity than the classic herbicides. In the registration process, the resulting yield advantage could balance a possible yield penalty of non‐sensitive genotypes. The introduction of a new system for weed control could improve application flexibility and control of troublesome weeds in sugar beet.     

贵州六盘水市猕猴桃病害调查及病原鉴定

针对2014年以来六盘水市猕猴桃病害日益严重的问题,分别于2015年10月、2016年4月及8月对六盘水市共12个乡镇26个代表性猕猴桃栽培园区病害进行了调查。采集了大量典型病害样本,综合生物学特性、分子鉴定及致病力测试对分离得到的病原菌进行鉴定。结果表明,六盘水市猕猴桃夏季和秋冬季病害主要为细菌性溃疡病和真菌性软腐病、灰斑病、褐斑病,其次是炭疽病和黑斑病,春季部分园区有细菌性溃疡病,真菌病害相对较轻。本文研究结果为六盘水市猕猴桃病害的预测预报及综合防治提供了理论依据。     

Damping-off of sugar beet caused by Rhizoctonia cerealis

Rhizoctonia cerealis was isolated from diseased sugar beet seedlings collected from crops in Ireland. In pathogenicity tests, isolates of R. cerealis from beet seedlings, and from sharp eyespot lesions on wheat crops, caused severe damping-off of sugar beet. Isolates from beet seedlings also caused symptoms of sharp eyespot on wheat.     

Dynamics of cercospora leaf spot disease determined by aerial spore dispersal in artificially inoculated sugar beet fields

Cercospora beticola is one of the most important fungal pathogens of sugar beet, causing cercospora leaf spot (CLS) disease. Due to the decreasing efficacy of various fungicides caused by resistance traits, the development of a sustainable disease management strategy has become more important. Therefore, detailed knowledge about the epidemiology of the pathogen is crucial. Until now, little was known about the spatiotemporal dispersal of C. beticola spores from the primary inoculum source. Rapid detection of C. beticola spores could facilitate a more precise and targeted disease control. Therefore, a TaqMan real-time PCR assay for detection and quantification of C. beticola spores caught with Rotorod spore traps was established. In 2016 and 2017, field trials were conducted to monitor C. beticola aerial spore dispersal and disease development within an inoculated field and in the adjacent noninoculated area. With the established detection method, C. beticola spores were successfully quantified and used as a measure for aerial spore dispersal intensity. The analysis of the spatiotemporal spread of C. beticola spores revealed a delay and decrease of aerial spore dispersal with increasing distance from the inoculated area. Consequently, disease incidence and severity were reduced in a similar manner. These results imply that spore dispersal occurs mainly on a small scale within a field, although long distances can be overcome by C. beticola spores. Moreover, secondary aerial spore dispersal from sporulating leaf spots seems to be the main driver for CLS disease development. These results provide an important basis for further improvement of CLS control strategies.     

Identification of the G143A mutation associated with QoI resistance in Cercospora beticola field isolates from Michigan,United States

BACKGROUND: Cercospora leaf spot (CLS), caused by the fungus Cercospora beticola, is the most serious foliar disease of sugar beet (Beta vulgaris L.) worldwide. Disease control is mainly achieved by timely fungicide applications. In 2011, CLS control failures were reported in spite of application of quinone outside inhibitor (QoI) fungicide in several counties in Michigan, United States. The purpose of this study was to confirm the resistant phenotype and identify the molecular basis for QoI resistance of Michigan C. beticola isolates. RESULTS: Isolates collected in Michigan in 1998 and 1999 that had no previous exposure to the QoI fungicides trifloxystrobin or pyraclostrobin exhibited QoI EC₅₀ values of ?0.006 µg mL^?1. In contrast, all isolates obtained in 2011 exhibited EC₅₀ values of > 0.92 µg mL^?1 to both fungicides and harbored a mutation in cytochrome b (cytb) that led to an amino acid exchange from glycine to alanine at position 143 (G143A) compared with baseline QoI‐sensitive isolates. Microsatellite analysis of the isolates suggested that QoI resistance emerged independently in multiple genotypic backgrounds at multiple locations. A real‐time PCR assay utilizing dual‐labeled fluorogenic probes was developed to detect and differentiate QoI‐resistant isolates harboring the G143A mutation from sensitive isolates. CONCLUSION: The G143A mutation in cytb is associated with QoI resistance in C. beticola. Accurate monitoring of this mutation will be essential for fungicide resistance management in this pathosystem. Copyright © 2012 Society of Chemical Industry     

Sugarbeet yield response to competition from Sinapis arvensis or Lolium perenne growing at three different distances from the beet and removed at various times during early growth

A sugarbeet field experiment was conducted in 1999 and 2000 to measure beet yield where Sinapis arvensis or Lolium perenne were growing in the crop row at 2, 4 or 8 cm from the beet plants. The weeds were removed by cutting once in the growing season in either late May, mid‐June or early July. The number of neighbouring beet plants to every target beet plant was recorded. Projected leaf cover of a subset of the data with non‐cut weeds was analysed by using image analysis to investigate whether this could be used to predict beet yield loss early in the growing season. Increasing the distance between beet and weed from 2 to 8 cm increased the beet yield significantly by an average of 20%, regardless of weed species. The dry weight of non‐cut and re‐growing weeds at harvest time decreased when cutting was postponed to the period between mid‐June and early July. The number of neighbours described a sigmoidal yield decline of the single beet plants. Results from image analysis showed that approximately 33 g of beet yield was lost in October/November for each per cent relative projected leaf cover of the weeds in May, despite variation in growing conditions. The results are discussed in relation to potentials for robotic in‐row weed control.     

Effects of cherry leaf spot on photosynthesis in tart cherry 'Montmorency' foliage

Results described here span a total of three field seasons and quantitatively depict the effects of an economically important fungal pathogen (Blumeriella jaapii) on tart cherry (Prunus cerasus 'Montmorency') leaf physiology. For the first time, leaf photosynthesis, stomatal conductance (g(s)), maximum ribulose-1,5-bisphosphate carboxylation rate (V(cmax)), and maximum electron transport (J(max)) were measured as functions of visible cherry leaf spot disease (CLS) severity. Defined as the proportion of chlorotic and necrotic tissue per leaf, CLS severity was estimated from leaves of mature 'Montmorency' trees in 2007, 2008, and 2009. Briefly, as visible disease severity increased, all of the leaf-level physiological parameters decreased significantly (P < 0.01) and disproportionately. Thus, the effects of visible symptoms on leaf photosynthetic metabolic function encroached upon asymptomatic tissue as well. Impairment of photosynthetic metabolism in 'Montmorency' tart cherry leaves due to CLS appears to be mediated through disproportionately large perturbations in g(s), V(cmax), and J(max). These findings offer a new perspective on the amount of damage that this serious disease can inflict.     

CERCBET 3 – a forecaster for epidemic development of Cercospora beticola*

Cercospora beticola is the most prevalent and damaging fungal disease in German sugar beet growing. Control strategies are based on action thresholds. A model has been developed which forecasts epidemic development (expressed as disease incidence) and signals when action thresholds are overridden. The plot‐specific model, CERCBET 3 uses as input meteorological parameters (temperature, relative humidity), easily accessible agronomic field characteristics and a single recording of C. beticola disease incidence. Extensive validation in 2001–03 showed that, in 80–95% of the cases, CERCBET 3 correctly forecasted the dates when thresholds were overridden. Cultivar diversity in German sugar beet growing is increasing, thus a module has been included into CERCBET 3 which reflects susceptibility to C. beticola by introducing a sporulation factor. In some cases a second or even third fungicide treatment could be necessary to control Cercospora leaf spot and so a further module which models fungicide efficacy has been elaborated. CERCBET 3 is available for sugar beet growers in an interactive form on the Internet platform ISIP, which is provided by the governmental crop protection services of Germany.     

<Emphasis Type="Italic">Clavibacter michiganensis</Emphasis> subsp. <Emphasis Type="Italic">michiganens</Emphasis>is strains virulence and genetic diversity. a first study in Argentina

Tomato leaves showing severe leaf spot symptoms have been observed and sampled in the central west and southwest Taiwan during 2015 and 2016. The symptoms were similar to those of bacterial leaf spot/late blight diseases, but only Stemphylium-like fungi were consistently isolated from the diseased tomato. Upon spray inoculation of tomato, Stemphylium-like isolates caused leaf spot symptoms identical to those of naturally infected plants, and the pathogenic isolates were successfully re-isolated from inoculated leaves. The tomato-pathogenic isolates were identified as S. lycopersici based on morphological characterization and molecular identification. S. lycopersici has been previously reported to cause gray leaf spot of tomato in the temperate regions, but the majority of S. lycopersici-caused lesions were black/dark brown rather than gray in our surveillance. Accordingly, it is suggested that S. lycopersici-caused disease of tomato is named Stemphylium leaf spot of tomato more appropriately than tomato gray leaf spot. Moreover, S. lycopersici-caused leaf spot disease on tomato has been distributed in major tomato production regions in Taiwan. The information provided by our study will be important for future breeding of tomato cultivars, especially for tomato producers in Taiwan.     

Spatial pattern of Cercospora leaf spot of sugar beet in fields in long- and recently-established areas

Spatial disease pattern of Cercospora beticola was characterised during natural epidemics of Cercospora leaf spot (CLS) in sugar beet. We applied linear regression and geostatistical analyses to characterise CLS spatial patterns in three field trials, in long-established and recently-established CLS-areas, during two consecutive years. Linear regression showed a positive influence of average disease severity of within-row neighbouring plants (0.38 < < 0.88). Semi-variograms modelled the spatial dependence of disease severity for two directions per week in both years. Disease severity displayed strong spatial dependence over time. The within-row spatial dependence was the largest, but across-row dependence was irregular and weaker. Both long- and recently established areas showed strong spatial dependence of disease severity within row, decrease in variability between years and within the second trial year and a relation between and the relative nugget. Observed differences were more field than area specific. These spatial and temporal analyses indicated that disease severities of adjacent plants were dependent; hence, we concluded that C. beticola is dispersed mainly over short distances from plant to plant.     

Characterization of cytochrome b from European field isolates of Cercospora beticola with quinone outside inhibitor resistance

Cercospora leaf spot (CLS), caused by the fungal pathogen Cercospora beticola, is the most important foliar disease of sugar beet worldwide. Control strategies for CLS rely heavily on quinone outside inhibitor (Q_OI) fungicides. Despite the dependence on Q_OIs for disease control for more than a decade, a comprehensive survey of Q_OI sensitivity has not occurred in the sugar beet growing regions of France or Italy. In 2010, we collected 866 C. beticola isolates from sugar beet growing regions in France and Italy and assessed their sensitivity to the Q_OI fungicide pyraclostrobin using a spore germination assay. In total, 213 isolates were identified with EC₅₀ values greater than 1.0???g?ml^?1 to pyraclostrobin, all of which originated from Italy. To gain an understanding of the molecular basis of Q_OI resistance, we cloned the full-length coding region of Cbcytb, which encodes the mitochondrial Q_OI-target enzyme cytochrome b in C. beticola. Cbcytb is a 1,162-bp intron-free gene with obvious homology to other fungal cytb genes. Sequence analysis of Cbcytb was carried out in 32 Q_OI-sensitive (<0.080???g?ml^?1) and 27 Q_OI-resistant (>1.0???g?ml^?1) isolates. All tested Q_OI-resistant isolates harboured a point mutation in Cbcytb at nucleotide position 428 that conferred an exchange from glycine to alanine at amino acid position 143 (G143A). A PCR assay developed to discriminate Q_OI-sensitive and Q_OI-resistant isolates based on the G143A mutation could detect and differentiate isolates down to approximately 25?pg of template DNA. Microsatellite analyses suggested that Q_OI resistance emerged independently in multiple genotypic backgrounds at multiple locations. Our results indicate that Q_OI resistance has developed in some C. beticola populations in Italy and monitoring the G143A mutation is essential for fungicide resistance management in this pathosystem.