First Report Of Strobilurin Resistance In \u3ci\u3eCercospora Beticola\u3c/i\u3e In Sugar Beet (\u3ci\u3eBeta Vulgaris\u3c/i\u3e) In Michigan And Nebraska, USA

Cercospora leaf spot (CLS) caused by Cercospora beticola Sacc. is the most important foliar disease of sugar beet (Beta vulgaris) worldwide (Jacobsen & Franc, 2009). CLS is controlled mainly with fungicides, including strobilurins (FRAC group 11). Resistance to strobilurins in C. beticola has not been reported in the field (Secor et al., 2010) but insensitive mutations have been artificially developed (Malandrakis et al., 2011). In 2011, fields from several areas in Michigan, USA treated with strobilurins had severe CLS and diminished control was also noted in small plot trials (Fig. 1). Individual leaf spot lesions were sampled from leaves and grown on sugar beet leaf extract agar (SBLEA). A conidium germination bioassay was done on SBLEA covered with water agar amended with pyraclostrobin, azoxystrobin or trifloxystrobin at 0, 0.001, 0.01, 0.1, 1, 10, or 100 μg/ml, supplemented with salicylhydroxamic acid (SHAM) to block the alternate oxidation pathway (Olaya et al., 1998). After 24 h incubation at 22°C, under ambient light, percentage germinated conidia (n = 50) was calculated from three replicates per treatment. Germination was recorded as positive when the germ tube was at least half the length of the conidium. A representative wild type isolate was unable to germinate over the 0.01 μg/ml concentration. EC50 values for each isolate were calculated by regression analysis of percentage growth inhibition vs. the log fungicide concentration using Sigmaplot Version 9.01 (Systat Software, Chicago). The EC50 for the sensitive isolate was \u3c0.01 μg/ml. Isolates from several counties in Michigan had uninhibited germination and EC50 values exceeded the highest concentration tested. Isolates also grew on spiral gradient dilution plates (Förster et al., 2004) amended with the three strobilurins (Fig. 2, for illustration of resistance response only). Two isolates were obtained from Nebraska and each showed similar response to strobilurin fungicides in amended plate assays. Pure cultures of four resistant isolates were grown in potato dextrose broth at 125 rpm, and DNA extracted. A fragment of the cytochrome b (CYTB) gene was amplified by PCR using the C. beticola primers of Malandrakis et al. (2011) to amplify the region of the CYTB gene likely to contain resistance mutations (Malandrakis et al., 2011). This fragment was sequenced at the Genomics Technology Support Facility (MSU, East Lansing, MI) and showed 99% identity with both the C. beticola cytochrome b mRNA, partial sequence (GenBank Accession No. EF176921.1) and the C. kikuchii mitochondrial gene for cytochrome b partial sequence (AB231863.1). Sequence results revealed that each resistant isolate contained a change in codon 143 that predicts to a substitution of G143A, which was demonstrated to confer QoI resistance in several other fungi (Ma & Michailides, 2005). All four isolates with the G143A mutation germinated at 100 μg/ml pyraclostrobin (50% of conidia), while sensitive isolates that lacked the mutation failed to grow. Isolates that contained the G143A mutation included representatives from Michigan and Nebraska, USA. These findings reveal that reduced Cercospora leaf spot control in some commercial sugar beet fields may be due to the development of resistance to strobilurins

Image_1_Diverse environmental bacteria displaying activity against Phakopsora pachyrhizi, the cause of soybean rust.pdf

The management of soybean rust (SBR) caused by the obligate fungus Phakopsora pachyrhizi mostly relies on the use of synthetic fungicides, especially in areas where the disease inflicts serious yield losses. The reliance on synthetic fungicides to manage this disease has resulted in resistance of P. pachyrhizi populations to most fungicides. In this study, bacteria isolated from diverse environments were evaluated for their biocontrol potential against P. pachyrhizi using soybean detached-leaf method and on-plant in the growth chamber, greenhouse, and field. Among 998 bacterial isolates evaluated using the detached-leaf method; 58% were isolated from plant-related materials, 27% from soil, 10% from insects, and 5% from other environments. Of the isolates screened, 73 were active (they had ⪖ 75% rust reduction) with an active rate of 7.3%. From the active isolates, 65 isolates were re-tested on-plant in the growth chamber for activity confirmation. In the confirmation test, 49 bacteria isolated from plant-related materials maintained their activity with a confirmation rate of 75%. The majority of bacteria with confirmed activity belonged to the taxonomic classes Bacilli and Gammaproteobacteria (70%). Active isolates were prioritized for greenhouse and field testing based on activity in the initial screen and confirmation test. Six bacterial isolates AFS000009 (Pseudomonas_E chlororaphis), AFS032321 (Bacillus subtilis), AFS042929 (Bacillus_C megaterium), AFS065981 (Bacillus_X simplex_A), AFS090698 (Bacillus_A thuringiensis_S), and AFS097295 (Bacillus_A toyonensis) were selected from those bacteria that maintained activity in the confirmation test and were evaluated in the greenhouse, and five among them were evaluated in the field. From the Alabama field evaluation, all bacterial isolates reduced rust infection as well as azoxystrobin (Quadris® at 0.3 L/ha) used as the fungicide control (P > 0.05). Moreover, the scanning electron micrographs demonstrated evidence of antagonistic activity of AFS000009 and AFS032321 against P. pachyrhizi urediniospores. Bacterial isolates that consistently showed activity comparable to that of azoxystrobin can be improved through fermentation and formulation optimization, developed, and deployed. These bacteria strains would provide a valuable alternative to the synthetic fungicides and could play a useful role in integrated disease management programs for this disease.</p

Table_1_Diverse environmental bacteria displaying activity against Phakopsora pachyrhizi, the cause of soybean rust.pdf

The management of soybean rust (SBR) caused by the obligate fungus Phakopsora pachyrhizi mostly relies on the use of synthetic fungicides, especially in areas where the disease inflicts serious yield losses. The reliance on synthetic fungicides to manage this disease has resulted in resistance of P. pachyrhizi populations to most fungicides. In this study, bacteria isolated from diverse environments were evaluated for their biocontrol potential against P. pachyrhizi using soybean detached-leaf method and on-plant in the growth chamber, greenhouse, and field. Among 998 bacterial isolates evaluated using the detached-leaf method; 58% were isolated from plant-related materials, 27% from soil, 10% from insects, and 5% from other environments. Of the isolates screened, 73 were active (they had ⪖ 75% rust reduction) with an active rate of 7.3%. From the active isolates, 65 isolates were re-tested on-plant in the growth chamber for activity confirmation. In the confirmation test, 49 bacteria isolated from plant-related materials maintained their activity with a confirmation rate of 75%. The majority of bacteria with confirmed activity belonged to the taxonomic classes Bacilli and Gammaproteobacteria (70%). Active isolates were prioritized for greenhouse and field testing based on activity in the initial screen and confirmation test. Six bacterial isolates AFS000009 (Pseudomonas_E chlororaphis), AFS032321 (Bacillus subtilis), AFS042929 (Bacillus_C megaterium), AFS065981 (Bacillus_X simplex_A), AFS090698 (Bacillus_A thuringiensis_S), and AFS097295 (Bacillus_A toyonensis) were selected from those bacteria that maintained activity in the confirmation test and were evaluated in the greenhouse, and five among them were evaluated in the field. From the Alabama field evaluation, all bacterial isolates reduced rust infection as well as azoxystrobin (Quadris® at 0.3 L/ha) used as the fungicide control (P > 0.05). Moreover, the scanning electron micrographs demonstrated evidence of antagonistic activity of AFS000009 and AFS032321 against P. pachyrhizi urediniospores. Bacterial isolates that consistently showed activity comparable to that of azoxystrobin can be improved through fermentation and formulation optimization, developed, and deployed. These bacteria strains would provide a valuable alternative to the synthetic fungicides and could play a useful role in integrated disease management programs for this disease.</p

Tackling agricultural diffuse pollution: What might uptake of farmer-preferred measures deliver for emissions to water and air?

Mitigation of agricultural diffuse pollution poses a significant policy challenge across Europe and particularly in the UK. Existing combined regulatory and voluntary approaches applied in the UK continue to fail to deliver the necessary environmental outcomes for a variety of reasons including failure to achieve high adoption rates. It is therefore logical to identify specific on-farm mitigation measures towards which farmers express positive attitudes for higher future uptake rates. Accordingly, a farmer attitudinal survey was undertaken during phase one of the Demonstration Test Catchment programme in England to understand those measures towards which surveyed farmers are most receptive to increasing implementation in the future. A total of 29 on-farm measures were shortlisted by this baseline farm survey. This shortlist comprised many low cost or cost-neutral measures suggesting that costs continue to represent a principal selection criterion for many farmers. The 29 measures were mapped onto relevant major farm types and input, assuming 95% uptake, to a national scale multi-pollutant modelling framework to predict the technically feasible impact on annual agricultural emissions to water and air, relative to business as usual. Simulated median emission reductions, relative to current practise, for water management catchments across England and Wales, were estimated to be in the order sediment (20%) &gt; ammonia (16%) &gt; total phosphorus (15%) ? nitrate/methane (11%) &gt; nitrous oxide (7%). The corresponding median annual total cost of the modelled scenario to farmers was £3 ha? 1 yr? 1, with a corresponding range of -£84 ha? 1 yr? 1 (i.e. a net saving) to £33 ha? 1 yr? 1. The results suggest that those mitigation measures which surveyed farmers are most inclined to implement in the future would improve the environmental performance of agriculture in England and Wales at minimum to low cost per hectare