Ustilaginoidea virens is an important fungus that causes rice false smut disease. This disease significantly reduces both grain yield and quality. Various methods have been developed for the detection of U. virens but most of these methods need sophisticated equipment such as a thermal cycler. Here, we present a loop-mediated isothermal amplification (LAMP) assay for the specific detection of U. virens. This assay used a specific region of the UvG-β1 gene (212-bp region) to design six LAMP primers. The LAMP assay was optimized by the combination of rapidity, simplicity, and high sensitivity for the detection of about 1 pg of target genomic DNA in the reaction whereas, with polymerase chain reaction (PCR), there was no amplification of DNA with concentrations less than 1 ng. Among the genomic DNA of 22 fungus species and two strains of U. virens, only the tube containing the DNA of U. virens changed to yellowish green with SYBR Green I. The color change was indicative of DNA amplification. No DNA was amplified from either the other 22 fungus species or the negative control. Moreover, 20 spikelets and 22 rice seed samples were used for the detection of rice false smut via LAMP. The results were comparable with conventional PCR. We conclude that gene UvG-β1 coupled with LAMP assay, can be used for the detection and identification of U. virens gene via LAMP.

New fungicides are tools to manage fungal diseases and overcome emerging resistance in fungal pathogens. In this study, a total of 121 Fusarium fujikuroi isolates were collected from various geographical regions of China and their sensitivity to a novel succinate dehydrogenase inhibitor (SDHI) fungicide ‘pydiflumetofen’ was evaluated. The 50% effective concentration (EC₅₀) value of pydiflumetofen for mycelial growth suppression ranged from 0.0101 to 0.1012 μg/ml and for conidial germination inhibition ranged from 0.0051 to 0.1082 μg/ml. Pydiflumetofen-treated hyphae showed contortion and increased branching, cell membrane permeability, and glycerol content significantly. The result of electron microscope transmission indicated that pydiflumetofen damaged the mycelial cell wall and the cell membrane, and almost broke up the cells, which increased the intracellular plasma leakage. There was no cross-resistance between pydiflumetofen and the widely used fungicides such as carbendazim, prochloraz, and phenamacril. Pydiflumetofen was found safe to seeds and rice seedlings of four rice cultivars, used up to 400 μg/ml. Seed treatment significantly decreased the rate of diseased plants in the greenhouse as well as in field trials in 2017 and 2018. Pydiflumetofen showed superb results against the rice bakanae disease (RBD), when used at 10 or 20 g a.i./100 kg of treated seeds, providing over 90% control efficacy (the maximum control efficacy was up to 97%), which was significantly higher than that of 25% phenamacril (SC) at 10 g or carbendazim at 100 g. Pydiflumetofen is highly effective against F. fujikuroi growth and sporulation as well as RBD in the field.

Blueberry, Vaccinium corymbosum L. (cultivar Sharpblue, highbush blueberry), is becoming an important fruit that has become more widely planted in Anhui Province of China in recent years. In the early summer of 2014, twig dieback was observed on the blueberry plants with the incidence of 8 to 14%, in several commercial production fields of Huangshan district, Anhui Province. Necrosis initiating from the stem, was observed and multiple lesions coalesced together on the twigs, resulting in death of the branches. Abundant, black, and solitary acervuli were observed on symptomatic twigs. Pieces of tissue (3 mm in size) from infected twigs were cut and surface disinfested in 75% ethanol for 1 min and washed in sterile water three times before being plating onto potato dextrose agar (PDA) and incubated at 28°C. Resulting fungal cultures produced white aerial mycelia with a rough surface, turning slightly yellow after 7 to 10 days. The cultures were covered with black and globular acervuli with a diameter of 100.4 to 200.1 μm (ave. 164.7 μm). The base of each conidiophore was swollen and globose with phialides growing from the apical end. Mature conidia consisted of 3 to 5 cells that were straight to fusiform, with a tapering base and 2 to 4 hyaline appendages (apical appendages measured 23.1 to 28.4 µm (ave. 25.2 µm) long and a single basal appendage was 3.0 to 3.6 µm (ave.3.3 µm) long). Conidia were 19.3 to 26.1 × 5.0 to 6.5 µm (ave. 24.4 × 5.8 µm) with median cells and two hyaline, cylindrical to conical apical cells typical of Pestalotiopsis clavispora (Guba, 1961). A total of eight isolates were obtained by isolation from eight symptomatic twigs collected in eight fields in the Huangshan district. Amplification of the ITS1 and ITS4 internal transcribed spacer (ITS) regions with universal primers (Luan et al., 2008) yielded 519 bp amplicons that were sequenced (GenBank Accession Number KT758727) and analyzed together with other Pestalotiopsis clavisporase sequences. BlastN analysis of the ITS sequence obtained from all eight isolates showed 100% similarity to each other and over 99% similarity to the isolates of Pestalotiopsis clavispora deposited in Genbank (JN943634). To test pathogenicity, twigs collected from field grown blueberry cv. Sharpblue were inoculated by placing 7-day-old mycelial plugs from one representative isolate grown on PDA on the young bark and covered with cotton saturated with sterile water (one plug per twig, six twigs per isolate). Sterile PDA plugs were used as controls in both tests. The inoculated plants were kept in a humidity chamber for 14 days at 26 °C. After 4 days, the inoculated twigs developed red brown lesions. After 10 days, the whole twig blighted and showed similar symptoms to natural infections. No lesions were observed on the control twigs. Furthermore, a fungus was re-isolated from all inoculated twig tissues and identified to be Pestalotiopsis clavisporaas as described above, thus fulfilling Koch's postulates. This pathogen has been previously reported to cause leaf spot (Luan et al., 2008) and twig blight (Zhao et al. 2014) on blueberry in China. To our knowledge, this disease is an emerging problem in Anhui Province and will necessitate the development of new disease-management practices to sustain commercial production of blueberry in these regions. References: E.F. Guba. Monograph of Monochaetia and Pestalotia. Harvard University Press, Cambridge, MA, 1961. Y.S. Luan et al. Plant Dis. 92:171, 2008. H.H. Zhao et al. Mycosystema 33: 577, 2014.

Kiwifruit (Actinidia spp.) is a fruit cultivated in many countries, and China is the largest producer of commercial kiwifruit worldwide. In the spring of 2015, a leaf spot disease of kiwifruit was observed in several orchards in Huangshan City, Anhui Provence, China. Early foliar symptoms were characterized by yellow to brown, irregular-shaped lesions between the veins of the leaves. The lesions later expanded to the entire leaf. Infected leaves turned dark brown and withered. A small portion (< 1 cm2) of the symptomatic leaf tissues was surface sterilized in 75% ethanol for 30 s, then rinsed three times with sterile distilled water, and plated on the water agar. Five fungal isolates were recovered on potato dextrose agar (PDA) from different samples. The fungal colonies were initially white in color that later turned grayish brown with the onset of sporulation. The conidia were black in color, spherical to subspherical in shape and single-celled (10 to14 ×18 to 20 μm, n=100), which were born on a hyaline...

Pomegranate crown rot caused by Coniella granati is one of the most severe diseases of pomegranate. No fungicides have been registered for controlling this disease in China. Pyraclostrobin, belonging to strobilurin fungicides, has a broad spectrum of activity against many phytopathogens. In this study, based on the mycelial growth and conidial germination inhibition methods, we investigated the biological activity of pyraclostrobin against C. granati in the presence of 50 μg/ml of salicylhydroxamic acid using 80 isolates collected from different orchards in China from 2012 to 2018. The EC₅₀ (50% effective concentration) values ranged from 0.040 to 0.613 μg/ml for mycelial growth and 0.013 to 0.110 μg/ml for conidium germination. Treated with pyraclostrobin, the hyphae morphology changed and conidial production of C. granati decreased significantly. The result of transmission electron microscope showed that treatment of pyraclostrobin could make the cell wall thinner and lead to ruptured cell membrane and formation of intracellular organelle autophagosomes. The pyraclostrobin showed good protective and curative activities against C. granati on detached pomegranate fruits. In field trials, pyraclostrobin showed excellent control efficacy against this disease, in which the treatment of 25% pyraclostrobin EC 1,000× provided 92.25 and 92.58% control efficacy in 2019 and 2020, respectively, significantly higher than that of other treatments. Therefore, pyraclostrobin could be a candidate fungicide for the control of pomegranate crown rot.

Pomegranate (Punica granatum L.) is an important commercial fruit rich in antioxidants grown worldwide and China has the biggest planting area in the world. In spring 2019, symptoms of fruit rot we...