A quantitative real‐time PCR assay for detection of Colletotrichum lindemuthianum in navy bean seeds

Bean anthracnose is a seedborne disease of common bean (Phaseolus vulgaris) caused by the fungal pathogen Colletotrichum lindemuthianum. Using seed that did not test positive for the pathogen has been proven to be an effective strategy for bean anthracnose control. To quantify the extent of anthracnose seed infection, a real‐time PCR‐based diagnostic assay was developed for detecting C. lindemuthianum in seeds of the commercial bean class navy bean. The ribosomal DNA (rDNA) region consisting of part of the18S rDNA, 5^.8S rDNA, internal transcribed spacers (ITS) 1, 2 and part of the 28S rDNA of seven races of C. lindemuthianum, 21 isolates of Colletotrichum species and nine other bean pathogens were sequenced with the universal primer set ITS5/ITS4. Based on the aligned sequence matrix, one primer set and a probe were designed for a SYBR Green dye assay and a TaqMan MGB (minor groove binder) assay. The primer set was demonstrated to be specific for C. lindemuthianum and showed a high sensitivity for the target pathogen. The detection limit of both assays was 5 fg of C. lindemuthianum genomic DNA. To explore the correlation between the lesion area and the DNA amount of C. lindemuthianum in bean seed, seeds of the navy bean cultivar Navigator with lesions of different sizes, as well as symptomless seeds, were used in both real‐time PCR assays.     

Resistance to Rhizoctonia solani AG‐2‐2 (IIIB) in creeping bentgrass plants transformed with pepper esterase gene PepEST

A pepper esterase (PepEST) gene was introduced into creeping bentgrass (Agrostis stolonifera) by Agrobacterium‐mediated transformation. Purified recombinant PepEST proteins were sufficient to inhibit the growth of the fungal pathogens Rhizoctonia solani AG2‐2 (IIIB) (causing brown patch) and Sclerotinia homoeocarpa (dollar spot), but not the oomycete responsible for pythium blight, Pythium aphanidermatum. PepEST proteins were most effective against R. solani. After genetic transformation of creeping bentgrass with PepEST, the genomic integration of transgenes bar and PepEST was confirmed by Southern blot analysis, and their expression was also validated by northern blot and western blot analyses. Disease severity on R. solani‐inoculated leaves of transgenic plants was <10% compared to ca. 50% in non‐transgenic plants. Microscopic observation of infected leaves indicated that PepEST inhibited the growth of hyphae upon fungal infection.     

Direct evidence of surface infestation of seeds and tubers by Plasmodiophora brassicae and quantification of spore loads

Using quantitative PCR, DNA of Plasmodiophora brassicae, the causal agent of clubroot, was detected and quantified on canola, pea and wheat seeds, as well as on potato tubers, all harvested from clubroot‐infested fields in Alberta, Canada. Quantifiable levels of infestation were found on seven of the 46 samples analysed, and ranged from <1·0 × 10³ to 3·4 × 10⁴ resting spores per 10 g seeds; the vast majority (80–100%) of resting spores on these samples were viable, as determined by Evan’s blue vital staining. However, the levels of infestation found were generally lower than that required to cause consistent clubroot symptoms in greenhouse plant bioassays. While the occurrence of P. brassicae resting spores on seeds and tubers harvested from clubroot‐infested fields suggests that seedborne dissemination of this pathogen is possible, practices such as commercial seed cleaning may be sufficient to effectively mitigate this risk.     

颗粒剂防治玉米钻心虫的研究

1962年在北京夏玉米上用顆粒剂防治玉米钻心虫的結果表明:心叶末期施5％DDT顆粒剂,防治一次,每株2克,在一般年份,从产量上来看是很有利的。同时对药剂残效,幼虫在玉米不同生育期植株上的分布,以及顆粒剂在心叶期施药至抽雄后在植株上的分布测定,并結合顆粒剂小区防治試驗。分析結果说明:心叶末期施药,影响药效的主要原因,不是5％DDT残效的問題,而是抽雄后药剂在植株上部分布較少,尤其是包在雄穗节外的第一叶腋没有药,而此时期,幼虫在植株上部分布較多,同时DDT速效也不如666。     

Assessment of the impact of resistant and susceptible canola on Plasmodiophora brassicae inoculum potential

The impact on clubroot severity of growing susceptible canola or mixtures of resistant and susceptible canola genotypes was examined. Bioassays revealed greater clubroot severity and incidence, and reduced plant height, where 100% of a susceptible cultivar had been grown. A higher proportion of susceptible plants within a resistant canola crop increased root hair and secondary infections. Regression analysis of root hair infection and the amount of Plasmodiophora brassicae DNA (as determined by quantitative PCR) revealed strong linear relationships between the two parameters. The linear relationships between root hair infection and P. brassicae DNA were stronger for the resistant cultivar than for the susceptible cultivar when regression analysis was conducted by cultivar over the sampling dates. In conclusion, the cropping of a resistant cultivar reduced clubroot severity, while the presence of susceptible volunteer canola increased inoculum potential. Quantitative PCR was a reliable tool for the quantification of root hair infection.     

Effects of Erysiphe pisi on protein profiles and ribulose-1,5-bisphosphate carboxylase content in resistant and susceptible pea (Pisum sativum × Pisum fulvum) plants

Journal of Plant Diseases and Protection - The fate of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) was examined in leaves of both resistant and susceptible plants from a Pisum sativum...     

Differences in the growth stages of Erysiphe pisi on cultivars of field pea (Pisum sativum L.)

Journal of Plant Diseases and Protection - The infection process of Erysiphe pisi on field pea cvs. ‘Highlight’ and ‘Tara’ carrying resistance gene erl, line...     

Quantifying resistance to Plasmodiophora brassicae in Brassica hosts

Plasmodiophora brassicae causes clubroot of crucifers. A quantitative PCR (qPCR)‐based protocol was developed to measure P. brassicae DNA in the roots of susceptible, intermediately susceptible, intermediately resistant and resistant Brassica hosts, and the non‐host wheat, at 5, 10, 15, 20 and 42 days post‐inoculation (dpi). The final reaction of each plant genotype was recorded as an index of disease at 42 dpi. Plasmodiophora brassicae DNA showed an increase in susceptible and moderately resistant hosts from 5 to 42 dpi, in contrast to a decrease in a highly resistant host and the non‐host wheat over the same period. Index of disease was significantly positively correlated with the amount of P. brassicae DNA in the roots at 5, 15, 20 and 42 dpi in one experiment, and at 10, 15, 20 and 42 dpi in a repeated experiment. Significant positive correlations also existed between the amounts of P. brassicae DNA in the roots at 42 dpi and those at 5, 10, 15 and 20 dpi in one experiment, and those at 10, 15 and 20 dpi in a repeated experiment. The results generated by the qPCR assay were validated by microscopic examination of roots inoculated with P. brassicae. The qPCR‐based protocol developed in this study allows for the accurate quantification of P. brassicae DNA in host root tissues as early as 5 dpi, and may serve as a useful tool to evaluate pathogen proliferation and development in the roots.     

Influence of prohexadione-calcium on growth and gibberellins content of Chinese cabbage grown in alpine region of South Korea

We investigated the effect of prohexadione-calcium (Pro-Ca) on growth characteristics and endogenous gibberellins content of Chinese cabbage grown in the alpine region of South Korea. Pro-Ca was applied at the rates of 200 ppm and 400 ppm, after 10, 15 and 20 days of transplanting seedlings in to the field. Application of Pro-Ca through foliage improved quality and quantity of yield by promoting Chinese cabbage head yield, number of head leaves, total soluble sugar content and compactness of head. The leaf size was reduced, while the chlorophyll content increased under the influence of elevated Pro-Ca application, when measured after 40 days of transplantation. The endogenous bioactive GA₁ and GA₄ contents of Chinese cabbage drastically decreased with elevated Pro-Ca, indicating that gibberellins (GAs) biosynthesis was blocked by this chemical. Current study suggests that both of GAs biosynthesis pathways are operational in Chinese cabbage, although non-C13-hydroxylation pathway was found to be the major pathway. GAs were quantified by gas chromatography/mass spectroscopy-selective ion monitoring (GC/MS-SIM).