A total of 618 isolates of corynespora leaf spot fungus (Corynespora cassiicola) collected from 24 commercial cucumber greenhouses in 12 cities in Ibaraki Prefecture, Japan, were tested for their sensitivity to boscalid. Boscalid-resistant isolates were detected in 17 out of 19 greenhouses with a history of use of this fungicide and detection frequencies of the resistant isolates exceeded 47% in nine greenhouses. Frequencies of very highly resistant (VHR) isolates with 50% effective concentration (EC 50 ) values of boscalid exceeding 30 lg mL )1 were higher than those of moderately resistant (MR) isolates with EC 50 ranging from 2AE0 to 5AE9 lg mL )1 in 11 greenhouses. Additionally, highly resistant (HR) isolates with EC 50 from 8AE9 to 10AE7 lg mL )1 were first detected. Furthermore, molecular characterization of genes encoding succinate dehydrogenase (SDH) subunits (SdhA, SdhB, SdhC and SdhD) was carried out to elucidate the amino acid substitution responsible for the resistance to boscalid. All 23 VHR isolates had the same mutation from CAC to TAC in the SdhB gene leading to the substitution of histidine with tyrosine at amino acid position 278 (B-H278Y). At the same position, the substitution to arginine conferred by a mutation to CGC (B-H278R) was detected in all four HR isolates. Some MR isolates showed a substitution from serine to proline at position 73 in SdhC (C-S73P), from serine to proline or from glycine to valine at position 89 (D-S89P) and 109 (D-G109V), respectively, in SdhD. There was no common mutation in SDH genes of all MR isolates.
Differential sensitivity to boscalid, penthiopyrad and fluopyram, clearly found in these two important pathogens of cucumber, may indicate involvement of a slightly distinct site of action for fluopyram from the two other SDHIs. This finding may lead to the discovery of unique SDHIs in the future.
An experiment was conducted to examine the accumulation and mobility of heavy metals (Zn and Cu) at different depths in three types of arable soils (Brown Lowland soil, Andosol, and Brown Forest soil) amended with cattle and pig farmyard manures for 5 years. Nitric‐perchloric acid digestion was performed for the determination of the total amounts of heavy metals, and 0.1 M hydrochloric acid extraction was performed for the determination of the amounts of soluble heavy metals. Results of the soil analysis indicated that pig farmyard manure application resulted in serious contamination of arable soils with Zn and potentially Cu. Especially, the Brown Forest soil displayed a high ability to accumulate heavy metals on the soil surface. Total‐Zn concentration in surface soils was considerably affected by the holding capacity of soluble‐Zn traction. Although the Andosol amended with pig farmyard manure showed higher concentrations of heavy metals related to the higher ability of retention on a weight basis, the soil did not contribute to high heavy metal accumulation because of its low bulk density. Heavy metals were easily leached in sandy soils such as Brown Lowland soil, and Cu was potentially stable compared with Zn. We suggest that long‐term pig farmyard manure application to the Brown Lowland soil and Andosol with a light soil texture is associated with a higher risk of groundwater pollution than the application to the Brown Forest soil.
An experiment was conducted to determine the accumulation of trace metals, zinc (Zn), chromium (Cr), manganese (Mn) and iron (Fe), in three types of arable soils (Brown Lowland soil, Andosol and Brown Forest soil) amended with chemical fertilizers (CHF), cattle farmyard manure (CFM) and pig farmyard manure (PFM) over a 5-year period. Two methods were used to determine the total amount of trace metals: the instrumental neutron activation analysis (INAA), as a non-destructive measurement, and the concentrated acid digestion method (CADM), as a destructive measurement. Results of soil analyses using both INAA and CADM indicated that PFM application resulted in a considerable increase in the amount of Zn in arable soils compared with CHF application, unlike CFM application. Results obtained from the two methods were not fully in agreement for Cr. Based on INAA, soil accumulation of Cr from CFM and PFM applications was not significant in any soil type. In contrast, based on CADM, Cr accumulation was relatively higher in the Brown Forest soil with PFM than with CHF application. The soil types and treatments exerted various effects on Cr, Mn and Fe concentrations. The 5-year application of animal manure did not result in a considerable increase in Cr, Mn and Fe concentrations in soils, whereas longer-term application may lead to a gradual accumulation of Cr and Fe depending on the soil and manure types. The use of CADM in the present study may not always be adequate to assess the accumulation of trace metals in soils amended with CFM and PFM because of the conditions of the measurements, such as sensitivity, dilution level or type of instrument, and not because of the digestion method. The types of soil and manure affect trace metal accumulation in arable soils, and elements in the manure relate to the ability of soils to retain trace metals. It is important that the results of non-destructive and destructive measurements, obtained using INAA and CADM, be evaluated properly.
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