Effect of treating soybean with 2,6-dichloroisonicotinic acid (INA) and benzothiadiazole (BTH) on seed yields and the level of disease caused by Sclerotinia sclerotiorum in field and greenhouse studies

Field or greenhouse grown soybeans were treated with 2,6-dichloroisonicotinic acid or benzothiadiazole and subsequently assessed for severity of white mold disease caused by Sclerotinia sclerotiorum. Three or four applications of 2,6-dichloroisonicotinic acid to field plots in 1993–1995 reduced severity of white mold after natural infection by 20–70% compared with water-treated controls in soybean cultivars Elgin 87 and Williams 82, which are considered to be highly susceptible to the disease. The effect was not as large in the cultivars Corsoy 79 and NKS19-90 which are more resistant to white mold. Two or four applications of benzothiadiazole to field plots in 1995 and 1996 reduced white mold severity by 20–60%, with the greatest reductions again observed in the more susceptible cultivars. Corresponding yields were increased compared with controls, particularly for the susceptible cultivars under conditions of high disease pressure. In greenhouse trials multiple applications of either compound resulted in significantly smaller lesion diameters following subsequent leaf inoculations with the fungus. The compounds did not result in observable phytotoxicity or inhibit growth of Sclerotinia sp. in vitro. We hypothesize that the decrease in disease severity following treatment with INA or BTH is a result of resistance induction.     

Prevalence of sclerotinia stem rot of soybeans in the north-central United States in relation to tillage, climate, and latitudinal positions

ABSTRACT Since the early 1990s, Sclerotinia stem rot, caused by Sclerotinia sclerotiorum, has caused considerable damage to soybean production in the north-central United States. To determine the extent of its distribution and associated factors, investigations were conducted in 1995 and 1996 in Illinois, Iowa, Minnesota, Missouri, and Ohio. Investigations also were conducted in 1997 and 1998 in Iowa, Minnesota, and Missouri. In each state, soybean fields were randomly selected in collaboration with the National Agricultural Statistics Service. From each field, 20 soybean stems 20 cm long (from the base) in 1995 and 1996 and full-length stems in 1997 and 1998 were sampled in a zigzag pattern. During the 4-year period, stem samples were collected from 1,983 fields and assessed for the presence or absence of the disease. Of the five states, Sclerotinia stem rot was most prevalent in north-central Iowa and southern Minnesota. Sclerotinia stem rot was not detected in Missouri during the 4-year investigation period. The disease was most prevalent in 1996 and least prevalent in 1995. The prevalence of the disease was strongly related to cumulative departures from normal maximum and minimum temperatures in July and August. The disease was more prevalent when yearly temperatures were below normal than when they were above normal. In 1996, a year with a cooler-than-normal summer, the disease was detected farther south than in 1995. In both years, the prevalence of the disease was exponentially related to latitudinal positions of the fields (R(2) = 0.93 and 0.83 for 1995 and 1996, respectively) reflecting the effect of the north-south variations in temperature. During the 4-year period, there was no relationship between precipitation and the prevalence of the disease. The lack of relationship may suggest that there was no shortage of moisture since it is one of the primary factors for disease development. The prevalence of Sclerotinia stem rot was less in no-till than in minimum-till or conventional-till fields (P = 0.001 and 0.007, respectively) and greater in minimum-till than in conventional-till fields (P = 0.07). Fields that had Sclerotinia stem rot, however, did not differ in incidence of the disease regardless of the tillage system.     

Sclerotinia rot losses in processing tomatoes grown under centre pivot irrigation in central Brazil

Sclerotinia rot caused by Sclerotinia sclerotiorum is one of the most important diseases of processing tomatoes in Central Brazil. Yield losses in tomato cultivars (cv.) IPA-5 were assessed in mature plants from 1995 to 1997, and related to different disease intensities, in a naturally infested area irrigated by centre pivot. Over the 3 years, there were no differences (Tukey at 5%) in fruit numbers between plants without symptoms (NS) and with intermediate symptoms (IS), which yielded higher numbers than plants with severe symptoms (SS). The greatest reduction in fruit number was 56.8% in 1997. Significant differences were observed in fruit weight and size among NS, IS and SS plants in 1995 and 1997. In 1996, NS and IS plants were similar, but different from SS, which yielded significantly less. Weight and size reductions in SS plants reached 84.3% and 62.0%, respectively, in 1997. In 1996 and 1997, yield losses related to time of symptom appearance and physiological age were also assessed. Significant correlations were found ( P  < 0.01), with nearly total losses observed when plants were infected from early to mid bloom, as opposed to plants infected near harvest, which had lower disease incidence and produced economically acceptable yields. Quadratic and exponential models best fitted the relationship between yield and time of symptom appearance, and yield vs physiological age could be explained by logistic and Gompertz functions.     

Assessment of the impact of mid-season late blight infection on disease development and yield of potato variety Russet Norkotah in Maine

Development of late blight of potatoes caused by Phytophthora infestans (US 8 fungal genotype, A2 mating type) was monitored in two Russet Norkotah commercial fields at Fort Fairfield in 1996 and Duncan Farm in 1997. Experimental plots representing various disease treatments (low, moderate, high and random late blight severity levels) were established in two fields in 1996. In 1997, only low and high disease treatments were established. The application of fungicides for late blight control was conducted in both years. Late blight incidence and severity were assessed in each plot of each treatment. Components of late blight disease development, tuber blight incidence and tuber yields were determined from each plot. Progress of late blight disease was rapid in 1996 but not in 1997. During the 1996 cropping season, mean disease incidence and severity in the random disease treatment plots were 84 and 21% respectively within 10 days of disease detection. In 1997, low levels of late blight severity were detected in the field plots. Average numbers of late blight leaf and stem lesions on infected plant and fungal sporangia on the diseased leaf were not significantly affected by disease treatment. Late blight foliar severity significantly affected potato tuber yields. Lowest tuber yield was obtained in plots with high disease levels and highest yields were recorded in plots with low late blight severity in 1996. Late blight severity was significantly correlated with tuber yield but not with per cent tuber blight.     

Phytoalexin accumulation during infection of bean and soybean by ascospores and mycelium of Sclerotinia sclerotiorum

Infection by ascospores of Sclerotinia selerotiorum caused hypersensitivity in epidermal cells in leaves and etiolated hypocotyls of bean and soybean. In bean, phaseollin and phaseollidin accumulated in leaves but kievitone alone in hypocotyls. In soybean, no phytoalexins were detected in leaves but glyceollin accumulated in hypocotyls.
Mycelial infection caused water-soaked spreading lesions in leaves and etiolated hypocotyls of both hosts. In bean, no phytoalexins were detected in leaves but kievitone alone accumulated in hypocotyls. In soybean, glyceollin accumulated in leaves but was not sought in hypocotyls.
Transfer of bean hypocotyls infected with mycelium from 18 to 28°C caused lesion limitation and marked accumulation of phaseollin and kievitone.
Phaseollin, kievitone and glyceollin inhibited ascospore germination and growth of hyphae from preformed germ-tubes and established mycelia, phaseollin being most active and glyceollin least active. Hyphal growth from mycelia was least affected by the phytoalexins.     

Biocontrol of sclerotinia stem rot (Sclerotinia minor) in sunflower by seed treatment with Gliocladium virens

An isolate of Gliocladium virens from disease-affected soil in a commercial sunflower field proved highly antagonistic to Sclerotinia minor in culture. Significant disease control was obtained when a conidial suspension was applied over a limited area of potting medium and lateral roots were inoculated outside the treated area at the edge of the pot. Most promising, in terms of minimizing the amount of G. viren s required for disease control, was the significant bioprotection obtained by seed treatment. In glasshouse experiments following inoculation of lateral roots with S. minor at 6–7 weeks after sowing, disease incidence was reduced from 60.0% to 15.8% in potting medium and from 66.7% to 12.5% in field soil. Seed treatment was also effective in field microplots, where disease incidence was reduced from 37.0% to 22.2%. G. viren s applied on seed did not affect the dry weight of the lateral root system.     

Effect of Coniothyrium minitans on sclerotial survival and apothecial production of Sclerotinia sclerotiorum in field-grown oilseed rape

In two field trials with oilseed rape, Coniothyrium minitans was applied to soil as a maizemeal-perlite preparation in order to determine its effect on sclerotial survival and apothecial production of Sclerotinia selerotiorum. The mycoparasite infected sclerotia and decreased sclerotial survival, carpogenic germination and production of apothecia. Effects were greatest when inoculum of C. minitans was applied in autumn, at the time of sowing, rather than when it was applied in spring. C. minitans survived in soil for 2 years and spread to adjacent control plots and infected sclerotia within those plots. However, despite the fact that the inoculum potential of S. selerotiorum was reduced by C. minitans treatment, no disease control was obtained either in trial 1, where disease levels were low (0-20% of plant stems affected), or in trial 2, where disease levels were high (up to 70% of plant stems affected). Possible reasons for this failure of C minitans to control sclerotinia disease in oilseed rape, and strategies to improve its efficacy in the field, are discussed.     

In vitro Effect of the Phytoalexin Glyceollin I on Polygalacturonase and Oxalic Acid Production ofSclerotinia sclerotiorum

The polygalacturonase (PG) activity in culture filtrates fromSclerotinia sclerotiorum is reduced when glyceollin I, the major soybean phytoalexin, is present in the culture medium. When the enzyme activities in the culture filtrates are expressed per unit of fungal growth, PG activity decreases with increasing concentration of glyceollin I in the culture medium. The phytoalexin does not influence the isoenzyme pattern. This suggests that glyceollin I may inhibit quantitative but not qualitative enzyme production. Only the highest glyceollin I concentration tested inhibits oxalic acid production. The inhibitory effect on mycelial growth is confirmed. The data suggest a further hypothesis about the role of phytoalexin during pathogenesis.     

Determination of agronomic practices for the management of blight of chickpea caused by <Emphasis Type="Italic">Ascochyta rabiei</Emphasis> in Turkey: 1. Appropriate sowing date

In order to determine the most appropriate dates for planting chickpea in central Anatolia, Turkey, six cultivars were planted at three sites that differed in disease pressure. In two of the sites, disease pressure from Ascochyta rabiei was promoted by spreading infected chickpea debris on the soil surface at the time of planting and, at one of these, sprinkle irrigation was applied. In the third site, where conditions were dryer, no artificial inoculum was provided. Plants from seeds sown in early March had the most disease and in the sprinkle irrigated plots the disease severity ranged from 7.8 on the most susceptible cv. Canitez to 3.3 on the least susceptible Gokce as scored on the 1–9 scale where 1 = no disease and 9 represents a plant killed by the fungus. There was an inverse relationship between disease severity and yield, production from blight resistant cultivars of around 2,000 kg ha⁻¹ being more than twice that of susceptible ones. Delaying planting for 3–5 weeks reduced the severity of ascochyta blight but also reduced the yields in four of the six cultivars. In contrast, reduction in disease severity by delayed sowing resulted in yield increases for the susceptible cvs Canitez and Local, although yield level was not as much as those of the less susceptible cvs sown early. Delay of 6–9 weeks almost eliminated ascochyta blight but yields of all cultivars were seriously compromised by drought stress. In consequence, chickpea farmers are recommended to use resistant or tolerant cultivars and sow early in March. For less resistant cultivars, sowing in early April is recommended. Further delay is not recommended unless irrigation is provided and fungicide spraying is recommended where signs of infection are present under conditions conducive to the disease.     

Impact of time between field application of Bacillus subtilis strains SB01 and SB24 and inoculation with Sclerotinia sclerotiorum on the suppression of Sclerotinia stem rot in soybean

This study evaluated the impact of time between the application of cell suspensions or cell-free filtrates of Bacillus subtilis strains SB01 or SB24 on soybean plants under field conditions and inoculation with Sclerotinia sclerotiorum on their effectiveness for suppression of S. sclerotiorum. The results showed that the cell suspensions of two strains provided greater effectiveness than the cell-free filtrates, but the suppression effectiveness decreased as the time between application in the field and S. sclerotiorum inoculation increased. The B. subtilis cell suspensions applied on soybean leaves for up to 10 days under field conditions were able to provide a significant (P < 0.01) reduction in disease severity by approximately 20–90% at 5 days after the S. sclerotiorum inoculation. When rated 15 days after S. sclerotiorum inoculation, plants treated with bacterial cells for ≤6 days reduced Sclerotinia stem rot severity by 15–70%. Most effectiveness was provided by the cell suspensions present on soybean leaves for <3 days under field conditions, which significantly (P < 0.01) reduced disease severity by 40–70% over 15 days. In comparison, the cell-free filtrates remaining on leaves for <6 days significantly (P < 0.01) reduced disease severity during the first 5 days after the inoculation, while the best cell-free filtrate treatments were those with ≤1-day intervals, which significantly (P < 0.01) reduced disease severity by 10–40% during 15 days after the inoculation. The effectiveness of B. subtilis was reduced when it rained after application.     

Effects of Herbicides on Fusarium solani f. sp. glycines and Development of Sudden Death Syndrome in Glyphosate-Tolerant Soybean

ABSTRACT Sudden death syndrome of soybean, caused by Fusarium solani f. sp. glycines, is a disease of increasing economic importance in the United States. Although the ecology of sudden death syndrome has been extensively studied in relation to crop management practices such as tillage, irrigation, and cultivar selection, there is no information on the effects of herbicides on this disease. Three herbicides (lactofen, glyphosate, and imazethapyr) commonly used in soybean were evaluated for their effects on the phenology of F. solani f. sp. glycines and the development of sudden death syndrome in four soybean cultivars varying in resistance to the disease and in tolerance to glyphosate. Conidial germination, mycelial growth, and sporulation in vitro were reduced by glyphosate and lactofen. In growth-chamber and greenhouse experiments, there was a significant increase in disease severity and frequency of isolation of F. solani f. sp. glycines from roots of all cultivars after application of imazethapyr or glyphosate compared with the control treatment (no herbicide applied). Conversely, disease severity and isolation frequency of F. solani f. sp. glycines decreased after application of lactofen. Across all herbicide treatments, severity of sudden death syndrome and isolation frequency were lower in disease-resistant than in susceptible cultivars. Results suggest that glyphosate-tolerant and -nontolerant cultivars respond similarly to infection by F. solani f. sp. glycines after herbicide application.     

Biocontrol of sclerotinia stem rot (Sclerotinia sclerotiorum) of soybean using novel Bacillus subtilis strain SB24 under control conditions

Sclerotinia stem rot (SSR), caused by Sclerotinia sclerotiorum, is a major disease of soybean in Canada. Laboratory and greenhouse experiments were conducted to evaluate potential effectiveness of cell suspensions, cell‐free culture filtrates and broth cultures of Bacillus subtilis strain SB24 for suppression of SSR. The SB24 cell suspensions and cell‐free culture filtrates significantly reduced mycelial growth of S. sclerotiorum by 50 to 75% and suppressed sclerotial formation by > 90%. The severity on soybean was negatively correlated (r < ?0·84, P < 0·01) to the concentrations of cell suspension, cell‐free culture filtrate and broth culture applied. The cell suspension and broth culture preparations significantly (P < 0·01) reduced SSR severity by 45 to 90% at concentrations ranging from 5 × 10⁶ to 10⁹ CFU mL^?1. The most effective concentration was 5 × 10⁸ CFU mL^?1 for all three preparations, reducing the severity by 60 to 90%. The B. subtilis SB24 was most effective in reducing disease severity when applied ≤ 24 h before plant inoculation with S. sclerotiorum and a significant effectiveness was observed up to 15 days after plant inoculation. The population density of B. subtilis on soybean leaves decreased by 1·5 to 2·5 log units over 15 days under field conditions, and by 0·8 log units over 5 weeks under control conditions. The decrease in population density was significantly correlated with rainfall in the field (r < ?0·93, P < 0·01), suggesting that the biocontrol bacteria may be washed away by rain.     

Biological Control of Pathogens Causing Root Rot Complex in Field Pea Using Clonostachys rosea Strain ACM941

ABSTRACT Pea root rot complex (PRRC), caused by Alternaria alternata, Aphanomyces euteiches, Fusarium oxysporum f. sp. pisi, F. solani f. sp. pisi, Mycosphaerella pinodes, Pythium spp., Rhizoctonia solani, and Sclerotinia sclerotiorum, is a major yield-limiting factor for field pea production in Canada. A strain of Clonostachys rosea (syn. Gliocladium roseum), ACM941 (ATCC 74447), was identified as a mycoparasite against these pathogens. When grown near the pathogen, ACM941 often was stimulated to produce lateral branches that grew directly toward the pathogen mycelium, typically entwining around the pathogen mycelium. When applied to the seed, ACM941 propagated in the rhizosphere and colonized the seed coat, hypocotyl, and roots as the plant developed and grew. ACM941 significantly reduced the recovery of all fungal pathogens from infected seed, increased in vitro seed germination by 44% and seedling emergence by 22%, and reduced root rot severity by 76%. The effects were similar to those of thiram fungicide, which increased germination and emergence by 33 and 29%, respectively, and reduced root rot severity by 65%. When soil was inoculated with selected PRRC pathogens in a controlled environment, seed treatment with ACM941 significantly increased emergence by 26, 38, 28, 13, and 21% for F. oxysporum f. sp. pisi, F. solani f. sp. pisi, M. pinodes, R. solani, and S. sclerotiorum, respectively. Under field conditions from 1995 to 1997, ACM941 increased emergence by 17, 23, 22, 13, and 18% and yield by 15, 6, 28, 6, and 19% for the five respective pathogens. The seed treatment effects of ACM941 on these PRRC pathogens were greater or statistically equivalent to those achieved with thiram. Results of this study suggest that ACM941 is an effective bioagent in controlling PRRC and is an alternative to existing chemical products.     

Control of take-all of wheat in the field with benzimidazole and triazole fungicides applied at seeding

Control of take-all ( Gaeumannomyces graminis var. tritici ) of wheat, with fungicides applied at seeding, was investigated at three naturally infested field sites. Fungicides were applied with seed either as granules or as pellets at rates ranging from 1-5 to 140 kg a.i./ha. Treatment with benzimidazole and triazole fungicides resulted in lower disease severity on seminal roots, up to 100% fewer deadheads and more than 200% greater yields.     

Predicting epidemics of yellow rust (Puccinia striiformis) on the upper canopy of wheat from disease observations on lower leaves

Data from field experiments were used to test whether disease observations on lower leaves of wheat were good predictors of future epidemic development in the upper canopy. Fungicide treatments to replicated plots in 1994 and 1995 caused variation in levels of initial inoculum of urediniospores of yellow rust ( Puccinia striiformis ). Observations of symptom severity were made on individual leaf layers throughout the season. Sporulating lesions on lower leaves were considered to be a measure of inoculum source strength for transfer to upper culm leaves. Low source strengths were associated with delays in epidemic development on the upper leaves, as quantified by a location parameter, t _m, the time at which severity reached half of the asymptote value in logistic fits to disease progress data on the upper leaves. However, there was much unexplained variation, probably due to extraneous variation in upward transfer efficiency of inoculum and rates of epidemic development. Vanderplank's sanitation ratio theory was used to account for variation in inoculum transfer and epidemic rate. The analysis revealed that if the aim is to predict disease on a newly emerged culm leaf, during the period when it can be treated effectively with fungicide, then observations of disease two leaves further down the culm were of the greatest predictive value ( R ² = 86%). These observations, however, need to be integrated with information about factors affecting upward inoculum transfer and rates of epidemic development if acceptable predictive precision is to be achieved.     

Environmental Factors Affecting Production, Release, and Field Populations of Conidia of Alternaria alternata, the Cause of Brown Spot of Citrus

ABSTRACT Alternaria brown spot, caused by Alternaria alternata pv. citri, affects many tangerines and their hybrids, causing loss of immature leaves and fruit and reducing the marketability of the remaining fruit. Conidial production of A. alternata was greatest on mature leaves moistened and maintained at near 100% relative humidity (RH) for 24 h, whereas leaves that had been soaked or maintained at moderate RH produced few conidia. Conidial release from filter paper cultures and infected leaves was studied in a computer-controlled environmental chamber. Release of large numbers of conidia was triggered from both substrates by sudden drops in RH or by simulated rainfall events. Vibration induced release of low numbers of conidia, but red/infrared irradiation had no effect. In field studies from 1994 to 1996, air sampling with a 7-day recording volumetric spore trap indicated that conidia were present throughout the year with periodic large peaks. The number of conidia captured was not closely related to rainfall amounts or average wind speed, but was weakly related to the duration of leaf wetness. Likewise, disease severity on trap plants placed in the field weekly during 1995 to 1996 was not closely related to conidial numbers or rainfall amounts, but was weakly related to leaf wetness duration. Sufficient inoculum appears to be available to allow infection to occur throughout the year whenever susceptible host tissue and moisture are available.     

Temporal and quantitative analyses of stem lesion development and foliar disease progression of peach rust in california

ABSTRACT The development of rust epidemics caused by Tranzschelia discolor on leaves and stems of cling peach was studied in California orchards. Sporulating stems lesions were only detected from late March until July in 1997 and 1998. When rust was present in the fall, the quadratic equation Y = -82.51 + 1.97JD - 0.01JD(2) using Julian day (JD) described the incidence of sporulating lesions on stems of cv. Andross (R(2) = 0.73; P /=117.3 mm of total precipitation and maximum temperatures of 相似文献   

Evaluation of soil solarization for control of root diseases of row crops in Victoria

The effect of soil solarization on the viability of plant pathogens and disease was evaluated in Victoria. The treatment was tested in NW and S Victoria with natural soil inoculated with high inoculum levels of Eusarium oxysporum, Plasmodiophora brassicae, Sclerotium cepivorum, Sclerotinia minor, Sclerotinia sclerotiorum, Verticillium dahliae and the nematodes Meloidogyne javanica and Pratylenchus penetrans. Other experiments were established at sites with a previous history of disease.
Solarization of artificially inoculated soils reduced inoculum levels to at least a depth of 10 cm and effectively controlled diseases caused by P. brassicae on broccoli, and S. minor and S. sclerotiorum on lettuce. The treatment reduced inoculum levels but not disease of carnations and watermelons affected by E. oxysporum , tomatoes affected by M. javanica , celery affected by P. penetrans , and onions affected by S. cepivorum. Results were inconclusive for tomatoes affected by V. dahliae.
Experiments in naturally infested soils established that solarization reduced disease and increased yields of Chinese cabbage affected by P. brassicae , celery affected by P. penetrans , lettuce affected by S. minor and watermelon affected by root rot.
Solarization reduced disease of onions affected by S. cepivorum but did not significantly increase yield. At all sites the treatment reduced the number of viable propagules of the pathogens to at least a depth of 10 cm.     

Using crop canopy modification to manage plant diseases

Modifying crop canopies can suppress plant diseases in some crops. For example, in carrot, lateral trimming of the canopy by 30–40 % after canopy closure reduced sclerotinia rot (Sclerotinia sclerotiorum) to zero under moderate disease pressure without the use of fungicides. Trimming reduced relative humidity within the carrot canopy and increased air and soil temperature, inhibiting the formation of apothecia of S. sclerotiorum. Trimming also severed infected petioles, which reduced the opportunity for infection to progress to the carrot crown. Trimming combined with application of foliar fungicide was even more effective. Trimming reduced carrot leaf blights (Alternaria dauci, Cercospora carotae) in 1 of 3 years, when disease pressure was low. However, there was no advantage of combining trimming and fungicide sprays for leaf blight control. Canopy modification also reduces disease in legume crops. Soybean cultivars with reduced height and lodging, and early maturity, had up to a 74 % reduction in apothecia of S. sclerotiorum within the crop, and up to an 88 % reduction in disease incidence at harvest. In field pea, artificially supporting plants to reduce lodging, in combination with fungicide application, reduced the severity of mycosphaerella blight (Mycosphaerella pinodes) on pods by 67 % and increased seed yield by 54 %. In chickpea, paired-row planting that opened the canopy increased seed yield by 12 %, likely by increasing fungicide deposition. Modifications of the crop canopy can reduce disease, the need for fungicide sprays, and sometimes improve fungicide efficacy, but the results are often pathosystem-specific.     

柑橘内生细菌YS45的鉴定、抗菌物质分析及其对油菜菌核病的防治作用

 从柑橘枝条中分离到26株对油菜菌核病菌(Sclerotinia sclerotiorum)具有拮抗作用的内生细菌,其中YS45菌株的拮抗活性最强。16S rRNA基因序列分析及形态学和生理生化鉴定结果表明YS45菌株为枯草芽孢杆菌(Bacillus subtilis)。高效液相色谱及质谱分析结果显示其主要的抑菌活性物质为一组fengycin同系物,包括fengycins A、fengycins B和一种稀少fengycin类型化合物。油菜离体叶片接种试验中,YS45菌株发酵液对油菜菌核病的防效在70%以上,与五氯硝基苯相当;田间小区接种试验表明,YS45菌株发酵液对油菜菌核病的防效也在50%以上。