The production by species of Allium of alkyl sulphides and their effect on germination of sclerotia of Sclerotium cepivorum Berk

A number of n -propyl and allyl sulphides were shown to stimulate germination of sclerotia of Sclerotium cepivorum in soil. Several of these sulphides were evolved by chopped garlic cloves and onion bulbs, and by their extracts, as well as by distillates and condensates prepared from extracts. Sulphides produced by garlic were methyl and allyl compounds, whereas those of onion were methyl and n -propyl derivatives. Few or no volatile sulphides were detected by gas chromatography of head spaces of intact Allium plants although it is considered that small quantities of sulphides may be produced from these plants in soil. The stimulatory activity of distillates and condensates appears to be entirely due to their alkyl sulphide content and that of extracts at least partially so.     

Fungi isolated from Sclerotia ofSclerotium cepivorum and from soil and their effects upon the pathogen

Summary Data are presented on the antagonistic effects of the fungi isolated from sclerotia ofSclerotium cepivorum and from nonrhizosphere soil taken from around the roots of infected onions upon mycelial growth and sclerotial germination ofS. cepivorum. Most of the isolated fungi especiallyPenicillium species were antagonistic to mycelial growth. Sclerotial germination was slightly inhibited by diffusates of these fungal isolates. Testing the antifungal effect of someAllium extracts against the fungal isolates by the inhibition zone method showed that garlic extract has the greatest antifungal effects and onion extract is the least potent. However, spore germination tests indicated that onion extract completely inhibits the spore germination of all test fungi. The role of host-plant extracts in stimulating sclerotial germination is discussed.     

STUDIES OF THE BIOLOGY OF SCLEROTIUM CEPIVORUM BERK.

Sclerotia from 6-week-old pure cultures of Sclerotium cepivorum germinated immediately in soil only after abrasion of their rinds, but after burial in soil for a month or more, unabraded sclerotia became capable of germination.
Marked stimulation of germination occurred in the presence of host plants (onion, leek and shallot). Little or no germination occurred in soil alone or in the presence of non-host plants (barley, cabbage, carrot and white clover). Sclerotial germination was observed in a number of soils of widely differing pH and over a wide range of soil water content. Germination of sclerotia on uninjured onion roots was greatest at the tip region. On artificially injured roots sclerotial germination was enhanced but the effect of position was lost.
Sclerotial germination was independent of contact between roots and sclerotia. It was induced by root extracts of all Allium spp. tested, but of no other plants. Boiling or autoclaving root extracts did not destroy the active principle and it is concluded that under field conditions sclerotia are induced to germinate by a thermostable chemical substance from Allium roots.
The process of germination of sclerotia is described.     

Induced tolerance of Sclerotium cepivorum to antibiotics in the presence of onion exudates

The onion white rot pathogen Sclerotium cepivorum was cultured on agar media containing 2% malt extract and one of the antifungal antibiotics, endomycin, griseofulvin, venturicidin and cycloheximide at concentrations that reduced but did not prevent growth of mycelium. When onion seeds or agar discs impregnated with diffusates from onion bulbs were placed on the antibiotic media, radial growth of the fungus was greatly increased, and there was a profuse development of aerial mycelium. Gaseous diffusates from onion tissue and from impregnated agar discs were also effective. On the antibiotic media, tomato, cabbage and radish seeds did not stimulate the growth of S. cepivorum and the onion exudates did not stimulate the growth of four other fungi. This and other evidence is considered to show that the stimulation of growth of S. cepivorum was not caused by any direct effect on the antibiotics but by a tolerance of the fungus to them, which was specifically induced by an exudate from its host plant. The phenomenon may be related to the reported reversal by onion extracts of the inhibitory effects of soil mycostasis on germination of sclerotia of the fungus.     

Production of volatile alkyl sulphides by microbial degradation of synthetic alliin and alliin-like compounds, in relation to germination of sclerotia of Sclerotium cepivorum Berk

Onion distillands proved more stimulatory than distillates towards germination of sclerotia of Sclerotium cepivorum. Synthetic allylcysteine and n-propylcysteine and their sulphoxides stimulated germination of sclerotia when added to soil. Solutions of these compounds in the presence of unsterile soil produced gaseous alkyl mercaptans, sulphides and disulphides which were detected by gas-liquid chromatography and which stimulated sclerotial germination when drawn through soil containing sclerotia. No volatile sulphides were detected above autoclaved solutions but the atmosphere over propyl and allylcysteines was slightly stimulatory although the corresponding autoclaved sulphoxides showed no such activity. The gases above all four solutions sterilized by membrane-filtration were not stimulatory. Six common soil bacteria were shown to be capable of degrading the synthetic precursors, causing the evolution of appropriate volatile alkyl sulphides.     

Botrytis sinoallii: a new species of the grey mould pathogen on Allium crops in China

A study was conducted to identify Botrytis spp. isolated from bulb onion, green onions, garlic, and garlic chives grown in Hubei Province of China. Based on colony morphology and conidial characteristics, 367 strains of Botrytis spp. were identified as five distinct species, namely, B. cinerea, B. squamosa, B. porri, B. byssoidea, and an undescribed Botrytis species (Botrytis sp.), which accounted for 64.3, 29.9, 3.3, 0.3, and 2.2%, respectively. The previously undescribed species is herein described as a new species, B. sinoallii sp. nov., which is characterized by production of numerous small sclerotia on potato dextrose agar. Phylogenetic analysis using partial sequences of three nuclear genes (G3PDH, HSP60, and RPB2) showed that B. sinoallii forms a unique lineage, which is closely related to B. squamosa, a well-known species on Allium crops, but distantly related to the other species of Botrytis on Allium crops, including B. cinerea, B. porri, B. aclada, B. allii, B. byssoidea, B. globosa, and B. sphaerosperma. Results of inoculation tests showed that B. sinoallii is a newly identified agent that can cause leaf blight of green onion, garlic, and garlic chives. Potential impact of B. sinoallii on production of Allium crops in China is discussed.     

The effect of species of Allium on germination in soil of sclerotia of Sclerotium cepivorum Berk

Sixty-three different plants, from a wide range of families, were tested for their effects on germination of sclerotia of Sclerotium cepivorum. Germination was stimulated only by members of the genus Allium. Water extracts of Allium species also stimulated germination of sclerotia, but extracts of other plants, and a number of nutrient solutions, had little effect. There was no evidence of inhibition of sclerotial germination by plants which failed to elicit a germination response.     

GarlicESTdb: an online database and mining tool for garlic EST sequences

Background  

Allium sativum., commonly known as garlic, is a species in the onion genus (Allium), which is a large and diverse one containing over 1,250 species. Its close relatives include chives, onion, leek and shallot. Garlic has been used throughout recorded history for culinary, medicinal use and health benefits. Currently, the interest in garlic is highly increasing due to nutritional and pharmaceutical value including high blood pressure and cholesterol, atherosclerosis and cancer. For all that, there are no comprehensive databases available for Expressed Sequence Tags(EST) of garlic for gene discovery and future efforts of genome annotation. That is why we developed a new garlic database and applications to enable comprehensive analysis of garlic gene expression.     

Variation within isolates ofTyphula incarnata from localities differing in winter climate

Isolates ofTyphula incarnata, a snow mold fungus, were collected from four localities with different winter climates. Their ecological traits such as mycelial growth rate, sclerotium size, carpogenic germination of sclerotia, and aggressiveness were compared between populations in order to reveal infraspecific differentiation associated with climatic differences. Population variability was evident only in sclerotium germination: isolates from more snowy localities germinated faster than those from less snowy localities.T. incarnata is regarded as a versatile pathogen with no specialized forms in contrast withT. ishikariensis. The germination rate of sclerotia is considered very critical in the life history ofT. incarnata.     

A direct observation technique for evaluating sclerotium germination by Macrophomina phaseolina and effects of biocontrol materials on survival of sclerotia in soil

Germination of sclerotia of Macrophomina phaseolina was quantified by direct microscopic observation following application of experimental treatments in vitro and incubation of sclerotia in soil. To assay germination, pieces of agar containing sclerotia were macerated in dilute, liquid cornmeal agar on glass slides; thinly spread; and incubated in a saturated atmosphere for 18–22 h. Germinated sclerotia then were identified by morphological features of germ hyphae. Frequencies of germination were similar in three dilute agar media. Germination was not affected by air-drying sclerotia for 2 weeks, but it was significantly reduced after 4 weeks and greatly reduced or eliminated after 6 or 8 weeks. Survival of sclerotia for 14 days in soil was greatest at 50, 75, and 100% moisture-holding capacity, less at 0 and 25%, and least at 125% (flooded soil). Incorporation of ground poultry litter into soil at 5% by weight reduced survival of sclerotia after 13 days, and incorporation of litter at 10% nearly eliminated it. These results indicate that the direct-observation technique may be used to evaluate animal wastes and other agricultural byproducts for biocontrol activity against sclerotia of M. phaseolina in soil.     

Carpogenic Germination of Sclerotia of Sclerotinia sclerotiorum after Periods of Conditioning in Soil

Periods of conditioning in soil reduced the length of the resting period needed before sclerotia of Sclerotinia sclerotiorum could germinate to form apothecia. Curves for germination of sclerotia were fitted by a form of the log-logistic equation and from this equation the time taken for 50% germination (x₅₀) was calculated. These x₅₀ values were used as the basis for comparing germinability of sclerotia collected from infected sunflower plants and others conditioned in soil, or moist vermiculite for various times. Sclerotia from sunflower roots germinated sooner than those from the stem cavities. Germinability increased with the length of the conditioning period. Conditioning in soil was more effective than in moist vermiculite.     

Colonization of Sclerotinia sclerotiorum sclerotia by a biocontrol isolate of Trichoderma harzianum,and effects on myceliogenic germination

Sclerotia of Sclerotinia sclerotiorum were incubated on cultures of Trichoderma harzianum. Myceliogenic germination decreased by 50% within 1 day and continued to decrease over time. Quantitative PCR showed a decrease in Sclerotinia DNA for older sclerotia, but not fresh sclerotia. Trichoderma DNA increased and persisted inside older sclerotia but not fresh sclerotia.     

The aqueous garlic, onion and leek extracts release nitric oxide from S-nitrosoglutathione and prolong relaxation of aortic rings

Garlic, onion and leek have beneficial effects in treatment of numerous health disorders. The aim of the present study was to investigate underlying molecular mechanisms. To test the potency of the aqueous garlic, onion and leek extracts to release NO from GSNO we have measured NO oxidation product, NO(2)-, by the Griess reagent method. Further, we studied the ability of garlic extract to relax noradrenaline-precontracted rat aortic rings in the presence of GSNO and effects of garlic extract on electrical properties of rat heart intracellular chloride channels. We have observed that: i) garlic, onion and leek extracts released NO from GSNO in the order: garlic > onion > leek; ii) the ability of garlic extract to release NO was pH-dependent (8.0 > 7.4 > 6.0) and potentiated by thiols (Cys > GSH = N-acetyl-cysteine > oxidized glutathione) at concentration 100 μmol/l; iii) the garlic extract (0.045 mg/ml) prolonged relaxation time of aortic rings induced by GSNO (50 nmol/l) and inhibited intracellular chloride channels. We suggest that NO-releasing properties of the garlic, onion and leek extracts and their interaction with Cys and GSH are involved in NO-signalling pathway which contributes to some of its numerous beneficial biological effects.     

The effect of species of Allium on soil bacteria in relation to germination of sclerotia of Sclerotium cepivorum Berk

Extracts made from Allium species were shown to exhibit marked antibiotic properties in a variety of tests. However, no evidence was obtained to suggest that intact Allium seedlings exude antibiotic compounds in sufficient quantity to cause inhibition of bacteria in the soil or in assay tests in the laboratory. Germination of sclerotia of Sclerotium cepivorum was induced by Allium extracts which were too dilute to cause antibiotic responses in laboratory tests using a number of bacteria. Synthetic methyl-methanethiolsulphinate had little effect on germination of sclerotia. The specific reversal, by Allium species and their extracts, of the mycostatic effect of unsterile soil on sclerotia of S. cepivorum does not appear to be due to antibiotic effects.     

Partial synchronization of cell division in root meristem induced by 5-aminouracil

5-aminouracil induces a partial synchronization of mitoses in barley, onion and garlic root tips. The highest degree of synchronization has been achieved in garlic where the mitotic index reached the value of about 36%, while in onion and barley the values equalled about 20%. The concentration causing the maximal synchronization in barley (400–750 ppm) was many times higher than in garlic (62.5 ppm) and onion (100 ppm). The occurrence of micronuclei was evaluated in garlic, under the conditions when synchronization was maximal. It was increased nearly tenfold as compared with the control.     

Biological control of Sclerotinia disease by Aspergillus sp. on oilseed rape in the field

Sclerotinia sclerotiorum causes serious yield losses to many crops worldwide. Aspergillus sp. Asp-4, previously shown to inhibit germination of sclerotia of S. sclerotiorum in vitro and in the field, was evaluated in field trials for suppression of this pathogen on oilseed rape. Spray application of Asp-4 to the soil prior to sowing rice in a rice–oilseed rape rotation resulted in a significant reduction in incidence of Sclerotinia stem rot on oilseed rape compared with the non-treated control in two field trials. This application of Asp-4 also resulted in a significant reduction in germination of sclerotia relative to the non-treated control in these field trials, suggesting that this reduction in sclerotial germination led to disease control. Microscopic examination demonstrated that Asp-4 could effectively colonise external and internal portions of sclerotia of S. sclerotiorum in vitro. Incubation of Asp-4 with sterile sclerotial material induced production of β-glucanase and chitinase activities by this isolate; β-glucanase and chitinase being potentially capable of degrading the glucan and chitin polymeric components of sclerotia. Incubation of Asp-4 with sterile sclerotial material also resulted in a significant reduction in dry weight of this sclerotial material relative to the non-treated control in 96?h in vitro experiments. Experiments reported here indicate that Aspergillus sp. Asp-4 has promise as a biological control agent for S. sclerotiorum on oilseed rape. Experiments reported here suggest that disease control results from inhibition of germination of sclerotial resting structures due to mycoparasitic colonisation by Asp-4.     

Effect of Moisture and Temperature on the Survival of Sclerotia of Sclerotium rolfsii in Soil

The sclerotia of Sclerotium rolfsii Sacc. survived in natural soil for 225 days under controlled moisture at 50% water holding capacity (WHC) after which there was a progressive reduction in the population of viable sclerotia. At 390 days only 48% were recovered. Sclerotia survived well at moisture contents upto 75% WHC but at 100% the population declined rapidly and none were recovered after 60 days. The contents of the sclerotia were found to lyse without germination leaving hollow rinds. Such lysis was found to be favoured between 25 and 40°C. At and below 20°C no such lysis was recovered and more than 80% sclerotia were recovered even after 60 days.     

Stimulation of sclerotial germination ofSclerotium cepivorum by host-plant extract

Summary Extract from onion bulbs and diffusate from roots of onion seedlings were fractionated by column chromatography. The stimulatory effects of the different fractions of onion extract on sclerotial germination ofSclerotium cepivorum were studied. The sugar fraction was the most stimulatory, whereas, the amino acid fraction was not effective. Paper chromatographic analysis revealed the presence of glucose, fructose and no amino acids in the root diffusate. These two sugars and 13 amino acids were identified in the onion extract. When various sugars and amino acids were supplied individually to autoclaved soil, only glucose, fructose, mannose and maltose effectively induced sclerotial germination. Partial stimulation occured in nonsterile soil amended with high glucose concentrations. Studies on the antibiotic effect of the different fractions against some soil fungi by the spore germination method showed that, the sugar fraction inhibits completely the spore germination of all the fungi, tested, whereas, the amino acid fraction was non-inhibitory. Both fractions did not show antibiotic activity when tested by the filter paper disc method. Attempts to extract inhibitory substances from soil which inhibit sclerotial germination were unsuccessful. It was suggested that onion extract plays a twofold role in stimulating sclerotial germination in natural soil: (a) a direct nutritional influence; (b) an antibiotic effect on soil mycoflora which reduces competition for nutrients.     

Onion and garlic extracts lessen cadmium-induced nephrotoxicity in rats

Cadmium (Cd) is a well-known nephrotoxicant inducing kidney damage via oxidative stress. Since kidney is the critical target organ of Cd toxicity, this study was designed to evaluate the protective effects of onion (Allium cepa L.) and garlic (Allium sativum L.) aqueous extracts on Cd-induced renal oxidative stress in male Wistar rats. The control group received double distilled water alone and Cd group was challenged with 3CdSO₄ · 8H₂O (as Cd) (1.5 mg/100 g bw/day per oral) alone. Extract-treated groups were pre-treated with varied doses (0.5 ml and 1.0 ml/100 g bw/day per oral) of onion and/or garlic extract for 1 week after which they were co-treated with Cd (1.5 mg/100 g bw/day per oral) for 3 weeks. The results showed that the levels of renal lipid peroxidation (LPO) and glutathione-S transferase (GST) were significantly (P < 0.001) increased in rats that received Cd alone relative to the control group. More so, the levels of renal glutathione (GSH), superoxide dismutase (SOD), catalase (CAT) and Na⁺/K⁺-ATPase were significantly (P < 0.001) decreased in rats that received Cd alone. Treatment of Cd-intoxicated rats with varied doses of onion and/or garlic extract significantly (P < 0.05) restored the alterations in these parameters relative to the group that received Cd alone. While treatment with high dose of onion extract exerted a significant dose-dependent restoration of these parameters, treatment with high dose of garlic elicited a pro-oxidant effect, relative to their respective low dose. Our study suggests that onion and garlic extracts may exert their protective effects via reduction in LPO and enhanced antioxidant defense. These extracts may, therefore, be useful nutritional option in alleviating Cd-induced renal damage.     

Sclerotia of Botrytis as a Source of Primary Inoculum for Bunch Rot of Grapes in New South Wales, Australia

Sclerotia of Botrytis cinerea were found to be a source of primary inoculum for bunch rot of grapes in the Hunter Valley of New South Wales. Conidia-bearing sclerotia were found on canes on grapevines. Resporulation of sclerotia was observed and sporogenic germination led to infection of flowers and berries. Sclerotia survived for eight months in untreated soil in the laboratory. Application of dicarboximides (procymidone, iprodione and vinclozolin) reduced infection of flowers and bunch rotcaused by the sporogenic germination of sclerotia.