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.     

桑椹肥大性菌核病病原菌生物学特性及流行性

【目的】对桑椹灾害性真菌病害——桑椹肥大性菌核病病原菌,即桑实杯盘菌(Ciboria shiraiana)的生物学特性进行研究,分析其流行性。【方法】采用人工接种、调查等研究方法,对C.shiraiana在无性生长阶段中菌丝侵染能力,菌核的休眠期,有性生长阶段中子囊孢子的结构、释放、数目以及萌发等进行研究,并对菌核萌发的物候期进行调查。【结果】C.shiraiana菌丝对桑雌花没有侵染能力;C.shiraiana菌核具有休眠期,低温处理6周以上的菌核才能萌发形成子囊盘;1个菌核可萌发1–15个子囊盘,直径为1.5 cm的子囊盘能产生高达(5.6–6.3)×10~7个子囊孢子;C.shiraiana子囊孢子在酸性环境中的萌发率明显高于在中性和碱性环境中的萌发率;C.shiraiana菌核萌发形成子囊盘产生子囊孢子的物候期,从1月下旬开始到4月中旬结束,其中在3月中旬萌发子囊盘的数目达到最高值。【结论】桑椹肥大性菌核病属于典型的流行性侵染病,在果桑栽培上容易造成毁灭性危害,生产上必须高度重视该病的防控。     

Prolific production of sclerotia in soil by Rhizoctonia solani anastomosis group (AG) 11 pathogenic on lupin

Rhizoctonia solani anastomosis group (AG) 11 causes serious damping‐off and hypocotyl rot of narrow‐leafed lupins (Lupinus angustifolius) in the northern grain‐belt of Western Australia. R. solani AG‐11 produced abundant sclerotia in sand overlaid on potato dextrose agar. Sclerotia were produced in larger numbers in natural Lancelin sand than in Geraldton loamy sand collected from the northern grain‐belt of Western Australia. The majority of the sclerotia produced were in >250 to <500 μam size range. The germination levels of sclerotia in the first two cycles of drying and germination were not significantly different. Sclerotia still retained 50% germination after four such cycles, indicating that they may have the ability to withstand the climatic cycles of the Mediterranean environment of southwestern Western Australia. The radial growth of the mycelium from sclerotia, however, declined with each drying and germination cycle. Inoculum potential of the pathogen increased with the size of sclerotia resulting in more severe lupin hypocotyl rot with larger sclerotia. The number of sclerotia produced in soil increased with increasing density of lupin seedlings. The results also indicate that R. solani AG‐11 can produce sclerotia on infected plant tissues as well as in soil. This is the first report of the prolific production of sclerotia by AG‐11 and their significant role in infection of lupins in soil in Western Australia.     

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.     

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.     

Germination of sclerotia of Botrytis tulipae, the cause of tulip fire

Sclerotia of Botrytis tulipae (Lib) Lind. buried in field soil germinated mainly in winter and early spring to produce conidiophores and conidia and then decayed. Except when buried very late in the year sclerotia germinated and decayed in the season following burial. In the laboratory, sclerotia from pure culture germinated in two ways. On sterile water-agar they formed colourless mycelium but no conidiophores; the rate of germination was greatest at 25 d?C. On unsterile filter paper and on soil they produced mainly conidiophores and conidia; the rate of germination was greatest at 5 d?C and was enhanced by previous cold treatment. It was unaffected by soil-moisture or pH. Primary infections of tulips arose from sclerotia placed in the soil but only when they were close to the shoot tip or developing stem. It is suggested that, because of their limited survival, soil-borne sclerotia are of importance only when tulips are planted in annual succession.     

Hydrocarbons,free fatty acids,and amino acids of sclerotia of Sclerotinia sclerotiorum

Summary Sclerotia of Sclerotinia sclerotiorum (Lib.) D By. were obtained from commercial pea-and bean-cleaning operations or grown on potato-dextrose agar and synthetic glucose-and sucrose-salts agar media. The crude fat (ether extract) content of sclerotia varied from 0.8 to 1.5%. Extraction and fractionation of the lipids followed by gas chromatographic analysis showed that sclerotia from pea cleanings contained one predominant hydrocarbon which was absent from sclerotia produced in the laboratory. Sclerotia from natural sources and grown in the laboratory contained a similar distribution of C₁₈ unsaturated free fatty acids, however, quantitative differences were noted. Palmitic, oleic and linoleic were the major free fatty acids of the laboratory-grown sclerotia while a high proportion of linoleic acid was also found in sclerotia from natural sources. Sclerotia were fractionated into water-soluble and water-insoluble fractions. After acid hydrolysis of the waterinsoluble fraction, both fractions were analyzed for amino acids. Twenty-one compounds, including 2 unknowns, were detected in the soluble fraction. The hydrolyzates contained 19 amino acids, including the same 2 unknowns. Two compounds tentatively identified as ornithine and -aminobutyric acid were found only in the water-soluble fraction. The relative amino acid composition of the water-insoluble fraction of sclerotia from various sources was fairly constant but the arginine content decreased on the synthetic media.     

The respiration of sclerotia of Sclerotium rolfsii

Summary The respiration of sclerotia ofS. rolfsii was investigated using the Warburg constant-volume respirometer to measure oxygen uptake. The effects of age of sclerotia, pH, and temperature were studied. Sclerotia produced on prune agar were ideal for respirometric studies, being uniformly round and of approximately equal size. On a dry weight basis, the respiration rates of sclerotia were considerably less than those of vegetative mycelium. Sclerotia showed a decrease in respiration with increasing age. This was accompanied by morphological changes in the outer hyphal rind of the sclerotium during maturation. The respiration rate of sclerotia was approximately the same at 30° and 40° C, but was significantly lower at 45° C. Respiration of sclerotia was not markedly affected by normally encountered hydrogen-ion concentrations. However, a pH of 8.0 markedly repressed oxygen uptake. Sclerotia produced in rye grain cultures were chemically analyzed. The nitrogen content was 4.7 %, the petroleum-ether-soluble lipid content was 0.7 %, and the crude glycogen content was 14.2 % of the oven dry weight of the sclerotia.Contribution No. 345 from The Department of Botany. Portion of a thesis presented by the senior author in partial fullfillment for the M.S. degree.     

Induction of edible sclerotia of Pleurotus tuber-regium (FR) Sing, in the laboratory

In the field, sclerotia of Pleurotus tuber-regium were found in dead trees and fallen logs of Daniella oliveri. Sclerotia of P. tuber-regium were produced in the laboratory when the fungus was cultured in moist drill dusts of D oliveri and Elaeis guineensis. The resulting sclerotia were viable and had features similar to those found in the natural habitat.     

Fungal species residing in the sclerotia of <Emphasis Type="Italic">Polyporus umbellatus</Emphasis>

Sclerotia of Polyporus umbellatus is a traditional Chinese herb. The sclerotia can survive in soils for long time and their growth depends upon a symbiotic association with Armillariella mellea. But it is unclear whether other fungi reside or play a role in the sclerotia. In this study, wild sclerotial samples were collected from seven provinces, which span southwest to northeast China. A total of 148 fungal isolates were recovered from the sclerotia of P. umbellatus and classified into 19 morphological taxa. Seventeen belonged to five genera: Fusarium, Eurotium, Penicillium, Geomyces and Mucor. The fungi found within the sclerotia varied depending on the province from which they were collected. The possible role of these fungi is discussed.     

Mycoparasitism by Coniothyrium minitans on Sclerotinia sclerotiorum and its Effect on Sclerotial Germination

Scanning electron microscopy showed that hyphae of Coniothyrium minitans produced appressorium-like swellings when they came in contact with Sclerotinia sclerotiorum in dual culture on PDA. The parasitized hyphae gradually skrank and collapsed, and hyphae of the mycoparasite were found inside the host hyphae. The mycoparasite hyphae grew inter- and intracellularly within the sclerotia of S. sclerotiorum. In the later stages of parasitism, hyphae of the mycoparasite proliferated extensively within the sclerotia and formed pycnidia near the sclerotial surface. At this stage, the sclerotia became flattened, soft and disintegrated. Sclerotia parasitized by C. minitans failed to germinate either myceliogenically or carpogenically.     

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.     

The effect of temperature on the infection of onions by Sclerotium cepivorum

Studies on Sclerotium cepivorum infection of salad onions in artificially infested soil showed that under field conditions infection was greatest in late spring and summer and declined to a low level in the late autumn and winter months. Change of infection levels was found to be correlated with the effect of soil temperature in the field on the germination of sclerotia. This was substantiated in laboratory studies which demonstrated that sclerotium germination, mycelial growth and seedling infection were all markedly influenced by temperature.     

Factors affecting the survival of sclerotia of Sclerotium cepivorum in the Fraser Valley of British Columbia

Sclerotia of Sclerotium cepivorum buried in muck soil in the Fraser Valley decayed with time. The rate of decay of sclerotia was influenced by local environmental conditions. A mixture of soil with sclerotia increased their survival but there was no difference in the rates of decay in three different soils. The decay was greatest during winter when Fraser Valley fields are often flooded. Sclerotial decay was also affected by pretreatment of the sclerotia. Dried sclerotia decayed significantly (P < 0.05) faster than sclerotia which had not been dried, a phenomenon which is apparently due to changes in micro-organisms on the sclerotia. Dried sclerotia which had been incubated in moist soil had fewer bacteria and more fungi than sclerotia which had been incubated in soil without being dried. The increase in fungi on the dried sclerotia was due to a dramatic increase in Trichoderma spp.     

<Emphasis Type="Italic">Coprinopsis cinerea</Emphasis> from rice husks forming sclerotia in agar culture

Sclerotia were formed in agar culture by a fungus with clamp connections isolated from rice husks at Tsukuba, Japan. The sclerotia were brown, globose to ellipsoidal, small, up to 200 μm in diameter, and composed of external rind tissue and internal medulla tissue. Such tiny sclerotia have not been commonly reported among basidiomycetous fungi in the literature. The fungus was identified as Coprinopsis cinerea on the basis of morphological characteristics together with molecular analyses. Three reference strains of C. cinerea formed sclerotia similarly under identical cultural conditions.     

Comparison of the effect of antibiotic substances on sclerotia of Mycosphaerella ligulicola and Colletotrichum coccodes

Summary Sclerotia ofColletotrichum coccodes tolerated much higher concentrations of actidione in agar than did sclerotia ofMycosphaerella ligulicola. With increase in concentration of the antibiotic sclerotia of both species took longer to germinate. Increased resistance of both species to actidione developed after growth of a single generation on media containing the antibiotic. Sclerotia ofC. coccodes survived 5 days immersion in a bacterial culture filtrate whereas scleroia ofM. ligulicola ceased to be viable after a similar period.Sclerotia ofC. coccodes andM. ligulicola exhibited strand and loose types of formation respectively. The degree of resistance of these sclerotia to antibiotic substances was correlated with both longevity in soil and type of formation, but, in general, there is unlikely to be a relationship between structure of the sclerotium and longevity.     

Factors Affecting Biological Control of Sclerotinia sclerotiorum by Fungal Antagonists

Studies were conducted to determine the effects of soil moisture (9, 16 or 24% w/w) and temperature (5, 15, 20 or 25°C) on the control of sclerotia of Sclerotinia sclerotiorum by five fungal agents in sterile and natural field soil. All five biocontrol agents were effective in reducing the survival of sclerotia of S. sclerotiorum in sterile soil under dry (9% moisture) or wet (24% moisture) conditions at 20°C, but only Coniothyrium minitans was effective in natural soil. Coniothyrium minitans was the most effective in reducing sclerotial viability at the temperature range of 15–25°C. Trichoderma virens was effective against sclerotia of S. sclerotiorum to a lesser extent than C. minitans , and in non-autoclaved soil, it performed best at 25°C. Although Epicoccum purpurascens , Talaromyces flavus and Trichothecium roseum were effective against sclerotia of S. sclerotiorum in some instances, they were less effective than C. minitans and T. virens . Sclerotia of S. sclerotiorum conditioned for myceliogenic germination were more vulnerable to attack by the biocontrol agents than dormant sclerotia. The implications are discussed with respect to enhancement of biological control of crop diseases caused by S. sclerotiorum in different geographic regions.     

Origin of Early Attacks of Sclerotinia Stem Rot on Winter Oilseed Rape (Brassica napus sub. sub. sp. oleifera var. biensis) in the UK

Studies in artificially infested field plots of winter oilseed rape showed that mycelia originating from both sclerotia and ascospores of Sclerotinia sclerotiorum can initiate early attacks of sclerotinia stem rot. Disease symptoms appeared in early November in plots previously infested with sclerotia in September. There was a 6–7 week period from artificially inoculating plants with ascospores in October to observing disease symtoms in December. The primary sites of infection for both typesof inocula were the laminae and petioles of senescent and frost-damaged leaves. Following primary infection, mycelia of the pathogen spread by plant to plant contact, causing secondary infections of neighbouring plants. Sclerotia developed on diseased plants and high numbers were returned to the soil. The importance of these findings to the epidemiology of the disease are discussed.     

THE EFFECT OF SOME PHYSIOLOGICAL AND ENVIRONMENTAL FACTORS ON SCLEROTIAL ASPERGILLI

Rudolph , Emanuel D. (Ohio State U., Columbus.) The effect of some physiological and environmental factors on sclerotial Aspergilli. Amer. Jour. Bot. 49(1): 71–78. Illus. 1962.—The effect of varying conditions of carbon-nitrogen balance, temperature, pH, and light upon the formation of sclerotia by 6 species of Aspergillus (A. alliaceus, A. avenaceus, A. flavus, A. quercinus, A. sclerotiorum and A. wentii) was studied. On Czapek's agar, optimal growth as well as maximum production of sclerotia and conidia took place at high sucrose and nitrate concentrations. In general, fewer sclerotia were formed with glucose than with sucrose, and very poor growth took place with lactose. Sclerotia were formed best at temperatures that were optimal or below optimal for mycelial growth. The ranges of pH through which sclerotia were formed were narrower than those through which conidia and mycelia were formed. Light had no effect upon sclerotium formation. The formation of sclerotia in A. alliaceus was found to represent the strand-type development. A number of UV-induced strains and a spontaneous mutant strain of A. alliaceus showing varying amounts of sclerotium and conidium production are characterized. It is suggested that the sclerotia in Aspergillus are sterile stromata.