Grouping of isolates in AG 2 ofRhizoctonia solani by total cellular fatty acid analysis

Total-cellular fatty acid compositions of 34 isolates ofRhizoctonia solani belonging to intraspecific groups (ISGs) of anastomosis group (AG) 2, i.e., AG 2-1, AG 2-2 IIIB (mat rush), AG 2-2 IV (sugar beet), AG 2-2 LP (turfgrass), and AG 2–3 (soybean), were compared. The major fatty acids identified were palmitic, stearic, and oleic acids. Principal component analysis based on the percentage composition of total cellular fatty acids revealed consistently low variability among isolates of a single ISG of AG 2. Average linkage cluster analysis showed that isolates obtained from turfgrass representing a newly proposed group, AG 2-2 LP, were differentiated from other AG 2 ISGs. Isolates of another newly proposed group AG 2–3, from diseased soybean were also closely related to AG 2-1 and AG 2-2 IIIB but distinguishable from the AG 2-1 and AG 2-2 LP isolates by the average linkage cluster analysis. These results suggested that the percentage composition of total-cellular fatty acids is a distinct characteristic for the five ISGs belonging to AG 2, and fatty acid analysis is useful for the differentiation and characterization of these ISGs of AG 2 inR. solani.     

Influence of soil factors,fertilizers and manures on pathogenicity ofRhizoctonia solani onVigna species

Rhizoctonia solani caused maximum mortality of mung bean seedlings at 20°C, and the disease incidence decreased with increase of temperature; 30° was optimum for mycelial growth of the fungusin vitro. The fungus grew best in nutrient broth of pH 5.5 but infected mung bean and pea seedlings more severely in neutral and alkaline river sand than in the sand adjusted to acidic reaction. The disease incidence was higher in adequately moist sandy loam and less in soil under moisture stress. Incidence of cowpea seedling rot was higher in heavy-textured loam and silt loam soils than in light-textured sandy- and loamy sand. Addition of montmorillonite and kaolinite in the sandy soil increased the disease incidence, but these clays reduced fungus growth in culture. More seedling rot occurred in the sandy soil fertilized with urea, potassium nitrate, monocalcium phosphate, or potassium dihydrogen phosphate while soil application of ammonium nitrate, potassium chloride, or potassium sulphate decreased the disease. In tests with combined soil application of N (as urea), P (as monocalcium phosphate) and K (as potassium chloride), disease incidence was more in all combinations having P. Among the six micronutrients tested, only boron reduced the disease incidence significantly both in presence and absence of NPK fertilizers. Farm-yard manure and biogas sludge aggravated seedling rot but their water extracts decreased it. Humic acid, extracted from farm-yard manure, increased the disease incidence but was inhibitory to fungus growth in culture. Green manure also resulted in more disease.     

Variability among Alternaria solani isolates associated with early blight of tomato

Variability among isolates of Alternaria solani, the causal agent of early blight of tomato, from Northern and Southern parts of India was determined based on conidial morphology, pathogenicity tests and random amplified polymorphic DNA (RAPD) techniques. The isolates varied with respect to size of conidia and number of septa. The average size of conidia varied from 150-224.9 microm x 12.4-17.2 microm. The number of horizontal (4-14), vertical (0-3) and beak (0-8) septa also varied among the isolates. The test isolates differed in the virulence pattern on ten tomato genotypes under screen house conditions. Based on disease severity, test isolates were categorized into three main groups. Isolates RAS (Rohtak) and HAS-I (Hisar) were more virulent than all other isolates. None of the genotypes were completely resistant to all the test isolates. The analysis of RAPD profiles showed that there was a high level of genetic variability among the isolates of A. solani. The cluster analysis based on similarity coefficients separated the ten A. solani isolates into two major clusters. There was no evidence for geographical clustering of isolates with high levels of genetic similarity, suggesting that isolates are widely spread across India.     

Build-up and decline ofRhizoctonia solani inoculum under field conditions

Summary Inoculum potential ofRhizoctonia solani Kühn was studied in an infested carnation field during two successive growth seasons. This inoculum potential was expressed as diseased carnation plants in the field and diseased bean seedlings planted in soil samples. Disease incidence in the field soil samples increased during the first season, up to 60% and 100%, respectively. Removing the carnation plants and keeping the soil wet for 45 days, resulted in a sharp decline in inoculum potential. Both inoculum potential and disease incidence in carnations were lower after plant removal. The use of either methyl bromide or vapam resulted in complete control of the disease and reduced inoculum potential. Results suggest possible reduction ofR. solani inoculum by maintaining the soil moist between growth periods.     

Heterokaryon formation with homokaryons derived from protoplasts ofRhizoctonia solani anastomosis group eight

Homokaryons were successfully recovered by regenerating protoplasts prepared from vegetative hyphae of field isolates ofRhizoctonia solani anastomosis group (AG) 8, the causal pathogen of bare-patch disease of cereals. A mating type incompatibility system, which is similar to that reported in AG 1 and AG 4, was demonstrated in AG 8. All homokaryons obtained in AG 8 were able to form tufts with their parent isolates and other heterokaryotic field isolates of AG 8 tested. Heterokaryons were readily recovered from tufts of pairing of certain homokaryon combinations. The synthesized heterokaryons formed tufts with both of the contributing homokaryons. The majority of hyphal tip cultures isolated from tufts resembled one of the contributing homokaryons. These nonheterokaryotic hyphae in tufts are attributed to transient heterokaryon effects.     

Morphological and pathological variations of rice sheath blight inciting south Indian Rhizoctonia solani isolates

Frequent assessment of pathogen diversity is one of the most important criteria in designing disease management programmes. A study on diversity of field isolates of Rhizoctonia solani from sheath blight-infected rice fields of south India has been carried out. A total of 236 R. solani isolates were obtained from 45 locations in the surveyed area. Sclerotial features such as colour, size and shape and distribution pattern were varying among isolates. However, no other morphological features found to differ among isolates. Majority of the R. solani isolates were fast growers as they attained complete mycelial growth within 2 days in a 90-mm Petri plate and the emergence of sclerotial structures was seen even in 4 days of incubation. Selected 10 R. solani isolates exhibited considerable variations in pathogenicity on three different rice cultivars. Vellai ponni was found to be the most susceptible rice cultivar to all the field isolates of R. solani.     

Bacterization of peanut withPseudomonas fluorescens for biological control ofRhizoctonia solani and for enhanced yield

Severity of stem-rot disease of peanut caused byRhizoctonia solani was reduced by 54.9 and 68% in plants of two cultivars treated in the greenhouse with antagonistic strains ofPseudomonas fluorescens. These strains were selected based on theirin vitro toxicity to mycelial growth and sclerotial germination ofR. solani. In field experiments, bacterization of peanuts withP. fluorescens resulted in taller plants (by 25.7%) and increased yields (by 59.0%).     

Conventional farming impairs Rhizoctonia solani disease suppression by disrupting soil food web

Intensive farming in agriculture raises questions in relation to environmental sustainability and the widespread use of agrochemicals. In the present work, we compare the impact of organic and intensive farming, in connection to the soil suppressiveness against the soilborne pathogen Rhizoctonia solani. Three farms were considered in the study: two practicing organic cultivation (for 10 and 20 years, respectively), and one applying conventional cultivation. Soil suppressiveness was assessed in a greenhouse bioassay with lettuce (Lactuca sativa). Soil microbiome was characterized by combining BIOLOG EcoPlates^? with high‐throughput sequencing of bacterial and eukaryotic rRNA gene markers. Suppressiveness towards R. solani was higher in organic than in conventional farming soil, but this property was lost after soil sterilization. Functional biodiversity was significantly higher in the two organic soils, and this parameter was predictive of the suppressiveness towards R. solani. According to our analyses, the overall microbial taxonomic diversity was unlinked to suppressiveness. A correlation analysis, carried out at the genus level for the most abundant bacterial and eukaryotic microbial taxa, showed that 58.7% of the genera had a statistically significant correlation with suppressiveness. In particular, the genera Flavisolibacter, Massilia, Pseudomonas, Ramlibacter, Rhizophus and the oligochaete worms belonging to the Enchytraeidae family positively correlated with the disease suppression.     

Effect of peat-bran inoculum ofTrichoderma species on biological control ofRhizoctonia solani in lettuce

A range of known biocontrol or plant growth-stimulating species ofTrichoderma orGliocladium were grown on peat-bran substrate to yield between 5×10⁷–3×10¹⁰ colony forming units (cfu's)g^–1 substrate after 14 days growth. Inocula were incorporated into peat:sand potting compost infested withRhizoctonia solani to give 7–8 × 10⁴ cfu's of antagonist g^–1 compost and assessed for biological control activity using lettuce seedlings. Six of the eight antagonists decreased daming-off and three of these consistently increased yield in comparison withR. solani treatment alone.Subsequently, peat-bran inoculum ofT. harzianum isolate TH1 was incorporated at 0.5% w/v intoR. solani infested potting compost. Both autoclaved and nonautoclaved inoculum ofT. harzianum TH1 decreased disease and increased yield. Incorporation of ethyl acetate-extracted autoclaved inoculum ofT. harzianum TH1 resulted in similar levels of biocontrol and improved plant growth as did incorporation of nonautoclaved and autoclavedT. harzianum TH1 inoculum. The need to standardize inocula and controls is emphasized.     

Studies on groundnut hypocotyl exudates and the behaviour ofRhizoctonia solani in influencing the disease

Summary The behaviour ofRhizoctonia solani on the surface of groundnut seedling hypocotyls was studied at different stages of growth. Characteristic variation in the growth and production on infection cushions was observed. No infection cushions were observed as the seedling reaches the age of three weeks. Hypocotyl exudates were collected at various stages of growth and analysed for total phenols, carbohydrates and ninhydrin positive compounds. The exudates were also analysed for individual amino acids, sugars and organic acids. Gradual decline in the total quantities of various compounds was observed with age. Quantitative and qualitative change in the individual compounds was also evident and some of the compounds were not at all detected as age increases. The possible correlation between the exudation and behaviour of the fungus on host surface is discussed.     

Stem canker (Rhizoctonia solani) of maincrop potatoes.

In two years, potato plants were sampled at 1- or 2- weekly intervals from plots planted with seed tubers bearing sclerotia of Rhizoctonia solani (black scurf) and with seed without sclerotia either infested or not with cultures of R. solani at planting. Sprouted King Edward seed was used in 1981 and sprouted and non-sprouted King Edward and Pentland Crown seed in 1982. In both years 60–80% of shoots from seed with sclerotia and 90% of shoots from seed inoculated at planting were affected with stem canker. Most disease developed before shoots emerged although it gradually increased later when new shoots arising both from seed tubers or as branches on shoots with damaged apices (pruned shoots) became infected before they emerged. Sprouting seed tubers bearing sclerotia decreased the disease on both cultivars but with soil-applied inoculum the disease was more severe on plants from sprouted than non-sprouted seed. Some stolons were infected by R. solani soon after they developed and incidence of infection later increased. Thirty to 50% of stolons were infected on plants from infected seed tubers and 60% on plants with soil-applied inoculum. With both cultivars and sources of inoculum about 70% of the infected stolons had their apices killed (pruned).     

Comparison between Rosellinia necatrix isolates from soil and diseased roots in terms of hypovirulence

The white root rot fungus, Rosellinia necatrix, is a devastating soil-borne pathogen of many plant species. Biocontrol with the hypovirulence factor is promising, but disease symptoms, signs or culture morphology of the pathogen cannot be reliably used as markers for hypovirulence in this fungus. We attempted to obtain hypovirulent isolates from soil rather than from diseased roots, based on the hypothesis that hypovirulent isolates were more likely to persist in soil as saprobes. Sixteen isolates, belonging to eight mycelial compatibility groups (MCGs), were obtained from soil in two active and one abandoned Japanese pear orchards. Comparison of these isolates based on clonality revealed that six MCGs were commonly recovered from both diseased roots and soil and two MCGs exclusively from soil. No MCG was found in more than one orchard. With two exceptions, isolates within the same MCG were similar in virulence, competitive saprophytic ability (CSA) and mycelial growth rate whether or not they carried dsRNA. The two exceptional isolates recovered from soil had multiple dsRNA segments that caused hypovirulence, weakened CSA and restricted mycelial growth on nutrient-rich media. They belonged to different MCGs, each including dsRNA-free isolates. Isolates from soil contained various dsRNAs (44%), including the hypovirulence factor, more frequently than isolates from diseased roots in the same fields (25%), which is much higher than the proportion of isolates with dsRNA from diseased roots (19%) in a total of 424 isolates from Japan examined so far. These results suggest that isolation of R. necatrix from soil is an effective method to obtain isolates with dsRNAs, including the hypovirulence factor.     

Reduced sensitivity of tomato early blight pathogen (Alternaria solani) isolates to protectant fungicides,and implication on disease control

The sensitivity of Alternaria solani isolates to the fungicides mancozeb and chlorothalonil was evaluated, to determine if inadequate disease management by these fungicides could be attributed to reduced sensitivity of A. solani isolates to these fungicides. The sensitivity of 60 isolates of A. solani was assessed using the inhibition of radial mycelial growth (RG) method, using fungicide concentrations of 0, 1.0, 10, 100, 500 and 1000 μg a.i ml^?1 medium. EC₅₀ was calculated for each isolate and fungicide combination. The EC₅₀ values of different A. solani isolates to mancozeb ranged from 9.05 to 712.65 μg ml^?1. EC₅₀ values of different isolates to chlorothalonil ranged from 4.25 to 849.4 μg ml^?1. The percentage of isolates with reduced sensitivity was 46.7 and 53.3% for mancozeb and chlorothalonil, respectively. Results of the in vivo tests demonstrated decline in disease control by the two fungicides with the reduced-sensitivity isolates compared to the sensitive ones.     

Relationship among isolates of rhizoctonia solani Kuhn

Aversion was observed between isolates ofRhizoctonia solani on inoculation in a culture plate. This phenomenon helped to study the relationship among fifty different isolates. Thirty three of the isolates showed aversion against some isolates and non-aversion against others. Seventeen of them, however, showed aversion against all the isolates. A digrammatic scheme is presented indicating the factors controlling this relationship.     

Quantification of rice sheath blight progression caused by Rhizoctonia solani

Rhizoctonia solani has a wide host range, including almost all cultivated crops and its subgroup anastomosis group (AG)-1 IA causes sheath blight in rice. An accurate measurement of pathogen’s biomass is a convincing tool for enumeration of this disease. Mycological characteristics and molecular diagnosis simultaneously supported that all six strains in this study were R. solani AG-1 IA. Heterokaryons between strains Rs40104, Rs40105, and Rs45811 were stable and viable, whereas Rs40103 and Rs40106 did not form viable fused cells, except for the combination of Rs40106 and Rs40104. A primer pair was highly specific to RsAROM gene of R. solani strains and the amplified fragment exists as double copies within fungal genome. The relationship between crossing point (CP) values and the amount of fungal DNA was reliable (R ² >0.99). Based on these results, we determined R. solani’s proliferation within infected stems through real time PCR using a primer pair and a Taqman probe specific to the RsAROM gene. The amount of fungal DNA within the 250 ng of tissue DNA from rice cv. Dongjin infected with Rs40104, Rs40105, and Rs45811 were 7.436, 5.830, and 5.085 ng, respectively. In contrast, the fungal DNAs within the stems inoculated with Rs40103 and Rs40106 were 0.091 and 0.842 ng. The sheath blight symptom progression approximately coincided with the amount of fungal DNA within the symptoms. In summary, our quantitative evaluation method provided reliable and objective results reflecting the amount of fungal biomass within the infected tissues and would be useful for evaluation of resistance germplasm or fungicides and estimation of inoculum potential.     

A selective medium for isolating Rhizoctonia solani from soil

Ko & Hora's (1971) selective medium (FM) for counting Rhizoctonia solani propagules from soil has been modified. FM medium minus gallic acid and fenaminosulf i.e. mineral antibiotic (MA) medium, was amended with inhibitors and fungicides at different concentrations. The modified medium consisting of MA + gallic acid (400 μg ml^-1) + fosetyl-Al (250μgml^-1) was found most efficient in selective isolation of R. solani from soil even in the presence of Macrophomina phaseolina (Rhizoctonia bataticold) and other soil mycoflora. This amended medium was effective in isolating R. solani from soil even when the number of propagules of M. phaseolina present was 10 times greater. The sensitivity of this medium was five times better than Ko & Hora's medium.     

Study of the aggressiveness of Rhizoctonia solani isolates

This paper presents an in vitro test to screen the pathogenicity of different Rhizoctonia solani isolates on a host range. The level of aggressivity of the different isolates was different for several host plants tested. There were significant differences between the crops and the isolates tested. In general, the disease level was higher on beans, lettuce and cabbage. In carrot and rye grass the level of infection was lower for the isolates of R. solani tested. The potato isolates of R. solani were less aggressive than the isolates coming from maize, fodder beet and sugar beet. The R. solani isolates were also biochemically characterized by pectic zymograms: the isolates Rs0401 (from maize) and Rs0504 (from sugar beet) belong both to the anastomosis group AG2-2.     

Impact of two soil-applied herbicides on damping-off of cowpea caused byRhizoctonia solani

Summary Alachlor was more inhibitory toRhizoctonia solani growth than fluchloralin in potato dextrose broth (PDB). Infective capacity of the pathogen was not altered by growing it in a medium containing either of the herbicides. Cowpea seedlings grown in alachlor-treated soil were more susceptible toR. solani than those treated with fluchloralin and the untreated seedlings. Pre-sowing application of alachlor in soil (5 l a.i./kg) aggravated damping-off whereas fluchloralin decreased the disease incidence to nearly half of that in untreated soil in greenhouse pot tests (av. temperature 31±5°C). Both herbicides reduced damping-off in pots kept at constant temperature of 30°C and increased the disease incidence at 20°C. Fungus growth in culture was stimulated at 20° but was strongly inhibited at 30°C by both herbicides. Growth inhibition by herbicides was maximum in PDB of pH 8 and decreased steadily up to pH 5.Impact of fluchloralin and alachlor onR. solani damping-off of cowpea appears to be due to the predisposing effect by the herbicides on the susceptibility of the host and is influenced by atmospheric temperature.