Genetic diversity of Rhizoctonia solani AG-3 from potato and tobacco in North Carolina

Anastomosis group 3 (AG-3) of Rhizoctonia solani (teleomorph = Thanatephorus cucumeris) is frequently associated with diseases of potato (AG-3 PT) and tobacco (AG-3 TB). Although isolates of R. solani AG-3 from these two Solanaceous hosts are somatically related based on anastomosis reaction and taxonomically related based on fatty acid, isozyme and DNA characters, considerable differences are evident in their biology, ecology, and epidemiology. However, genetic diversity among field populations of R. solani AG-3 PT and TB has not been documented. In this study, the genetic diversity of field populations of R. solani AG-3 PT and AG-3 TB in North Carolina was examined using somatic compatibility and amplified fragment length polymorphism (AFLP) criteria. A sample of 32 isolates from potato and 36 isolates from tobacco were paired in all possible combinations on PDA plus activated charcoal and examined for their resulting somatic interactions. Twenty-eight and eight distinct somatic compatibility groups (SCG) were identified in the AG-3 PT and AG-3 TB samples, respectively. AFLP analyses indicated that each of the 32 AG-3 PT isolates had a distinct AFLP phenotype, whereas 28 AFLP phenotypes were found among the 36 isolates of AG-3 TB. None of the AG-3 PT isolates were somatically compatible or shared a common AFLP phenotype with any AG-3 TB isolate. Clones (i.e., cases where two or more isolates were somatically compatible and shared the same AFLP phenotype) were identified only in the AG-3 TB population. Four clones from tobacco represented 22% of the total population. All eight SCG from tobacco were associated with more than one AFLP phenotype. Compatible somatic interactions between AG-3 PT isolates occurred only between certain isolates from the same field (two isolates in each of four different fields), and when this occurred AFLP phenotypes were similar but not identical.     

Control of Fusarium solani rot of potato tubers with fungicides

Tecnazene (up to 33 ppm) and dichloran (up to 500 ppm) had little effect on germination of spores or growth of Fusarium solani isolated from and causing a rot of potato tubers; they also did not decrease rotting when applied to wounds later inoculated with the pathogen. Benomyl and thiabendazole (up to 500 ppm) also had little effect on spore germination but did greatly decrease growth at 5 ppm. A pronounced pink coloration developed in cultures growing slowly in the presence of benomyl; a similar though less striking effect appeared in agar cultures containing thiabendazole. Benomyl suspended in water or diluted with Fuller's earth gave good control of rotting when applied to wounds inoculated later with F. solani. Still better control was obtained with thiabendazole; dusts containing 1% a.i. substantially decreased rots and those containing 10 % a.i. gave almost complete control when applied to wounds shortly before inoculation. Thiabendazole was also very effective when used 24 h after inoculation and a fair measure of control was obtained when it was applied 24 h later. Benomyl and thiabendazole placed on apparently intact surfaces of tubers caused tissue 5 mm deep to become toxic to F. solani 10 days later, and, unexpectedly, this tissue prevented spore germination. Fuller's earth alone substantially decreased rotting. The results obtained suggest that dusts containing thiabendazole have some promise for the control of Fusarium rots of potato tubers, especially of early crops.     

Soil suppressiveness to Rhizoctonia solani and microbial diversity

Rhizoctonia solani anastomosis group 2-2IIIB causes damping-off, black root rot and crown rot in sugar beet (Beta vulgaris). Based on experiences of growers and field experiments, soils can become suppressive to R. solani. The fungus may be present in the soil, but the plant does not show symptoms. Understanding the mechanisms causing soil suppressiveness to R. solani is essential for the development of environmentally friendly control strategies of rhizoctonia root rot in sugar beet. A bioassay that discriminates soils in their level of disease suppressiveness was developed. Results of bioassays were in accordance with field observations. Preliminary results indicate an active role of microbial communities. Our research is focused on the disentanglement of biological mechanisms causing soil suppressiveness to R. solani in sugar beet. Therefore, we are handling a multidisciplinary approach through experimental fields, bioassays, several in vitro techniques and molecular techniques (PCR-DGGE).     

Stem canker (Rhizoctonia solani) on five early and seven maincrop potato cultivars I. Infection of shoots, stolons and tubers

In 1983 and 1984, potato seed tubers of five early and seven maincrop cultivars were inoculated with cultures of Rhizoctonia soluni during planting to simulate severe seed infection. Shoot and stolon infection was assessed in June-August and black scurf on tubers recorded after harvest in October. Almost all shoots of all cultivars had stem canker in both years and disease on shoots, stolons and tubers was more severe in 1984 than in 1983. In 1983 similar amounts of disease developed on all early cultivars and between 11% (Ulster Sceptre) and 32% (Maris Peer) shoots were pruned off. Maris Peer had a stem canker score lower than other cultivars in 1984 but more than half the shoots were pruned off. Shoot pruning on Estima, Ulster Prince and Ulster Sceptre was more common on plants from sprouted than non-sprouted seed. Between 30 and 50% of stolons were pruned off. After harvest in 1985, black scurf was least prevalent on Arran Comet and Maris Peer tubers and in 1984 on Arran Comet and Estima tubers from non-sprouted seed. Of the maincrop cultivars, King Edward plants from sprouted seed had many shoots pruned off in both years. Shoot pruning was also prevalent on Maris Piper and Pentland Squire plants from non-sprouted seed. Record had fewest pruned shoots and stolons and the lowest stem canker score. The disease was more severe on Pentland Crown and Maris Piper plants from non-sprouted than sprouted seed. Black scurf was most common on Cara and King Edward tubers in 1983 and on King Edward and Record tubers in 1984. In both years few shoots but many tubers were infected on plants from non-inoculated seed and the significance of this is discussed.     

Luminescence-based detection of Erwinia carotovora associated with rotting potato tubers

A luminescence-based marker system has been used to follow rotting of potato tubers by Erwinia carotovora. The early stages of infection could be detected by visual observation of areas of luminescence developing from the point of inoculation. During later stages luminescence was limited to peripheral regions and to central regions where the structure of the potato was destroyed. Luminometry demonstrated reduction in aerobic activity of infecting cells due to oxygen limitation. The system enables rapid and sensitive detection of active marked populations and distinction between aerobic and fermentative activity.     

Diversity of Rhizoctonia solani associated with pulse crops in different agro-ecological regions of India

Four hundred seventy Rhizoctonia solani isolates from different leguminous hosts originating from 16 agro-ecological regions of India covering 21 states and 72 districts were collected. The disease incidence caused by R. solani varied from 6.8 to 22.2 % in the areas surveyed. Deccan plateau and central highlands, hot sub-humid ecoregion followed by northern plain and central highlands and hot semi-arid ecoregion showed the highest disease incidence. R. solani isolates were highly variable in growth diameter, number, size and pattern of sclerotia formation as well as hyphal width. The isolates obtained from aerial part of the infected plants showing web blight symptoms produced sclerotia of 1–2 mm in size whereas, the isolates obtained from infected root of the plants showing wet root rot symptoms produced microsclerotia (<1 mm). Majority of R. solani isolates showed <8 μm hyphal diameter. Based on morphological characters the isolates were categorized into 49 groups. Seven anastomosis groups (AGs) were identified among the populations of R. solani associated with the pulse crops. The frequency (25.6 %) of AG3 was the highest followed by AG2–3 (20.9 %) and AG5 (17.4 %). The cropping sequence of rice/sorghum/wheat-chickpea/mungbean/urdbean/cowpea/ricebean influenced the dominance of AG1 (16.3 %). Phylogenetic analysis utilizing ITS-5.8S rDNA gene sequences indicated high level of genetic similarity among isolates representing different AGs, crops and regions. ITS groups did not correspond to the morphological characters. The sequence data from this article has been deposited with NCBI data libraries with JF701707 to JF701795 accession numbers.     

Control of pre- and post-emergence damping-off of potato seedlings caused by Rhizoctonia solani in Peru

Fungicide treatment of soil and true potato seed and the use of subsoil as a growth medium significantly reduced pre-emergence damping-off caused by Rhizoctonia solani. Attempts were made to determine safe levels of fungicides for soil and seed treatments. For seed, safe levels varied between 0–9% a.i. for pentachloronitrobenzene (PCNB) and 3-0% a.i. for mancozeb (dithane M-45). Fungicides only marginally reduced post-emergence damping-off of transplants raised from true potato seed but gave significant increase in yield. The prospects of using fungicide treatment of true seed to control damping-off in seed beds and in direct field sowing are discussed.     

Methylation and aging in Rhizoctonia solani

Our earlier studies had shown that as fungi age, many of their vital functions decrease; in Rhizoctonia solani, protein synthesis is one of the functions so affected. We now find that the ability to methylate tRNA, a vital component of the protein synthesizing system, also decreases with age. This methylation of Escherichia coli tRNA by R. solani methylase preparations increased with the concentration of enzyme and with time of incubation; in both cases the rate of increase was considerably higher for preparations from young cells than for those from old cells. The methylation reaction also increased with the concentration of substrate tRNA, with temperature, at least to 45° C, and with pH to 9.0. Methylase preparations from R. solani methylated both exogenous E. coli tRNA and yeast tRNA, but were only weakly active on isolated R. solani tRNA. However, acid-precipitated methylases from R. solani were very effective in methylating the homologous exogenous tRNA. Regardless of the source of the tRNA used as substrate, the methylases from older cells were always less active than those from young cells from the same mycelium. No methylase inhibitor was detected in the fungus.     

Acetone Inhibition of Rhizoctonia solani Growth

Acetone (5% v/v) inhibited growth of four isolates of Rhizoctonia solani which differ in their pathogenicity on squash seedlings. Acetone (5% v/v) showed best inhibition on the most aggressive isolate 4 followed by the less aggressive ones (1, 2 and 3, respectively); IAA had no effect on the growth of all R. solani isolates compared to the controls.     

Genetic Variability in Pectic Enzymes of Rhizoctonia solani Isolates Causing Bare-patch Disease of Cereals

Field isolates of Rhizoctonia solani obtained from three discrete bare patches in a wheat field in Western Australia were characterized by pectic zymogram grouping. The genetic background of pectic enzymes was analysed by comparing the zymograms of asexual homokaryons and sexual progenies derived from field isolates. The 170 field isolates obtained from the field site produced indistinguishable pectic zymograms. However, variations among field isolates of the same zymogram group were detected, on the basis of zymograms of their resultant protoplast-regenerated cultures. Asexual sibling homokaryons derived from each of the field isolates were heterogeneous for their pectic enzymes. Homokaryons with a common heterokary on incompatibility factor, obtained from a field isolates were homogeneous for pectic enzymes. Basidiospore progenies of a field isolate segregated widely in pectic zymograms. It appeared that the expression of pectic enzymes by field isolates involved multiple genetic factors. The variation of zymograms among homokaryotic strains suggests that each field isolate of R. solani contains two types of nuclei, although cells of vegetative hyphae are multinucleate.     

Overexpression of snakin-1 gene enhances resistance to Rhizoctonia solani and Erwinia carotovora in transgenic potato plants

Snakin-1 (SN1), a cysteine-rich peptide with broad-spectrum antimicrobial activity in vitro, was evaluated for its ability to confer resistance to pathogens in transgenic potatoes. Genetic variants of this gene were cloned from wild and cultivated Solanum species. Nucleotide sequences revealed highly evolutionary conservation with 91-98% identity values. Potato plants (S. tuberosum subsp. tuberosum cv. Kennebec) were transformed via Agrobacterium tumefaciens with a construct encoding the S. chacoense SN1 gene under the regulation of the ubiquitous CaMV 35S promoter. Transgenic lines were molecularly characterized and challenged with either Rhizoctonia solani or Erwinia carotovora to analyse whether constitutive in vivo overexpression of the SN1 gene may lead to disease resistance. Only transgenic lines that accumulated high levels of SN1 mRNA exhibited significant symptom reductions of R. solani infection such as stem cankers and damping-off. Furthermore, these overexpressing lines showed significantly higher survival rates throughout the fungal resistance bioassays. In addition, the same lines showed significant protection against E. carotovora measured as: a reduction of lesion areas (from 46.5 to 88.1% with respect to the wild-type), number of fallen leaves and thickened or necrotic stems. Enhanced resistance to these two important potato pathogens suggests in vivo antifungal and antibacterial activity of SN1 and thus its possible biotechnological application.     

立枯丝核菌营养菌丝多型性观察

采用了2种不同的方法对立枯丝核菌营养菌丝的形态进行了观察和比较,观察到2种不同的营养菌丝的形态,即菌核类和假分生孢子类。为以后进一步研究立枯丝核菌营养菌丝的多型性奠定了一定的基础。     

Quantitative proteomic analysis of differentially expressed proteins in tubers of potato plants differing in resistance to Dickeya solani

Plant and Soil - This study aims the detection of proteins associated with increased resistance of tubers to necrotrophic bacteria Dickeya solani in tetraploid and diploid potato plants....     

Fungicidal control of Rhizoctonia solani in soil amended with organic manures

Amendments of nutrient-deficient soil with three organic manures and one non-edible oil-cake reduced the disease controlling potential of methoxyethyl mercury chloride (MEMC), quintozene and carbendazim used as seed treatments on cowpea and cotton against seedling rot caused by Rhizoctonia solani. Biogas sludge (BGS) and farm yard manure (FYM) nullified the activity of MEMC and quintozene and reduced markedly the efficacy of carbendazim. Humic acid extracted from BGS inactivated MEMC and carbendazim but had little effect on quintozene. Green manure (Sesbania aculeata) slightly reduced the efficacy of MEMC only. Soil amendment with mahua (Madhuca indica) cake and soil drench with its aqueous extract greatly reduced the efficacy of the three fungicides.     

Molecular diversity analysis of Rhizoctonia solani isolates infecting various pulse crops in different agro-ecological regions of India

Genetic diversity of 89 isolates of Rhizoctonia solani isolated from different pulse crops representing 21 states from 16 agro-ecological regions of India, 49 morphological, and 7 anastomosis groups (AGs) was analyzed using 12 universal rice primers (URPs), 22 random amplified polymorphic DNA (RAPD), and 23 inter-simple sequence repeats (ISSR) markers. Both URPs and RAPD markers provided 100?% polymorphism with the bands ranging from 0.1 to 5?kb in size, whereas ISSR markers gave 99.7?% polymorphism with the bands sizes ranging from 0.1 to 3?kb. The marker URP 38F followed by URP13R, URP25F, and URP30F, RAPD marker R1 followed by OPM6, A3 and OPA12 and ISSR3 followed by ISSR1, ISSR4, and ISSR20 produced the highest number of amplicons. R. solani isolates showed a high level of genetic diversity. Unweighted pair group method with an arithmetic average (UPGMA) analysis grouped the isolates into 7 major clusters at 35?% genetic similarity using the three sets of markers evaluated. In spite of using three different types of markers, about 95?% isolates shared common grouping patterns. The majority of the isolates representing various AGs were grouped together into different sub-clusters using all three types of markers. Molecular groups of the isolates did not correspond to agro-ecological regions or states and crops of the origin. An attempt was made for the first time in the present study to determine the genetic diversity of R. solani populations isolated from different pulse crops representing various AGs and agro-ecological regions.