A PCR-based method to distinguish fungi of the rice sheath-blight complex, Rhizoctonia solani, R. oryzae and R. oryzae-sativae

Identification of Rhizoctonia solani, R. oryzae and R. oryzae-sativae, components of the rice sheath disease complex, is extremely difficult and often inaccurate and as a result may hinder the success of extensive breeding programmes throughout Asia. In this study, primers designed from unique regions within the rDNA internal transcribed spacers have been used to develop a rapid PCR-based diagnostic test to provide an accurate identification of the species on rice. Tests on the specificity of the primers concerned showed that they provide the means for accurate identification of the Rhizoctonia species responsible for sheath diseases in rice.

Studies on diseases of bamboos and nursery management of rhizoctonia web bligi-it in kerala

Bamboos are vulnerable to various diseases which affect them in nurseries, plantations as well as in natural stands. In India, rot and blight of emerging culms have already been identified as the limiting factor of the bamboo production in many bamboo growing areas, especially in the coastal belts of Orissa (Jamaluddin et a1., 1992). Similarly, foliage blight and rust have been recorded to pose threat to nursery as well as outplanted seedlings which are in the early establishnent phase (Bakshi et a1., 1972; Harsh et a1., 1989). With the increased emphasis and priority on raising multipurpose tree species, large—scale planting of bamboos has been initiated recently in the State. Limited experience in raising the bamboo seedlings together with the lack of information on bamboo diseases and their control measures often resulted in partial to complete failure of many nurseries. Also, poor handling of bareroot seedlings for outplanting affected seriously the planting programme. This was clearly reflected by the large-scale nortality of outplanted young seedlings reported from many plantations. So far, no systanatic attempt has been made to study the diseases affecting bamboos in nurseries, plantations and natural stands in the country. Hence, the present investigation was taken up to conduct a systematic study of the diseases affecting bamboos in Kerala.

Biological control of Rhizoctonia solani Damping-off with a bacterium symbiotically associated with Steinernema abbasi

Rhizoctonia solani is a causal agent of damping-off of may cultivated plants. An isolate of the bacterium Pseudomonas oryzihabitans, symbiotically associated with the entomopathogenic nematode Steinernema abbasi, strongly inhibited the pathogen in vitro. The bacterium was firmly attached onto fungus mycelia and degraded the cell walls of the pathogen. In greenhouse experiments, bacterial suspension in sterile water applied in the soil, effectively controlled damping-off of radish.

Characterization and origin of infection of Rhizoctonia solani associated with Brassica oleracea crops in the UK

An extensive study was conducted to determine where in the production chain Rhizoctonia solani became associated with UK module-raised Brassica oleracea plants. In total, 2600 plants from 52 crops were sampled directly from propagators and repeat sampled from the field. Additional soil, compost and water samples were collected from propagation nurseries and screened using conventional agar isolation methods. No isolates of R. solani were recovered from any samples collected from propagation nurseries. Furthermore, nucleic acid preparations from samples of soil and compost from propagation nurseries gave negative results when tested for R. solani using real-time PCR. Conversely, R. solani was recovered from 116 of 1300 stem bases collected from field crops. All the data collected suggested R. solani became associated with B. oleracea in the field rather than during propagation. Parsimony and Bayesian phylogenetic studies of ribosomal DNA suggested the majority of further classified isolates belonged to anastomosis groups 2-1 (48/57) and AG-4HGII (8/57), groups known to be pathogenic on Brassica spp. in other countries. Many R. solani isolates were recovered from symptomless plant material and the possibilities for such an association are discussed.

Molecular tools to investigate Rhizoctonia solani distribution in soil

Real-time PCR protocols were developed to detect and discriminate 11 anastomosis groups (AGs) of Rhizoctonia solani using ribosomal internal transcribed spacer (ITS) regions (AG-1-IA, AG-1-IC, AG-2-1, AG-2-2, AG-4HGI+II, AG-4HGIII, AG-8) or beta-tubulin (AG-3, AG-4HGII, AG-5 and AG-9) sequences. All real-time assays were target group specific, except AG-2-2, which showed a weak cross-reaction with AG-2tabac. In addition, methods were developed for the high throughput extraction of DNA from soil and compost samples. The DNA extraction method was used with the AG-2-1 assay and shown to be quantitative with a detection threshold of 10-7 g of R. solani per g of soil. A similar DNA extraction efficiency was observed for samples from three contrasting soil types. The developed methods were then used to investigate the spatial distribution of R. solani AG-2-1 in field soils. Soil from shallow depths of a field planted with Brassica oleracea tested positive for R. solani AG-2-1 more frequently than soil collected from greater depths. Quantification of R. solani inoculum in field samples proved challenging due to low levels of inoculum in naturally occurring soils. The potential uses of real-time PCR and DNA extraction protocols to investigate the epidemiology of R. solani are discussed.

Distinctively variable sequence-based nuclear DNA markers for multilocus phylogeography of the soybean- and rice-infecting fungal pathogen Rhizoctonia solani AG-1 IA

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Binucleate Rhizoctonia sp AG G causing root rot in yacon (Smallanthus sonchifolius) in Brazil

A new rot caused by a binucleate Rhizoctonia sp. affecting the tuberous root cortex of the domesticated yacon (Smallanthus sonchifolius) has been observed in Brazil. Isolates of a binucleate Rhizoctonia sp. were collected from roots with rot symptoms and characterized by the number of nuclei per cell, hyphal anastomosis, RAPD molecular markers, ITS-5.8S rDNA sequence and pathogenicity tests. All isolates had a mean of 1.9-2.2 nuclei per cell and anastomosed with the binucleate Rhizoctonia sp. AG G-tester strain. RAPD analysis was carried out between 11 isolates recovered from yacon and 11 AG (A, Ba, Bb, Bo, C, D, F, G, O, P, Q) standard testers of binucleate Rhizoctonia sp. Genetic similarities of 94.8-100% were observed among isolates of the binucleate Rhizoctonia sp. from yacon and all isolates were genetically more closely related to the AG G tester than other strains according to UPGMA analysis using RAPD markers. Homologies of complete ITS nucleotide sequences were 100% between binucleate isolates of Rhizoctonia sp. from yacon and the AG G tester. According to pathogenicity tests, the isolates caused typical rot symptoms of yacon tubers 90 days after inoculation.

Identification of Rhizoctonia solani AG 1-IB in lettuce, AG 4 HG-I in tomato and melon, and AG 4 HG-III in broccoli and spinach, in Brazil

Fungi isolated in Brazil, from lettuce, broccoli, spinach, melon and tomato, were identified as Rhizoctonia solani. All lettuce isolates anastomosed with both AG 1-IA and IB subgroups and all isolates from broccoli, spinach, melon and tomato anastomosed with AG 4 subgroup HG-I, as well as with subgroups HG-II and HG-III. DNA sequence analyses of ribosomal internal transcribed spacers showed that isolates from lettuce were AG 1-IB, isolates from tomato and melon were AG 4 HG-I, and isolates from broccoli and spinach were AG 4 HG-III. The tomato isolates caused stem rot symptoms, the spinach, broccoli and melon isolates caused hypocotyl and root rot symptoms on the respective host plants and the lettuce isolates caused bottom rot. This is the first report on the occurrence in Brazil of R. solani AG 4 HG-I in tomato and melon, of AG 4 HG-III in broccoli and spinach and of AG 1-IB in lettuce.

Caracterização citomorfológica, cultural, molecular e patogênica de Rhizoctonia solani Kühn associado ao arroz em Tocantins, Brasil

No Estado do Tocantins, no Norte do Brasil, a incidência de rizoctoniose no arroz é importante, causando danos significativos em lavouras de arroz irrigado. O principal objetivo deste trabalho foi determinar o grupo de anastomose (AG) de isolados de R. solani associados ao arroz naquela região, testando a hipótese de que esses isolados pertencem ao grupo padrão de anastomose AG-1 IA, que também é o agente causal da mela em soja em áreas úmidas do Norte do Brasil. Todos os quatro isolados de arroz foram caracterizados, através de fusão de hifas, como AG-1 IA. A caracterização cultural, em função das temperaturas basais (mínimas, máximas e ótimas), evidenciou que os isolados de R. solani de arroz apresentaram perfis semelhantes aos padrões AG-1 IA, AG-1 IB e AG-1 IC. Os isolados de arroz foram caracterizados como autotróficos para tiamina assim como os isolados padrões AG-1 IA, IB, IC, AG-4 HGI e o isolado da mela da soja. O teste de patogenicidade em plantas de arroz cultivar IRGA-409 e de patogenicidade cruzada à cultivar IAC-18 de soja (suscetível à mela), indicou que além de causar a queima da bainha em arroz, esses isolados causam mela em soja. da mesma forma, o isolado SJ-047 foi patogênico ao arroz. As seqüências de bases de DNA da região ITS-5.8S do rDNA dos isolados do arroz foram similares às seqüências do AG-1 IA, depositadas no GenBank® - NCBI. A filogenia do ITS-rDNA indicou um grupo filogenético comum formado pelos isolados do arroz, o isolado da soja e o isolado teste do AG-1 IA. Assim, com base em características citomorfológicas, culturais, filogenéticas e patogênicas, foi confirmada a hipótese de que os isolados de R. solani patógenos de arroz do Estado do Tocantins pertencem ao grupo de anastomose AG-1 IA, além da indicação de que esses isolados podem também causar a mela em soja.

Highly polymorphic in silico-derived microsatellite loci in the potato-infecting fungal pathogen Rhizoctonia solani anastomosis group 3 from the Colombian Andes

Fourteen polymorphic microsatellite DNA markers derived from the draft genome sequence of Rhizoctonia solani anastomosis group 3 (AG-3), strain Rhs 1AP, were designed and characterized from the potato-infecting soil fungus R. solani AG-3. All loci were polymorphic in two field populations collected from Solanum tuberosum and S. phureja in the Colombian Andes. The total number of alleles per locus ranged from two to seven, while gene diversity (expected heterozygosity) varied from 0.11 to 0.81. Considering the variable levels of genetic diversity observed, these markers should be useful for population genetic analyses of this important dikaryotic fungal pathogen on a global scale.

Genetic Structure of Populations of the Rice-Infecting Pathogen Rhizoctonia solani AG-1 IA from China

Sheath blight disease (SBD) on rice, caused by Rhizoctonia solani AG-1 IA, is one of the most devastating rice diseases on a global basis, including China (in Eastern Asia), the world's largest rice-growing country. We analyzed the population genetics of nine rice-infecting populations from China using nine microsatellite loci. One allopatric population from India (Southern Asia) was included in the analyses. In total, 300 different multilocus genotypes were found among 572 fungal isolates. Clonal fractions within rice fields were 16 to 95%, suggesting that sclerotia were a major source of primary inoculum in some fields. Global Phi(ST) statistics (Phi(ST) = 42.49; P <= 0.001) were consistent with a relatively high level of differentiation among populations overall; however, pairwise comparisons gave nonsignificant R(ST) values, consistent with contemporary gene flow among five of the populations. Four of these populations were located along the Yangtze River tributary network. Gene flow followed an isolation-by-distance model consistent with restricted long-distance migration. Historical migration rates were reconstructed and yielded values that explained the current levels of population subdivision. Except for one population which appeared to be strictly clonal, all populations showed evidence of a mixed reproductive mode, including both asexual and sexual reproduction. One population had a strictly recombining structure (all loci were in Hardy-Weinberg equilibrium) but the remaining populations from China and the one from India exhibited varying degrees of sexual reproduction. Six populations showed significant F(IS) values consistent with inbreeding.

Genetic structure of populations of Rhizoctonia solani anastomosis group-1 IA from soybean in Brazil

The Basidiomycete fungus Rhizoctonia solani anastomosis group (AG)-1 IA is a major pathogen of soybean in Brazil, where the average yield losses have reached 30 to 60% in some states in Northern Brazil. No information is currently available concerning levels of genetic diversity and population structure for this pathogen in Brazil. A total of 232 isolates of R. solani AG1 IA were collected from five soybean fields in the most important soybean production areas in central-western, northern, and northeastern Brazil. These isolates were genotyped using 10 microsatellite loci. Most of the multilocus genotypes (MLGTs) were site-specific, with few MLGTs shared among populations. Significant population subdivision was evident. High levels of admixture were observed for populations from Mato Grosso and Tocantins. After removing admixed genotypes, three out of five field populations (Maranhao, Mato Grosso, and Tocantins), were in Hardy-Weinberg (HW) equilibrium, consistent with sexual recombination. HW and gametic disequilibrium were found for the remaining soybean-infecting populations. The findings of low genotypic diversity, departures from HW equilibrium, gametic disequilibrium, and high degree of population subdivision in these R. solani AG-1 IA populations from Brazil are consistent with predominantly asexual reproduction, short-distance dispersal of vegetative propagules (mycelium or sclerotia), and limited long-distance dispersal, possibly via contaminated seed. None of the soybean-infecting populations showed a reduction in population size (bottleneck effect). We detected asymmetric historical migration among the soybean-infecting populations, which could explain the observed levels of subdivision.

Divergence Between Sympatric Rice- and Maize-Infecting Populations of Rhizoctonia solani AG-1 IA from Latin America

The basidiomycetous fungus, Rhizoctonia solani anastomosis group (AG)-1 IA is a major pathogen in Latin America causing sheath blight (SB) of rice Particularly in Venezuela. the fungus also Causes banded leaf and sheath blight (BLSB) oil maize, which is considered all emerging disease problem where maize replaced traditional rice-cropping areas or is now planted in adjacent. fields Our goals in this study Were 10 elucidate (i) the effects of host specialization on gene flow between sympatric and allopatric rice and maize-infecting fungal populations and (ii) the reproductive mode of the fungus, looking for evidence of recombination in total, 375 isolates of R. solani AG1 IA sampled from three sympatric rice and maize fields in Venezuela (Porutuguesa State) and two allopatric rice fields from Colombia (Meta State) and Panama (Chiriqui State) were genotyped Using, 10 microsatellite loci Allopatric populations from Venezuela. Colombia. and Panama were significantly differentiated (Phi(ST), of 0 16 to 0 34). Partitioning of the genetic diversity indicated differentiation between sympatric populations from different host species, with 17% of the total genetic variation distributed between hosts while only 3 to 6% wits distributed geographically among the sympatric Venezuelan Fields We detected symmetrical historical migration between the rice- and the maize-infecting populations from Venezuela Rice- and maize-derived isolates were able to infect built rice and maize but were more aggressive Oil their original hosts, consistent with host specialization. Because the maize- and rice-infecting populations are still cross-pathogenic, we postulate that the genetic differentiation was relatively recent and mediated via a host shift. An isolation with nu.-ration analysis indicated that the maize-infecting population diverged from the rice-infecting population between 40 and 240 years ago Our findings also suggest that maize-infecting Populations have a mainly recombining reproductive system whereas the rice-infecting Populations have a Mixed reproductive system in Latin America

Highly polymorphic microsatellite loci in the rice- and maize-infecting fungal pathogen Rhizoctonia solani anastomosis group 1 IA

Ten polymorphic microsatellite loci were isolated and characterized from the rice- and maize-infecting Basidiomycete fungus Rhizoctonia solani anastomosis group AG-1 IA. All loci were polymorphic in two populations from Louisiana in USA and Venezuela. The total number of alleles per locus ranged from four to eight. All 10 loci were also useful for genotyping soybean-infecting R. solani AG-1 isolates from Brazil and USA. One locus, TC06, amplified across two other AG groups representing different species, showing species-specific repeat length polymorphism. This marker suite will be used to determine the global population structure of this important pathogenic fungus.