Deleterious effects of fungi isolated from paddy soils on seminal root of rice

Soil samples were collected from rice paddies at 22 locations in northeastern Honshu, Japan. In 20 of the samples, seedling growth of rice was improved by soil pasteurization (aerated steaming at 60°C for 30 min), although no typical disease symptoms were observed in the seedlings grown in the untreated soil samples. In most locations, rice seedlings grew better in a potting medium containing root material from plants grown in the pasteurized portion of a soil sample than in a medium with root material from plants grown in the unpasteurized portion of the same sample. The results suggest that microorganisms that restrained rice seedling growth may be common in the soils of rice paddies. Approximately 800 isolates were obtained from seedling roots grown in unpasteurized soils and grouped by cultural and microscopic morphologies. The deleterious effects of 79 isolates from 21 major groups, most of which were soil-dwelling taxa, were examined after direct inoculation of the seminal roots. Isolates of Curvularia sp., Cirrenalia sp., Eppicoccum nigrum, Fusarium graminearum, F. oxysporum, Gliocladium virens, Humicola sp., Penicillium sp., Rhizoctonia oryzae-sativae, Sclerotium hydrophilum, Trichoderma aureoviride, and T. harzianum inhibited root growth, suggesting that deleterious root-infecting fungi were more common in paddy soil than previously thought. These fungi may be involved in the restraint of rice seedling growth.     

A case study of infection of rice roots by deleterious fungi and minor pathogenic<Emphasis Type="Italic"> Pythium</Emphasis> species in a paddy field

Roots of rice plants grown in paddy fields in a transplant culture system were collected seven times between 8 and 22 weeks after transplanting, for two crop seasons. Rice seedlings grown in a potting medium amended with the collected roots were significantly shorter than those grown in the same medium either without the addition of the roots or amended with pasteurized, collected roots indicating that seedlings were inhibited by heat-labile microorganisms on the collected rice roots. Ninety-five and 172 pure cultures of Pythium spp. and fungi, respectively, were isolated from the rice roots collected 5 or 7.5 weeks after transplanting in the fields. Among these microorganisms, Pythium aristosporum inhibited seedling growth in greenhouse experiments, and Acremonium sp., Alternaria sp., Epicoccum nigrum, Fusarium sp., Massarina sp., Penicillium spp., Rhinocladiella sp., Stemphylium sp., Trichocladium sp., and several unidentified fungi inhibited seminal root growth in in vitro experiments. These microorganisms might be involved in the inhibition of seedling growth in soils amended with the rice roots collected from the paddy field. Thus, roots of rice plants at the middle stage of growth transplanted into paddy fields can harbor pathogenic or deleterious fungi or Pythium sp(p). The effects of these microorganisms on rice growth in paddy fields are discussed.     

水稻旱育秧立枯病致病菌鉴定及药剂防治研究

从水稻旱育秧病苗上分离到67个菌株 ,经鉴定分属于镰刀菌58个、腐霉菌7个、丝核菌2个。经回接测定其致病性 ,结果表明致病的镰刀菌主要是串珠镰刀菌 (Fusarium moniliforme) ;腐霉菌中主要是盐腐霉 (Pythium salinum)、间生腐霉 (P .interedium)和顶生腐霉 (P .acrogenum) ;丝核菌为立枯丝核菌 (Rhizoctonia solani)。接种试验表明串珠镰刀菌在6～8d龄幼苗的根中部侵染发病率最高 ,腐霉菌和丝核菌在一叶一心期茎基部侵染发病率最高。药剂试验表明以浸种灵(二硫氰基甲烷)、土菌消(hymexazol)、甲霜灵(metalaxyl)等种子处理加土壤处理 ,防效优于单独种子处理或土壤处理。     

Suppression of Fusarium wilt of spinach with hypovirulent binucleate <Emphasis Type="Italic">Rhizoctonia</Emphasis>

 Four isolates of hypovirulent binucleate Rhizoctonia (HBNR) were evaluated for their ability to control Fusarium wilt of spinach (FWS) caused by Fusarium oxysporum f. sp. spinaciae (FOS). Fourteen-day-old spinach seedlings grown in paper pots with HBNR-amended soil (1% w/w ground barley grain inoculum) were transferred to artificially pathogen-infested soil. Treatments with HBNR isolates significantly (P = 0.05) reduced disease and discoloration severity by 56%–100% and 52%–100%, respectively. The numbers of colony-forming units of FOS per gram fresh weight in petioles or roots were reduced significantly (P = 0.01) in the plants treated with HBNR. HBNR isolates were well reisolated from the roots inside paper pots where they were inoculated, whereas inconsistent colonization of HBNR was recorded from the roots outside paper pots where only pathogen was inoculated. Root extracts from HBNR-treated and pathogen-challenged plants significantly inhibited germination and germling length of FOS. The fresh weight of spinach leaves in the HBNR-treated plants increased significantly (P = 0.01), as much as 53%–63%, over the untreated and pathogen-challenged plants. This is the first report of biocontrol of FWS by HBNR. Received: July 18, 2002 / Accepted: October 22, 2002 Acknowledgments We are grateful to Dr. Komada for providing nonpathogenic Fusarium F13. The senior author (A.M.) thanks the Ministry of Education, Culture, Sports, Science, and Technology (Monbukagakusho) Japan, for financial assistance.     

Comparative pathogenicity of Pythium species associated with poor seedling establishment of rice in Southern Australia

Isolates of Pythium attributable to thirteen named species and to species group F (sensu van der Plaats-Niterink) were isolated from roots of field-grown rice seedlings showing poor root development and lack of vigour. P. arrhenomanes and group F were the most commonly isolated species. The roots of 15-week-old rice plants yielded only P. flevoense, P. vanterpoolii, P. rostratum and Pythium group G of which the latter two were not obtained from seedling roots.
In pot tests with representative Pythium isolates, P. arrhenomanes was most pathogenic to rice seedlings, causing pre- and post-emergence death and average reductions in shoot and root growth in surviving seedlings of 48 and 70%, respectively. P. irregulare killed seedlings before but not after emergence, whilst the response of rice seedlings to P. myriotylum apparently depended on the length and timing of a cold shock treatment. P. pyrilobum did not reduce seedling number or shoot growth but reduced root growth. P. vanterpoolii and group F were not pathogenic in the tests. P. coloratum, P. echinulatum, P. flevoense, P. oedochilum, P. oligandrum, P. periilum, P. pyrilobum and P. rostratum were isolated infrequently and had no adverse effects on seedlings in pathogenicity tests, although P. myriotylum, P. oligandrum , and P. periilum were associated with significant increases in shoot growth. P. tracheiphilum was isolated from one site but its pathogenicity was not tested.
This is the first record of P. coloratum, P. echinulatum, P. flevoense, P. irregulare, P. oedochilum, P. oligandrum, P. periilum, P. pyrilobum and P. tracheiphilum from rice roots.     

Role of the α subunit of heterotrimeric G-protein in probenazole-inducing defense signaling in rice

 Certain cellular responses to stresses and stimuli are regulated by a G-protein-mediated signaling pathway. A rice dwarf mutant that is defective in the α subunit of the heterotrimeric G-protein was found to be fully protected from blast fungus by the plant activator probenazole (PBZ) despite the 1-day delay in induction of the PR-10 gene PBZ1 by PBZ. These results suggest that the signaling pathway for protection by PBZ is not via the G-protein, although G-protein is involved in the induction of PBZ1 by PBZ. Received: March 27, 2002 / Accepted: August 20, 2002     

Ratio of rice reflectance for estimating leaf blast severity with a multispectral radiometer

 Rice reflectance was measured to determine the spectral regions most sensitive to leaf blast infection with a multispectral radiometer. As disease severity increased, reflectance also increased in the 400–500 nm (blue), 570–700 nm (red), and 900–2000 nm regions but decreased in the 500–570 nm and 700–900 nm regions. The increased reflectance in the blue and red regions may be attributed to decreased chlorophyll and carotenoid contents in response to the blast infection. The maximum and minimum reflectance differences occurred at 680 nm and 760 nm for the nondiseased and diseased rice, respectively. The spectral location of maximum sensitivity was 675 nm regardless of disease severity. Rice reflectance ratios were evaluated as indicators of leaf blast severity. Two ratios, R550/R675 (reflectance at 550 nm divided by reflectance at 675 nm), and R570/R675 quantified the significant disease severity. These wavelengths were selected based on the sensitivity minima and maxima. The ratios of nondiseased rice plants varied depending on growth stage. The variation in ratios must be considered when they are used to estimate leaf blast severity. Received: April 2, 2002 / Accepted: August 12, 2002     

Management of bakanae and bacterial seedling blight diseases in nurseries by irradiating rice seeds with atmospheric plasma

The control of seedborne rice seedling diseases in the seed beds is important to avoid epidemics in rice nurseries and paddies, which may result in severe yield loss. Recently, irradiation with plasma containing electrons, creating positive or negative ions and neutral species, has been shown to have an antimicrobial effect, probably via generation of reactive oxygen species. This study examines whether two seedborne rice seedling diseases, bakanae disease caused by the fungal pathogen Fusarium fujikuroi, and bacterial seedling blight caused by Burkholderia plantarii, are suppressed by irradiation of infected rice seeds with atmospheric plasma. Seed germination and seedling growth were not inhibited in plasma‐treated healthy seeds. When F. fujikuroi‐infected rice seeds were irradiated with plasma after being immersed in sterile distilled water, bakanae disease severity index and the percentage of plants with symptoms were reduced to 18.1% and 7.8% of non‐irradiated control, respectively, depending on the duration of plasma irradiation. The bacterial seedling blight disease index was also reduced by plasma irradiation in vacuum‐inoculated seeds to 38.6% of the non‐irradiated control, and in infected seeds harvested from spray‐inoculated heads of rice plants to 40.1% of the control. Therefore, plasma irradiation seems to be effective in controlling two independent seedborne rice seedling diseases.     

Rapid detection of chitosanase activity in chitosanase gene-transformed strains of <Emphasis Type="Italic">Enterobacter cloacae</Emphasis> by lytic infection of specific bacteriophages

 In combination with lytic infection by virulent phages, a simple method for monitoring transgenic strains of Enterobacter cloacae was developed in this study. First, 15 strains of E. cloacae were used as indicator bacteria to isolate virulent phages with different host ranges. Of the phages isolated, five isolates (EcP-22, -35, -45, -55, and -70) were used to construct a set of virulent phages corresponding to all strains of E. cloacae. Using this phage set, a rhizosphere strain (KRM-055E) of E. cloacae was effectively screened from field soil. KRM-055E was transformed with a prokaryotic chitosanase gene csnSM1 and infected with the phage EcP-03, which can lyse the strain most effectively. The lysis of KRM-055E/csn occurred 2 h after inoculation, and the chitosanase activity was simply detected by dropping the lysate onto an agar plate containing glycol chitosan. The positive signal for chitosanase activity was detected in the 2-h lysates, and the signal intensity reached a maximum in the 5-h lysate. The present assay was simple, rapid, inexpensive, easy to perform, and applicable to another strains. Received: August 2, 2002 / Accepted: October 31, 2002 Acknowledgments This work was supported in part by a grant (no. 99L01205) from the “Research for the Future” program of the Japan Society for the Promotion of Science. We are grateful to Dr. M. Sato, National Institute of Agrobiological Science, Dr. H. Okamoto, Fukui Agricultural Experiment Station, and Dr. K. Tsuda, Kyoto Prefectural Institute of Agricultural Biotechnology, for kindly providing E. cloacae strains. We thank Dr. P. Park, Kobe University, for technical support with the electron microscopic observations.     

Colonization of the vegetative stage of rice plants by the false smut fungus Villosiclava virens,as revealed by a combination of species‐specific detection methods

Rice false smut disease caused by the ascomycete fungus Villosiclava virens (Clavicipitaceae) reduces rice yield worldwide. It invades rice panicles and forms dark‐green false smut balls composed of thick‐walled conidia. Although the infection process during the booting stage is well studied, its infection route before this is unclear. It was hypothesized that the thick‐walled conidia in soil penetrate rice roots, and the fungus latently colonizes roots and tiller buds at the vegetative stage. This hypothesis was tested using species‐specific detection methods. First, real‐time PCR with species‐specific primers and probe was used to estimate thick‐walled conidial number in the paddy field soil. Secondly, nested PCR with species‐specific primers showed that fungal DNA was detected in roots and shoot apices of rice plants in the vegetative stage. Thirdly, colourimetric in situ hybridization with a species‐specific oligonucleotide probe targeting 18S rRNA suggested that sparse mycelia or tightly condensed mycelia were present on the external surface of tiller buds enveloped by juvenile leaf sheaths at the vegetative stage. Thin hyphae were found around leaf axils at the surface of elongated stems at the heading stage, and the fungal hyphae grew in the rice root tissues. In addition, it was demonstrated that eGFP‐tagged transformants of the fungus invaded rice roots and colonized the surface of roots and leaf sheaths under artificial conditions.     

Root growth inhibition of rice by bensulfuron

Bensulfuron inhibited root growth of waterseeded rice (Oryza sativa L.) plants in a solution culture system. Rice root growth was more sensitive to bensulfuron than was shoot growth. A1 2.5 × 10^-9 M bensulfuron, plant height, number of leaves and number of roots were not significantly affected, while root growth was reduced, Seedlings treated 3 or 4 days after emergence showed a significant reduction of total root length by the second day after treatmeni. Root growth inhibition was proportional to duration of treatment; however, treatment for 2 days was necessary to affect root growth, Bensulfuron decreased the mitotic index and the mitotic height in root tips. At 5 days after treatment, only 0.7% of the cells were dividing and mitotic height was 68% of control     

Transmission of rice blast from seeds to adult plants in a non‐systemic way

Rice blast, caused by the fungal pathogen Magnaporthe oryzae, is a serious threat to rice production worldwide. In temperate regions, where rice is not cultivated for several months each year, little is known about the initial onset of the disease in the field. The main overwintering and primary inoculum sources reported are infested residues and seeds, but the subsequent steps of the disease cycle are largely unknown, even though a systemic infection has been proposed but not demonstrated. The present work follows rice blast progression in infected seeds from germination to seedling stage, with direct and detailed microscopic observations under both aerobic conditions and water seeding. With the use of GFP‐marked M. oryzae strains, it was shown that spores are produced from contaminated seeds, infect emerging seedling tissues (coleoptile and primary root) and produce mycelium that colonizes the newly formed primary leaf and secondary roots. Using different rice cultivars exhibiting distinct levels of resistance/susceptibility to M. oryzae at the 2/4‐leaf stage, it was observed that resistance or susceptibility of a considered genotype is already established at the seedling stage. The results also showed that when plants are inoculated either at ripening stage (mature panicles), heading stage (flowering/immature panicles) or even before heading (flag leaf fully developed), they produce infested seeds. These seeds produce contaminated seedlings that mostly die and serve as an inoculum source for healthy neighbouring plants, which gradually develop disease symptoms on leaves. The possible rice blast disease cycle was reconstructed on irrigated rice in temperate regions.     

Laboratory studies on the influence of the earthworm Eukerria saltensis (Beddard) (Oligochaeta: Ocnerodrilidae) on overlying water quality and rice plant establishment

Dense infestations of a peregrine oligochaete worm, Eukerria saltensis (Beddard), have been linked to rice crop failures in southern New South Wales, Australia. The influence of E. saltensis on water quality and rice plant establishment was investigated in a series of laboratory experiments using a flooded Riverina clay soil. Worm densities of 20 and 40 per container (2548 and 5096 worms/m², respectively) significantly increased water turbidity after 7 days incubation. Longer incubation periods led to turbidity levels of over 500 NTU being achieved (40 worms per container, 21 days incubation). Water pH was significantly reduced by densities of 10, 20, and 40 worms per container after 7 days under cyclical illumination, however in continuous darkness significant changes in pH related to worm density were only found after 21 days incubation. Nitrogen as NH₄+ and total phosphorus increased significantly in the overlying water in response to increasing worm densities after 21 days incubation, however nitrogen as nitrate/nitrite and soluble phosphorus did not. Algal production (measured as extracted chlorophyll a concentration) was unaffected by the worms, reflecting the low concentrations of available phosphorus in all treatments. Rice plants grown in containers with worms produced significantly longer and heavier shoots than control plants. Root systems were unaffected, and there was no evidence of root abrasion. Evaluation of rice seed stratification in the soil profile indicates that rice seeds can be passively transported below the soil surface by the feeding and tunnelling activity of E. saltensis. Our results suggest that E. saltensis impedes the establishment of aerially-sown rice crops primarily by increasing water turbidity. Plants respond to high turbidity by partitioning more of their growth into shoot production, and consequently become vulnerable touprooting through wave action, particularly as the soil loses compaction because of worm activity. Maintaining the lowest possible water levels during rice crop establishment has helped farmers to minimize these effects.     

Characterization of fungi (Fusarium and Rhizoctonia) and oomycetes (Phytophthora and Pythium) associated with apple orchards in South Africa

Several species of fungi and oomycetes including Fusarium, Rhizoctonia, Phytophthora and Pythium have been reported as root pathogens of apple where they contribute to a phenomenon known as apple replant disease. In South Africa, little is known about specific species in these genera and their pathogenicity toward apple. Therefore, these aspects were investigated along with the development and optimization of qPCR tests for detection and quantification of the most virulent oomycete species. In eight investigated orchards, the oomycete Phythophthora cactorum was widely distributed, while nine Pythium species were differentially distributed among the orchards. Pythium irregulare was the most widely distributed and the most virulent species along with P. sylvaticum, P. vexans and Ph. cactorum. Seven binucleate Rhizoctonia anastomosis groups (AGs) were also differentially distributed among the orchards, with the majority appearing to be non-pathogenic while certain AG-I and AG-F isolates exhibited low virulence on apple. In the genus Fusarium, F. oxysporum was widely distributed, but isolates were non-pathogenic. Fusarium solani and F. avenaceum were less frequently encountered, with only some isolates having low virulence. qPCR data obtained from seedling roots inoculated with the most virulent Pythium species (P. irregulare, P. sylvaticum and P. vexans) and the genus Phytophthora were not always reproducible between trials, or isolates of the same species. In general, seedling growth inhibition was associated with the presence of a low amount of pathogen DNA (±40 fg μl⁻¹ to 2 pg μl⁻¹) in roots. Pythium irregulare, although having the lowest DNA concentrations in roots, was the only species for which a significant negative correlation was found between seedling weight and pathogen DNA concentration.     

Components of early competition between upland rice (Oryza sativa L.) and Brachiaria brizantha (Hochst. ex A.Rich) Stapf.

Abstract

In Latin America early weed competition is a constraint upon upland rice production, leading to intensive herbicide use. Breeding for rice competitiveness may help reduce the need for herbicides. Two consecutive screenhouse pot experiments were conducted with 13 upland cultivars grown alone or in mixtures with Brachana brizantha to identify plant characteristics for early rice competitiveness which could serve as selection criteria. Rice growth reduction from B. brizantha competition was evident in all cultivars by 30 d.a.e., and the traits most related to cultivar competitiveness were: total above‐ground plant biomass > leaf weight > stem weight > leaf area. Plant height and tillering were least affected by competition. Rice and B. brizantha partially avoided competition since the competing canopies did not fully intercept the incoming light. Traits measured in monoculture did not correlate well with the competition‐induced growth reductions. In breeding for competitiveness, selection according to the above traits should be conducted on rice growing in competition, rather than in monoculture. A measure of plant biomass or leaf area at 30 d.a.e. appears to provide the most convenient estimates of early interference.     

Leaf spot of <Emphasis Type="Italic">Aster savatieri</Emphasis> (Makino) Kitam. caused by <Emphasis Type="Italic">Septoria chrysanthemella</Emphasis> Saccardo

 In May 1998 leaf spot caused by Septoria chrysanthemella was found on Aster savatieri in Kanagawa Prefecture, Japan. This is the first report of leaf spot on A. savatieri caused by S. chrysanthemella. Received: September 13, 2002 / Accepted: October 18, 2002 Acknowledgments The authors thank Dr. T. Kobayashi, formerly of Tokyo University of Agriculture, for his advice on identifying the fungus.     

Induced systemic resistance of Chinese cabbage to bacterial leaf spot and <Emphasis Type="Italic">Alternaria </Emphasis>leaf spot by the root endophytic fungus, <Emphasis Type="Italic">Heteroconium chaetospira</Emphasis>

 The root endophytic fungus Heteroconium chaetospira isolate OGR-3 was tested for its ability to induce systemic resistance in Chinese cabbage against bacterial leaf spot caused by Pseudomonas syringae pv. maculicola and Alternaria leaf spot caused by Alternaria brassicae of the foliar diseases. Chinese cabbage seedlings planted in soil infested with an isolate of H. chaetospira were incubated in a growth chamber for 32 days. The first to fourth true leaves of the seedlings were challenge-inoculated with P. syringae pv. maculicola or A. brassicae. Chinese cabbage planted in soil infested with H. chaetospira showed significant decreases in the number of lesions of bacterial leaf spot or Alternaria leaf spot when compared to the control plants not treated with H. chaetospira. The results indicated that colonization of roots by H. chaetospira could induce systemic resistance in Chinese cabbage and reduce the incidence of bacterial leaf spot and Alternaria leaf spot. Received: April 24, 2002 / Accepted: August 9, 2002     

Rice nematode problems in Nigeria: Their occurrence,distribution and pathogenesis

Abstract

In a survey of plant parasitic nematodes associated with or affecting rice throughout Nigeria, some important nematode pests, especially the white tip disease nematode, Aphelenchoides besseyi and the rice root nematodes Hirschmanniella spinicaudata and H. oryzae were identified from seed, soil and root samples from swamp rice fields respectively. The sugarcane cyst nematode, Heterodera sacchari occurred in swamp rice fields only around the major sugarcane estates in Nigeria. The root‐knot Meloidogyne incognita and the root lesion nematode, Pratylenchus brachyurus were also encountered in upland (rainfed) rice fields. The white tip disease nematode, A. besseyi occurred at low levels in soils and rice seeds throughout the country. High population levels of H. spinicaudata and H. oryzae were encountered especially in areas where monoculture of rice is practised. General chlorosis, poor tillering and significantly reduced yield have been observed due to H. spinicaudata. Rice plants attacked by H. sacchari also showed intense chlorosis, delayed and reduced tillering and reduced grain yield. The roots of attacked plants were twiggy, very necrotic and blackened. The root‐knot M. incognita and the root lesion nematodes P. brachyurus have both been observed to reduce rice yields. Rice cultivars screened for reactions to the nematodes showed varying degrees of susceptibilities. Some varieties were however resistant to the root‐knot nematode, M. incognita.     

Susceptibility to imazamox in Italian weedy rice populations and Clearfield® rice varieties

The introduction of imidazolinone‐tolerant rice varieties has made selective Oryza sativa (weedy rice) control possible. We hypothesised that Italian weedy rice populations have variable degrees of susceptibility to imazamox prior to imidazolinone‐tolerant variety introduction. To this end, 149 Italian weedy rice populations collected from fields never before cultivated with imidazolinone‐tolerant varieties were tested in a glasshouse‐based, whole‐plant response screening study. Imazamox was applied to all populations post‐emergence at a rate of 70 g a.i. ha^?1, resulting in 70–90% shoot biomass reduction in the majority of cases. The results prompted a second study of the seedling dose response of four weedy rice populations from the initial study group. Three imidazolinone‐tolerant and one conventional rice variety were also included. The seedling roots were cut six days after germination and exposed to different concentrations of imazamox. The root regrowth associated with each concentration‐exposure was then measured. Imazamox concentrations to inhibit weedy rice root growth by 50% varied by about two orders of magnitude, or between 0.0018 and 0.12 mm . Even with this result, imidazolinone‐tolerant varieties were at least 31.8 times less susceptible than weedy rice populations, suggesting that Italian weedy rice populations were not tolerant to imazamox before introduction of these varieties.