Helminthosporium Sacchari Research Articles

Appraisement of chemotherapy, plant defense activators, and genetic resistance against eyespot disease in sugarcane

Sugarcane (Saccharum officinarum) is a crucial agro-industrial crop that faces numerous biotic and abiotic challenges. One of the most severe biotic stresses affecting sugarcane is eyespot disease, caused by Bipolaris sacchari. This study focused on managing eyespot disease using resistant varieties, chemicals, and plant defense activators. Disease samples were collected from Lalian, Chiniot, and Faisalabad, and the pathogen was isolated and identified from the infected leaves. In the screening experiment, 10 sugarcane varieties were evaluated. CPF-246 exhibited high resistance, while CPF-247 and CPF-77400 showed resistance to eyespot disease. The poisoned food technique was used to assess fungicides at different concentrations (50, 100, and 150 ppm) under in vitro conditions. Tilt (Propiconazole) exhibited the maximum growth inhibition (4.645 mm) under these conditions. Plant activators were tested at different concentrations (0.25%, 0.5%, and 0.75%) in greenhouse conditions. Salicylic acid resulted in the minimum disease incidence (13.236%, 7.483%, and 6.473%) after 15, 30, and 45 days, respectively. Under field conditions, the combination of Salicylic acid and Tilt (12.395%) showed the lowest disease incidence. By employing these newly investigated management strategies, the farming community can effectively combat the significant threat posed by eyespot disease to the sugarcane industry.

Fungicidal spectrum and biological properties of a new fungicide, pyriofenone.

Pyriofenone is a new fungicide developed by Ishihara Sangyo Kaisha, Ltd. To determine the fungicidal spectrum of pyriofenone, in vivo pot tests and in vitro mycelial growth-inhibition tests were conducted. Pyriofenone showed excellent activity against wheat and cucumber powdery mildew and moderate efficacy against rice blast in the pot tests. In the mycelial growth-inhibition tests, most fungi were not affected by pyriofenone except for Botrytis cinerea, Helminthosporium sacchari, Pseudocercosporella herpotrichoides, Pyricularia oryzae, Rosellinia necatrix, and Verticillium dahliae. The fungicidal properties of pyriofenone on powdery mildew in cucumber and wheat were evaluated precisely. Pyriofenone exhibited excellent preventive and residual activities. It had high rainfastness in the cucumber leaves against powdery mildew. Pyriofenone also showed inhibitory activity on lesion development upon application until 2 days after inoculation, and the lesion expansion and sporulation of the cucumber powdery mildew fungus were effectively controlled. Furthermore, pyriofenone showed translaminar and vapor activities.

Evaluation of Fungicides and Plant Extracts Against Pathogenic Fungi Associated With Basella Spp.

Five fungicides viz., CM 75, Dithane M 45, Knowin 50 WP, Nativo 75 WG and Rovral 50 WP were evaluated against Colletotrichum lindemuthianum, Drechslera sacchari and Fusarium semitectum following poisoned food technique. Out of five fungicides complete inhibition of radial growth of C. lindemuthianum was observed in Nativo 75 WG at 100 ppm. On the other hand, the complete inhibition of the growth of D. sacchari was observed with Rovral 50 WP at 400 ppm, whereas Nativo 50 WP showed complete growth inhibition at 500 ppm. CM 75 WP, Knowin 50 WP and Nativo 75 WG showed complete growth inhibition of F. semitectum at 100 ppm. Five different plant leaf extracts viz., Azadirachta indica A. Juss., Heliotropium indicum L., Lippia alba L., Michelia champaca L. and Thuja occidentalis L. were tested against the test pathogens. Of the five plant leaf extracts, Lippia alba showed the highest growth inhibition in C. lindemuthianum, D. sacchari and F. semitectum at 20% concentration.
 Asiat. Soc. Bangladesh, Sci. 46(1): 81-89, June 2020

Prevalence and Pathogenic Potentiality of Fungi Associated With Leaf Spot of Basella Alba and B. Rubra

Two major species of Indian spinach, namely Basella alba L. and B. rubra L. were selected to find out associated mycoflora and their pathogenic potentiality. Ten fungi viz., Alternaria alternata, Aspergillus flavus, A. fumigatus, A. niger, Colletotrichum dematium, C. lindemuthianum, Curvularia lunata, Drechslera sacchari, Fusarium semitectum and Penicillium sp. were found to be associated with the selected leaves of Basella spp. Among the isolated fungi C. lindemuthianum, D. sacchari and F. semitectum were found to be pathogenic to Basella spp. Association of C. lindemuthianum and D. sacchari with Basella spp. are recorded first time from Bangladesh.
 Dhaka Univ. J. Biol. Sci. 29(2): 183-189, 2020 (July)

Exploitation of new endophytic bacteria and their ability to promote sugarcane growth and nitrogen nutrition.

Few studies have evaluated endophytic bacteria in relation to plant growth promotion, nitrogen uptake and biological control. The aim of this study was to molecularly and physiologically characterize thirteen endophytic bacteria strains, evaluate their biological control properties and their ability to promote plant growth and plant N nutrition. All the strains produced indole acetic acid and promoted increase of plant biomass, N accumulative amount and N-use efficiency index. None of the strains carries the nifH gene. Four strains stimulated plant nitrate reductase activity, four solubilized phosphate, nine produced siderophores and none produced HCN. Seven strains inhibited Bipolaris sacchari growth and one was antagonistic to Ceratocystis paradoxa. The pathogens were inhibited by the production of diffusible and volatile metabolites by the bacterial strains. Moreover, this is the first study to demonstrate the effect of Delftia acidovorans on sugarcane plant growth, nitrogen metabolism improvement and antagonism to B. sacchari. The most efficient strains in promoting plant growth and exhibiting antagonistic activities towards fungal pathogens were Herbaspirillum frinsingense (IAC-BECa-152) and three Pantoea dispersa strains (IAC-BECa-128, IAC-BECa-129, and IAC-BECa-132). These bacteria show potential to be used as inoculants for sustainable agricultural management, mainly at the seedling production phase.

Registration of ‘CP 04‐1935’ Sugarcane

‘CP 04‐1935’ (Reg. No. CV‐154, PI 667660) sugarcane (a complex hybrid of Saccharum spp.) was developed through cooperative research conducted by the USDA‐ARS, the University of Florida, and the Florida Sugar Cane League and was released to growers in Florida on 20 Sept. 2011. CP 04‐1935 was selected from the cross CP 94‐2059/CP 84‐1322 made at Canal Point, FL on 8 Dec. 1998. In the final stage of selection, CP 04‐1935 was tested for yield performance at two sand‐soil locations along with 15 other genotypes across three crop years and for freeze tolerance in northern Florida for two crop years. CP 04‐1935 produced an 11% higher cane yield, a 3.5% higher sucrose content, and a 14.5% higher sucrose yield than the reference sand cultivar CP 78‐1628. CP 04‐1935 is resistant to brown rust (caused by Puccinia melanocephala H. & P. Sydow), orange rust (caused by Puccinia kuehnii E.J. Butler), mosaic (caused by Sugarcane mosaic virus strain E), to smut (caused by Ustilago scitaminea H. & P. Sydow), and to eyespot [caused by the Bipolaris sacchari (E.J. Butler) Shoemaker]; it is moderately resistant to leaf scald (caused by Xanthomonas albilineans Ashby, Dowson), and to ratoon stunt (caused by Leifsonia xyli subsp. xyli Evtushenko et al.), but it is susceptible to Sugarcane yellow leaf virus. CP 04‐1935 performed significantly worse under freeze conditions than CP 89‐2143, the Florida industry standard for acceptable freeze tolerance. With its profitability predicted to be 19% higher than that of CP 78‐1628, combined with a good disease profile, CP 04‐1935 was recommended for planting on sand soils in Florida.

First report of leaf spots on banana in Brazil caused byBipolaris sacchari

A new leaf spot disease caused by Bipolaris sacchari is reported for the first time on banana (Musa × paradisiaca)

Lower temperature during the dark cycle affects disease development on Lygodium microphyllum (Old World climbing fern) by Bipolaris sacchari

Growth chamber experiments were conducted to examine environmental parameters affecting disease development by the indigenous pathogen Bipolaris sacchari isolate LJB-1L on the invasive weed Lygodium microphyllum (Old World climbing fern). An initial experiment examined three different temperature regimes (20/15, 25/20, 30/25°C; light/dark cycle) and three different periods of leaf wetness (12, 24, 48h). Disease incidence was significantly reduced by incubation of plants at the highest temperature regime (30/25°C) and with the shortest period of leaf wetness (12h). Subsequent experiments concentrated on the effect of temperature, as determination of an optimal temperature for disease development might determine the best time of year for field inoculations with this fungus. When the light cycle temperature was kept constant at 30°C, a dark cycle temperature of 15°C was more conducive for disease development than 25°C based on disease incidence and area under the disease progress curve (AUDPC). A qualitative examination of conidia germination and appressorial formation in vivo indicated that these initial steps toward infection were probably not influenced by the dark cycle temperature. In vitro, temperature (range of 15–35°C) did not significantly affect conidia germination, but hyphal growth was significantly affected, with optimal growth between 25 and 30°C.

First report of Bipolaris sacchari causing wheat stem‐base disease in Iran

During May and June 2003, wheat plants in several fields in Hamada, Iran showed symptoms of root rot and crown necrosis. Stem-base and root samples were submerged in 0·5% sodium hypochlorite for 3–5 min, rinsed with sterile distilled water, transferred to Petri dishes containing potato dextrose agar (PDA), and incubated at 22–25°C for 7 days. Bipolaris spp. was the most prevalent fungus isolated. Based on the host origin, conidia and conidiophore morphology, number of pseudosepta and hilum characteristics, the pathogen was identified as Bipolaris sacchari (Sivanesan, 1987). Inoculum for pathogenicity tests was produced by seeding 200 g sterile wheat stubble in an Erlenmeyer flask with four 5-mm-diameter mycelial plugs from 5-day-old fungal colonies grown on PDA, and incubated at room temperature (20°C) for 3 weeks. The inoculated wheat straw was incorporated into sterile soil (1% w/w), then transferred to five 25-cm pots and planted with eight wheat seeds per pot. Control plants were grown in soil containing 1% sterile wheat straw. The treatments were replicated five times. Plants were grown in a glasshouse for 75 days at 24°C, then scored for disease (Singleton et al., 1992). Symptoms were similar to those observed in the field, and no disease was found in the control plants. The fungus was reisolated from the infected parts. In a separate study using wheat cvs Alvand and Roushan, the pathogen caused a significant reduction in seed germination and subsequent plant height. This is the first report of root and stem rot disease of wheat caused by B. sacchari in Iran. Further field studies are needed on the ecology and pathogenicity of B. sacchari to formulate steps for controlling root and stem rot of wheat. Financial support for this work by the Research Council of Bu-Ali Sina University, Hamadan, Iran, is gratefully acknowledged.

Cellular target of helminthosporium sacchari Toxin in sugarcane tissue

Spindles from sugarcane (Saccharum spp. hybrid) tissue were challenged using labeledHelminthosporium (Bipolaris) sacchari toxin to determine the general effect on different organelles and structures. Toxin labeling was characterized by biochemical and physiological behaviour of the fungal pathogen (dry weight, pH, refractive index, invertase and β -galactosidase activity and the biological activity of the toxin), allowing to improve the metabolic uptake of a labeled structural precursor of the toxin. A more efficient toxin labeling than previously reported was obtained. The labeled toxin was used as a tracer for determination of interactions at mitochondrial level. Binding activity reached saturation in time, confirming respiratory and ultrastructural analysis of the toxin effect.

First Report of Bipolaris sacchari Causing Leaf Spot on Lygodium japonicum and L. microphyllum in Florida.

Lygodium japonicum (Thunb.) Sw. (Japanese climbing fern) and L. microphyllum (Cav.) R.Br. (Old World climbing fern) are invasive, noxious weeds in Florida. During 2001, L. japonicum sporelings were collected from natural sites in Hamilton, Highland, and Madison counties and transported to Broward County (Fort Lauderdale) for research use. During February 2002, leaf spots were observed on pinnules (leaflets) of these containerized plants growing in a shadehouse. A fungus with Bipolaris-like spores was isolated from affected pinnules and purified and stored for future evaluation. In early 2005, the fungus was grown on 1.5% water agar, with sterile, wheat straw pieces embedded in the agar surface, at 26°C and a 12-h photoperiod using cool-white fluorescent and Verilum full-spectrum bulbs. Conidia were 80.5 ± 14.5 μm (range 53.2 to 123.4 μm) × 15.1 ± 1.6 μm (range 12.1 to 19.4 μm), pale brown, slightly curved, narrowly ellipsoid, without a protuberant hilum, distoseptate (8 ± 1, range 6 to 10), and germinated from both polar cells. Conidiophores were septate and smooth. Conidiogenous nodes were smooth. On the basis of these characteristics, the fungus was identified as Bipolaris sacchari (E. Butler) Shoem. (1,2). Pathogenicity toward L. japonicum and L. microphyllum was determined using conidia produced on potato dextrose agar at 26°C and a 12-h photoperiod. A 1 × 106 conidia ml-1 suspension was sprayed until runoff on healthy plants grown in 450-ml containers. Control plants were sprayed with sterile water. There were four replicate plants per treatment. Plants were covered with plastic bags to maintain high humidity and placed in a growth chamber with a 12-h photoperiod at 28°C (light cycle) and 22°C (dark cycle). Bags were removed after 72 h, and small (1 to 2 mm), water-soaked spots were evident throughout the plant canopy on both Lygodium spp. Plants were incubated for three more weeks under the same photoperiod and temperatures with ˜70% relative humidity (light cycle) and ˜45% relative humidity (dark cycle), and then evaluated for disease. At least 50% of L. microphyllum pinnules and 25% of L. japonicum pinnules on each inoculated plant had small (1 to 2 mm), brown leaf spots or larger (approximately 5 mm) necrotic spots. B. sacchari was reisolated from both types of spots from both Lygodium spp.; there was no evidence of fungal sporulation on the plants. No symptoms were apparent on control plants. To our knowledge, this is the first report of B. sacchari on a non-Poaceae host in Florida.

Identification of sugarcane genes induced in disease-resistant somaclones upon inoculation with Ustilago scitaminea or Bipolaris sacchari

To understand the molecular basis of a specific plant-pathogen interaction, it is important to identify plant genes that respond to the pathogen attack. Amplified fragment length polymorphism (AFLP) analysis of cDNA was used to identify sugarcane genes differentially expressed in disease-resistant but not in susceptible sugarcane somaclones in response to inoculation with either Ustilago scitaminea or Bipolaris sacchari (also known as Helminthosporium sacchari or Drechslera sacchari), causal agents of smut and eyespot respectively. In total 62 differentially regulated genes were identified, of which 10 were down-regulated and 52 were induced. Of these 52, 19 transcript derived fragments showed homology to known plant gene sequences, most of them related to defense or signaling. The total set of differentially expressed sugarcane genes can be an important resource for further studies aimed at understanding sugarcane pathogen defense.

Evaluation of Fungal Pathogens as Biological Control Agents for Cogongrass (Imperata cylindrica)1

Based on field surveys and evaluations in the greenhouse, two fungal pathogens,Bipolaris sacchari and Drechslera gigantea, were identified as promising biological control agents for cogon- grass. In greenhouse trials, the application of spore suspensions of these fungi containing 10 5 spores/ ml in a 1% aqueous gelatin solution to cogongrass plants and their incubation in a dew chamber for 24 h resulted in disease symptoms that ranged from discrete lesions to complete blighting of leaves. Disease severity (DS), based on a rating scale for southern corn leaf blight with 50% as the maximum DS rating, ranged from 42 to 49%. In greenhouse experiments, the application of spores formulated in an oil emulsion composed of 4% horticultural oil, 10% light mineral oil, and 86% water resulted in higher levels of foliar blight with no dew exposure or shorter periods of dew exposure (4, 8, or 12 h) as compared with the application of spores formulated in 1% gelatin. Field trials demonstrated that under natural conditions, the application of a spore and an oil emulsion mixture containing 10 5 spores/ml of either fungus could cause foliar injury from disease and phytotoxic damage from the oil emulsion. Depending on the application rate (100 or 200 ml/plot), the level of foliar injury ranged from 40 to 86% (based on a field assessment scale of 0 to 100% foliar injury) with B. sacchari as the test fungus. However, with D. gigantea as the test fungus, foliar injury ranged from 9 to 70% depending on the application volume and the oil concentration used. Although B. sacchari and D. gigantea were capable of causing foliar blight on cogongrass, the regenerative ability of the rhizomes allowed cogongrass to recover from the damage caused by these fungi. However, the level of injury caused by these fungi is sufficient to support their use as components for integrated management of cogongrass. Nomenclature: Cogongrass, Imperata cylindrica (L.) Beauv. # 3 IMPCY; corn, Zea mays L. Additional index words: Bioherbicide, biological control, Bipolaris sacchari (E.J. Butler) Shoe- maker, Drechslera gigantea (Heald & F.A. Wolf) Ito. Abbreviations: CRD, completely randomized design; DAI, days after inoculation; DS, disease se- verity; RH, relative humidity; SOE, spore and oil emulsion mixture; WAI, weeks after inoculation; WM, weed mortality.

Responses ofEchinochloaSpecies and Rice (Oryza sativa)to Indigenous Pathogenic Fungi

Six pathogenic fungal species were isolated from naturally infected Echinochloa species and evaluated as biological control agents of Echinochloa species in rice. Curvularia lunata var. aeria and Exserohilum oryzae were pathogenic to both rice and Echinochloa species and were not evaluated further. Bipolaris sacchari, Curvularia geniculata, Dactylaria dimorphospora, and Exserohilum monoceras were pathogenic only to Echinochloa species and were further compared for virulence under controlled environment conditions in the greenhouse. When provided a 24-h dew period, Exserohilum monoceras killed seedlings of all three Echinochloa species tested: E. crus-galli, E. colona, and E. glabrescens; B. sacchari resulted in 100% mortality of seedlings of E. colona and E. glabrescens; C. geniculata killed seedlings of only E. colona; and D. dimorphospora did not cause any plant death. When given a 12-h dew period, Exserohilum monoceras still killed the three Echinochloa species, whereas the other fungi did not cause plant death. Echinochloa seedlings at the one- and two-leaf stages were more susceptible to these fungi than were seedlings at the three- and four-leaf stages. B. sacchari, Exserohilum monoceras, and E. oryzae produced phytotoxins that caused 100% leaf area chlorosis and wilting of intact seedlings of the Echinochloa species placed in cell-free culture filtrates.

Characterization of Sugarcane Response toBipolaris sacchari: Inoculations and Host-Specific HS-Toxin

Twenty-seven sugarcane clones inoculated with Bipolaris sacchari, causal agent of eyespot disease, were classified as susceptible or resistant based on the presence or absence of distinct chlorotic halos surrounding lesions on leaves. Under the inoculation conditions used in this study, all lesions on resistant plants were smaller than 3 mm; the proportion of lesions larger than 3 mm on susceptible clones ranged from 0.2 to 82.2%. The total number of lesions was not related to resistance or susceptibility. All clones classified as resistant to B. sacchari were insensitive to a host-specific toxin (HS-toxin) produced by the pathogen, whereas all susceptible plants produced a water-soaked streak when 1 yl of partially purified HS-toxin was placed in a hole punched in a leaf segment [...]

Effect of Drechslera (Helminthosporium) sacchari Toxin upon the Permeability of Sugarcane Leaf and Callus Tissues

AbstractSugarcane leaves and calli from highly susceptible and resistant varieties to eyespot disease were used to evaluate the Drechslera sacchari toxin effect at different concentrations and incubation times by measuring electrolyte leakage. This expression of disease resistance was observed not only in the leaf but also in the callus. Furthermore, the growth of resistant calli MS medium supplemented with DS toxin, was higher when compared to the susceptible ones. The possibility of obtaining resistant somaclones is confirmed.

Die wirkung des toxischen metaboliten von Drechslera sacchari(Butler) Subram. & Jain auf das blattgewebe von Saccharum officinarum L. nach unterschiedlicher applikationsweise

Summary The biological activity of the toxic component from Drechslera sacchari was detected by means of visible leaf symptoms (biotest) and cytological abnormities. After applicating the culture filtrate of the pathogen respectively a partially purificated crude toxin-preparation as well as after inoculating the leaves of a susceptible sugarcane-variety qualitatively vast equivalent injurities were found. In all cases characteristic red-brown coloured stripes were induced on the leaves. On ultrastructural level the chloroplasts and mitochondriae were damaged and plasmolysis in leaf-mesophyll cells was observed. In-vitro-plants reacted in a similar way too after exposition with the host-specific toxin.

Interconversions of aglycone and host-selective toxin from Helminthosporium sacchari

Helminthosporium sacchari, a fungus that causes disease in sugarcane, produces oligosaccharide-sesquiterpene toxins (HS toxins A, B, and C) that are required for infection and disease development. Two free sesquiterpenes were isolated from mycelium but not from culture fluids of the fungus. One sesquiterpene was identified by HPLC and mass spectrometry as an aglycone of HS toxin C and could be obtained by enzymatic hydrolysis of this toxin. The other sesquiterpene appeared to be the 2-keto form of the first compound. The aglycone from toxin C hydrolysis was labelled with tritium by successive treatments with active manganese dioxide, sodium boro[ 3H]hydride, and lithium aluminium hydride. The labelled compound was fed to cultures of H. sacchari, radioactivity was incorporated into HS toxin C and into lower molecular weight homologues. The results suggest a metabolic route (aglycone → metabolite Y, → HS toxin → metabolite X) for the biosynthesis of HS toxin; metabolites X and Y are lower molecular weight homologues of the toxin.

Effect of Helminthosporium sacchari-toxin on cell membrane potential of susceptible sugarcane

The effect of the host-specific toxin produced by Helminthosporium sacchari (HS-toxin) on cell membrane potential of susceptible sugarcane was investigated. Membrane depolarization was detected at concentrations as low as 50 n m HS-toxin. The onset of membrane depolarization was observed after a lag phase of 4–10 min depending on the toxin concentration. The energy-dependent component of membrane potential (E m–p) was inhibited by the toxin. However, membrane repolarized, i.e. E m–p, was re-established after removing the toxin from the bathing solution. At low toxin concentrations E m–p was lost in the dark but was partially recovered in the light and more fully in the light with the addition of fusicoccin. The HS-toxin-induced membrane depolarization was prevented by temperatures above 30°C but subsequent lowering of temperature to 26°C resulted in membrane depolarization. Treatments with a nontoxic lower homologue of HS-toxin (lacking one galactose unit in the molecule) protected cells against HS-toxin-induced membrane depolarization. Pretreatments of tissues with galactose or raffinose did not prevent HS-toxin-induced depolarization. It is postulated that HS-toxin is causing a loss of H + gradient across the plasmalemma. However, it is unlikely that the H + pump is the site of the toxin action since in the presence of the toxin the pump can be activated by light and fusicoccin.

Selective Toxins and Analogs Produced by Helminthosporium sacchari

Helminthosporium sacchari toxin and several lower molecular weight, nontoxic analogs were isolated from culture filtrates. Three isomers of the toxin (A, B, and C), each with four galactose units, were separated by high performance liquid chromatography. Isomer C had the highest and isomer A had the lowest toxicity to H. sacchari-susceptible sugarcane; resistant clones were not affected. Each toxin isomer was partially digested with a beta-galactofuranosidase and the resulting analogs (seven from each toxin isomer) were separated by reverse phase high performance liquid chromatography and identified. Each isomer of the analogs with 3 galactose units per mole also was partially digested and the arrangement of galactose units was determined. The compound with one galactose attached to position 2 of the bicyclic sesquiterpene and with 2 galactose units attached to position 13 (analog A(1,2)) was highly toxic to some but not to all clones of H. sacchari-susceptible sugarcane. Toxin analogs protected sensitive tissue against active toxin; protective effects of the analogs differed, but at least a 10-fold excess of analog was required. Analog C(2,1) was more effective at preventing toxin C-induced electrolyte losses than was any other analog. Each of the 3-galactose analog isomers protected better than did any of the 2-galactose compounds. The 1,1 analogs did not protect as well as did the 2,0 or 0,2 analogs. Thus, the sesquiterpene isomer, the number of galactose units, and the galactose arrangement pattern determine the effectiveness of the compound in induction of electrolyte loss and in prevention of toxininduced loss from sugarcane tissues.