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Abstract: Coniothyrium minitans is a mycoparasite and a promising biocontrol agent of Sclerotinia sclerotiorum, the causal of Sclerotinia stem rot of oilseed rape. In order to assess whether soil application of C. minitans can be combined with fertilizer application during cultivation of oilseed rape, a study was conducted to determine the compatibility of C. minitans with commercial N–P–K compound fertilizer. Results showed that the compound fertilizer at concentrations of 0.1–10% inhibited conidial germination and mycelia growth of C. minitans in a dosage–response manner in vitro. Simultaneous application of C. minitans and the compound fertilizer at various concentrations significantly (P <0.05) reduced the number of apothecia produced by sclerotia of S. sclerotiorum in both pot and field plot experiments. No negative effects of the compound fertilizer on the ability of C. minitans to infect sclerotia of S. sclerotiorum in vitro or to suppress carpogenic germination of sclerotia of S. sclerotiorum were observed. It is demonstrated that C. minitans is compatible with the compound fertilizer when applied at planting of oilseed rape. Application of C. minitans together with the compound fertilizer would be advantageous in saving labor cost, thus increasing production efficiency of oilseed rape. [Copyright &y& Elsevier]

Abstract: Bacterial fruit blotch (BFB) caused by Acidovorax avenae subsp. citrulli (Aac) is a serious disease of hami melon (Cucumis melo var. saccharinus) in Northern China. A study was conducted to screen plant epiphytic yeasts for use as biocontrol agents of BFB. Results showed that 24 out of 463 yeast strains isolated from leaves or flowers of plants collected from three provinces in China were antibiotic against Aac on agar medium and eight antagonistic yeast strains including strain 0732-1 formed inhibition zones larger than 18mm in diameter. Spray application of strain 0732-1 isolated from watermelon grown in Xinjiang was effective in reducing incidence and severity of disease caused by Aac on leaves of hami melon. Treatment of hami melon seeds with cell-free cultural filtrates of the yeast strain 0732-1 resulted in a significant reduction in severity of seedling blight caused by seedborne Aac, and the efficacy was not significantly different (P >0.05) from that of chemical seed treatments including streptomycin sulfate (0.1%, w/v) and hydrochloric acid (2%, v/v). Based on morphological and physiological characteristics and analysis of the DNA sequence of the internal transcribed spacer of ribosomal DNA, the yeast strain 0732-1 was identified as Pichia anomala Kurtzman. This study suggests that the yeast strain 0732-1 is an agent with potential for biocontrol of BFB of hami melon caused by Aac. [Copyright &y& Elsevier]

Abstract: Headspace volatile substances (VS) produced by Streptomyces platensis F-1 were preliminarily identified using GC–MS. The effects of VS released by S. platensis F-1 on the control of leaf blight/seedling blight of rice caused by Rhizoctonia solani, leaf blight of oilseed rape caused by Sclerotinia sclerotiorum and fruit rot of strawberry caused by Botrytis cinerea, as well as on the growth of these three pathogenic fungi, were investigated. Results showed that sixteen volatile compounds were tentatively identified in 1-week-old cultures of S. platensis F-1 grown on autoclaved wheat seeds. They could be chemically grouped into alcohols, esters, acids, alkanes, ketones and alkenes. The most abundant composition in volatiles of S. platensis F-1 is geosmin, an earthy-muddy–smelling compound. Two antifungal compounds, phenylethyl alcohol and (+)-epi-bicyclesesquiphellandrene, were detected in the volatile profile of S. platensis F-1. Consistent fumigation of healthy tissues of rice, oilseed rape and strawberry to VS of S. platensis effectively reduced the incidence and/or the severity of leaf blight/seedling blight of rice (R. solani), leaf blight of oilseed rape (S. sclerotiorum) and fruit rot of strawberry (B. cinerea). A significant (P <0.05) suppression of the mycelial growth of R. solani, S. sclerotiorum and B. cinerea by the VS of S. platensis was observed. The potential of using VS of S. platensis F-1 as a biofumigant to control plant fungal diseases is discussed. [Copyright &y& Elsevier]

Abstract: Degradation of oxalic acid (OA) by the mycoparasitic fungus Coniothyrium minitans (Cm), and the effects of OA degradation on production and activity of β-1,3-glucanase of Cm were investigated. Results showed that OA was degraded by 86–92% by Cm grown at 20°C for 15 days in modified potato dextrose broth (mPDB) containing OA at 0.8–28mM. Ambient pH values of these cultures were increased from 3.0–4.8 to 8.3–8.6 due to the degradation of OA and/or accumulation of ammonia by Cm. In dual cultures of Cm and Sclerotinia sclerotiorum (Ss) on potato dextrose agar amended with 0.1% (w/v) bromophenol blue (a pH indicator), spread of Cm onto colonies of Ss was correlated with the elevation of the ambient pH from 2.9 to 6.6, implying that OA degradation by Cm may occur during the interaction between the two fungi. Increase of the ambient pH was also observed on flower petals of oilseed rape inoculated with Cm (5.4×104 conidia/petal) and Ss (5.4×103 ascospores/petal) when they were incubated on water agar amended with 0.1% (w/v) bromophenol blue for 6 days, compared to those inoculated with Ss alone. Meanwhile, leaf blight incidence of oilseed rape caused by flower petals inoculated with Cm (1.5×103–5 conidia/petal) and Ss (4.2×103 ascospores/petal) was significantly lower (P <0.05) than that caused by flower petals inoculated with Ss alone. Results also showed that OA degradation was correlated with the enhanced production of β-1,3-glucanase by Cm and the stimulated activity of this enzyme. In cultures of Cm grown in mPDB amended with OA (16mM) for 9, 12 and 15 days, OA was degraded by 90%, 91% and 94%, respectively, and the yield of β-1,3-glucanase produced by Cm was increased by 61%, 23% and 52%, respectively, compared to that in Cm cultures grown in mPDB alone. Results in buffered PDB media showed that the yield of β-1,3-glucanase produced by Cm was positively correlated (R =0.9439, P <0.01) with the ambient pH ranging from 3 to 8, implying that the increase in ambient pH caused by OA degradation may be responsible for the enhanced production of β-1,3-glucanase by Cm in OA-containing media. Inhibition by OA of the activity of β-1,3-glucanase produced by Cm was observed and the degree of inhibition was positively correlated to the concentration of OA ranging from 4 to 32mM, which inversely conditioned the ambient pHs to a range from 1.5 to 3.5. In contrast, the activity of β-1,3-glucanase produced by Cm was not affected by sodium oxalate ranging from 2 to 32mM, which conditioned the ambient pH to a range from 5.9 to 6.1. The optimum ambient pH for the enzymatic reaction of β-1,3-glucanase of Cm ranged from 4.0 to 6.0. This study suggests that degradation of OA by Cm can nullify the pH effect conditioned by OA and may improve mycoparasitism of Cm on Ss by stimulating production of β-1,3-glucanase by Cm and/or the activity of this enzyme. Degradation of OA by Cm might also be a mechanism by which Cm can protect plants from infection by Ss. [Copyright &y& Elsevier]

Abstract: Antifungal activity (AA) of the mycoparasite Coniothyrium minitans (CM) against Sclerotinia sclerotiorum (SS), designated as AACM–SS, was detected by measuring the effect of cell-free cultural filtrates of CM on inhibition of growth (IG) of SS on potato dextrose agar. Using this bioassay method, effects of ambient pH and nutritional factors on AACM–SS in CM cultures grown in various liquid media were investigated. Results showed that production of AACM–SS by CM was closely related to ambient pH. After incubation for 9–15 days, AACM–SS was high in modified Czapek–Dox broth (MCD) with IG values being 78–90%, but low in potato dextrose broth (PDB) with IG values being 9–18%. In contrast, pH value was as low as 3.3–3.6 in the MCD cultures, whereas as high as 6.0–7.2 in the PDB cultures. In the pH-buffered experiment, AACM–SS was detected in MCD cultures of CM buffered at pH 2–8 with the optimum pH being 3–5, under which AACM–SS in PDB cultures of CM was greatly enhanced with the IG values reaching up to 46–67%. Results also indicated that AACM–SS in CM cultures can be affected by nutritional factors and their effects were also related to ambient pH. In the basal medium Czapek mineral salts, most carbon and nitrogen sources investigated were favourable to CM to produce AACM–SS with the IG values being 73–91% and the final pH values being 2.7–4.1. In the other basal medium potato extracts, a few carbon and nitrogen sources were favourable for CM to produce AACM–SS with the IG values being 40–60% and the final pH values being 2.7–5.5. In the pH-buffered experiments with selected carbon and nitrogen sources, or with mycelial or sclerotial extracts of SS, AACM–SS was much higher in CM cultures buffered at pH 3 than that buffered at pH 6. Additional experiments showed that AACM–SS could be induced by the phytotoxin oxalic acid (OA), but not by sodium oxalate, further suggesting the importance of acidic pH in production of AACM–SS by CM. Meanwhile, leaf blight incidence on Chinese cabbage caused by SS was greatly reduced in the treatment of CM conidia pre-germinated in PDB plus OA (16mM) than that in the treatment of CM conidia pre-germinated in PDB alone. This study suggests that ambient pH plays an important role in regulation of the antifungal activity of CM against SS. [Copyright &y& Elsevier]

Bacterial fruit blotch (BFB) of watermelon caused by Acidovorax citrulli (Ac) is a seedborne disease. Seed treatment with bacterial disinfectants is considered as an important measure for suppression of Ac infection. This study was performed to detect the antibacterial activity of the cultural filtrate (CF) of the saprophytic fungus Aspergillus niger Y-1 against Ac. The six-day-old CF, citric acid (CA, 4 mmol/L) and oxalic acid (OA, 60 mmol/L) were determined for in vitro antibacterial activity against Ac. CF and OA were determined as seed disinfectants for suppression of seedborne infection by Ac. Results showed that production of CA and OA by A. niger in potato dextrose broth was consistently detected by HPLC. The CF, CA and OA inhibited growth of Ac and their inhibitory effect disappeared when the pH values of the three solutions was adjusted to 7.0. OA was more effective than CA in suppression of Ac . The potting experiment showed that both CF and OA applied on watermelon seeds effectively suppressed BFB incidence on seedlings. The efficacy was comparable to the seed treatment with HCl (1%, v/v). This study suggests that the CF of A. niger and OA can be used as seed disinfectants for elimination of seedborne Ac. [ABSTRACT FROM AUTHOR]

This study was conducted to optimize the cultural medium and conditions for production of antifungal substances by Streptomyces platensis 3-10 and to evaluate its efficacy in suppressing Plasmodiophora brassicae (PB), the causal agent for clubroot of oilseed rape. The optimization was done using the one-factor-at-a-time (OFAT) method, followed by orthogonal designing. The optimized medium contained starch at 3% (w/v), peptone at 0.75%, yeast extract at 0.025%, soybean meal at 1% (w/v) and minor/trace elements (K + , Mg 2+ , Mn 2+ , Zn 2+ ), initial pH 6.5. After incubation in this medium at 28 °C/150 rpm for 72 h, isolate 3-10 showed high antifungal activity. The cultural filtrates (CF 3-10 ) produced an average inhibition-zone diameter of 50 mm against Aspergillus niger . The live spores and metabolites of 3-10 in CF 3-10 or its crude extracts (CE 3-10 ) suppressed PB infection of oilseed rape roots by 3–83%, compared to the treatment with PB alone. The efficacy was positively correlated (correlation co-efficiency: 0.9526–0.9873, P < 0.05) to the applied dose of each active component. Germination of PB resting spores was inhibited by 75% and 80% by CE 3-10 (1000 μg/mL, w/v) and CF 3-10 (5%, v/v), respectively. These findings suggest that S. platensis 3-10 is a promising biocontrol agent against PB. [ABSTRACT FROM AUTHOR]

Seventy-two Trichoderma isolates were obtained from Hubei Province of China and identified to 11 species based on rDNA-ITS. The isolates were initially tested for invasive growth on Botrytis cinerea colony in dual cultures with B. cinerea on potato dextrose agar at 20 °C. The resulting 28 isolates were further determined for biocontrol potential using three criteria: invasive growth on Botrytis cinerea colonies at 15 °C, antifungal activity and promotion of tomato seed germination. Performance of the Trichoderma isolates was scored and the total scores were used to select isolates with the highest biocontrol potential. Four selected isolates ( T. harzianum : T-21 and T-68; T. koningiopsis : T-35 and T-51) were evaluated for growth-promoting and resistance-inducing effects on tomato seedlings, and for sporulation-suppressing effect on B. cinerea on dead tomato leaves. All the four isolates significantly ( P < 0.05) promoted tomato growth possibly through producing gibberellins and indole-3-acetic acid, and induced systemic resistance against B. cinerea . Isolates T-21, T-51 and T-68 suppressed B. cinerea sporulation by 62.8, 66.1 and 63.7%, respectively, compared to the control treatment. This study suggests that the multiple criteria-based procedure is rational for screening of Trichoderma isolates against B. cinerea , and T. koningiopsis T-51 is a promising biocontrol agent. [ABSTRACT FROM AUTHOR]

Highlights: [•] Oilseed rape harbors diversified endophytic fungi. [•] Four antagonists effectively suppressed S. sclerotiorum. [•] VOCs of F. oxysporum highly suppressed B. cinerea. [•] Three endophytic fungi promoted growth of oilseed rape. [Copyright &y& Elsevier]

Abstract: Volatile substances produced by Streptomyces globisporus JK-1 grown on autoclaved wheat seed inhibited Botrytis cinerea growth and development both on media and in inoculated tomato fruit. The volatiles suppressed mycelial growth of various plant pathogens in vitro, especially that of B. cinerea and Sclerotinia sclerotiorum. Conidial germination and sporulation of B. cinerea were also suppressed. Disease incidence and severity on wound-inoculated tomato fruit (Lycopersicon esculentum) were inhibited when fumigated with 120g wheat seed culture of S. globisporus JK-1 per liter of airspace in treatment containers. Suppression of the infection process of B. cinerea on tomato fruit was observed via scanning microscopy, showing inhibition of conidial germination and of appressorial formation on tomato fruit, as well as abnormal morphology of appressoria and conidia. The viability of the conidia obtained from the volatile-treated and non-treated disease lesions was tested with the vital stains fluorescein diacetate (FDA) and propidium iodide (PI). Conidia fumigated with 30, 60 or 120g/L wheat seed culture of S. globisporus JK-1 at 20°C for 6days showed 46.0%, 69.8%, or 80.9% reduction in viability, respectively. Transmission electron microscopy of fumigated and untreated B. cinerea showed excessive vesiculation or thickened cell walls in exposed conidia and increased vesiculation or strong retraction of plasma membrane in exposed hyphae. These results provide a better understanding of the mode of action of volatiles from JK-1 on B. cinerea. The inhibition growth of B. cinerea both in vitro and in vivo showed that volatiles from S. globisporus JK-1 have the potential for control of postharvest grey mold of tomato fruits through fumigant action. [Copyright &y& Elsevier]

Abstract: An isolate, JK-1, was obtained as a contaminant from a fungal culture, and identified as Streptomyces globisporus. In dual cultures on solid media, this strain suppressed mycelial growth of numerous plant pathogenic fungi, especially that of Magnaporthe oryzae, Bipolaris maydis and Cryphonectria parasitica. Culture filtrates of JK-1 inhibited mycelial growth of M. oryzae, and histological investigations showed that conidial germination and appressorial formation of M. oryzae were suppressed on detached rice leaves. When applied at 15μl per 5-cm-long detached leaf, washed cells of JK-1 at 108 CFU/ml could suppress disease incidence of rice blast caused by M. oryzae co-inoculated at 5×105 spores/ml, but disease incidence was not reduced when washed cells of JK-1 were used at 106 or 107 CFU/ml. Applying the culture filtrate on rice seedlings in the greenhouse at 2h post inoculation (HPI) with M. oryzae showed 88.3% disease reduction of rice blast, while culture filtrate application before pathogen inoculation showed even higher rates of disease reduction. Suppression of the infection process of M. oryzae on detached rice leaves was observed by light and scanning electron microscopy, showing inhibited conidial germination and reduced appressorial formation on rice leaves sprayed with culture filtrate before 3 HPI. Application of JK-1 as cells or culture filtrates to plant tissues did not cause any noticeable negative effects. These results indicate that the antifungal substance(s) in the culture filtrate of S. globisporus JK-1 can exhibit an inhibitory effect on M. oryzae and a suppressive effect on rice blast disease. [Copyright &y& Elsevier]

• Hypovirulent isolate QT5-19 of B. cinerea has strong competitiveness. • QT5-19 suppressed infection by virulent isolates of B. cinerea by competition. • QT5-19 produced volatiles with high antifungal activity against B. cinerea. • Improved volatile production is a mechanism for competitiveness of B. cinerea. This study determined the competitive saprophytic ability (CSA) of the hypovirulent isolate QT5-19 of Botrytis cinerea on potato dextrose agar (PDA) at 20 °C. Meanwhile, importance of the QT5-19 CSA in biocontrol against virulent isolates of B. cinerea and Sclerotinia sclerotiorum (a close relative of B. cinerea), and the mechanisms for the QT5-19 CSA were elucidated. The results showed that QT5-19 had higher CSA than the virulent isolates 08168 (B. cinerea) and EP-1PNA367 (S. sclerotiorum) on potato dextrose agar. The hyphal fragments of QT5-19 exhibited an effective suppression against infection of leaves of oilseed rape by the hyphae of the two virulent fungal pathogens through competition. QT5-19 was detected to be able to produce volatile organic compounds (VOCs) with antifungal activity against B. cinerea and S. sclerotiorum , whereas the VOCs from the virulent isolate RoseBc-3 of B. cinerea had no detectable antifungal activity. The QT5-19 VOCs effectively reduced disease severity of strawberry fruit rot caused by two virulent fungal pathogens. These results suggest that QT5-19 may achieve success of competition through extensive mycelial growth and production of antifungal volatiles. This study provided an example of using hypovirulent isolates to control virulent isolates of B. cinerea and S. sclerotiorum. [ABSTRACT FROM AUTHOR]