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Trichoderma spp. are widely used in plant disease control and growth promotion due to their high efficacy and multiple biocontrol mechanisms. Trichoderma koningiopsis T-51 is an effective biocontrol agent against gray mold disease by direct contact. However, the indirect physical contact biocontrol potential of Trichoderma spp. is not clear. In this study, the volatile organic compounds (VOCs) produced by T-51 showed high inhibitory activity against plant pathogenic fungi Botrytis cinerea and Fusarium oxysporum. The percentage of B. cinerea and F. oxysporum mycelial growth inhibition by T-51 VOCs was 73.78% and 43.68%, respectively. In both B. cinerea and F. oxysporum, conidial germination was delayed, and germ tube elongation was suppressed when exposed to T-51 VOCs, and the final conidial germination rate of B. cinerea decreased significantly after T-51 treatment. The VOCs from T-51 reduced the Botrytis fruit rot of tomato compared with that noted when using the control. Moreover, the T-51 VOCs significantly increased the size and weight of Arabidopsis thaliana seedlings. Twenty-four possible compounds, which were identified as alkenes, alkanes, and esters, were detected in VOCs of T-51. These results indicate that T. koningiopsis T-51 can exert biological control by integrating actions to suppress plant disease and promote plant growth. [ABSTRACT FROM AUTHOR]

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: 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]

Eight fungal strains (P1 to P8) were isolated from rapeseed stems (Brassica napus) infected with the blackleg pathogen Leptosphaeria biglobosa (Lb). They formed pycnidia with similar morphology to those of Lb, and thus were considered as Lb relatives (LbRs). The species-level identification of these strains was performed. Their virulence on rapeseed and efficacy in the suppression of Lb infection were determined, and the biocontrol potential and biocontrol mechanisms of strain P2 were investigated. The results showed that the LbRs belong to two teleomorphic genera in the family Didymellaceae, Didymella for P1 to P7 and Boeremia for P8. Pathogenicity tests on rapeseed cotyledons and stems indicated the LbRs were weakly virulent compared to L. biglobosa. Co-inoculation assays on rapeseed cotyledons demonstrated that P1 to P7 (especially P1 to P4) had a suppressive effect on Lb infection, whereas P8 had a marginal effect on infection by L. biglobosa. Moreover, D. macrostoma P2 displayed a more aggressive behavior than L. biglobosa in the endophytic colonization of healthy rapeseed cotyledons. Cultures of P2 in potato dextrose broth (PDB) and pycnidiospore mucilages exuded from P2 pycnidia showed antifungal activity to L. biglobosa. Further leaf assays revealed that antifungal metabolites (AM) of strain P2 from PDB cultures effectively suppressed infection by L. biglobosa, Botrytis cinerea (gray mold), and Sclerotinia sclerotiorum (white mold). An antifungal metabolite, namely penicillither, was purified and identified from PDB cultures and detected in pycnidiospore mucilages of strain P2. This study suggests that the LbRs are a repertoire for screening biocontrol agents (BCAs) against rapeseed diseases, and D. macrostoma P2 is a multi-functional BCA, a penicillither producer, and an endophyte. [ABSTRACT FROM AUTHOR]

A fungal strain (JC-1) of Simplicillium was isolated from a pod of oilseed rape (Brassica napus) infested with the blackleg pathogen Leptosphaeria biglobosa. This study was done to clarify its taxonomic identity using morphological and molecular approaches, to characterize its antifungal activity through bioassays and genome-based identification of antifungal metabolites, and to determine its efficacy in inducing systemic resistance (ISR) in oilseed rape. The results showed that JC-1 belongs to Simplicillium lamellicola. It displayed a strong antagonistic relationship with L. biglobosa, Botrytis cinerea (gray mold) and Sclerotinia sclerotiorum (stem rot). The cultural filtrates of JC-1 showed a high efficacy in suppressing infection by S. sclerotiorum on detached leaves of oilseed rape. Genome analysis indicated that JC-1 has the capability of producing multiple antifungal metabolites, including aureobasidin A1, squalestatin S1 and verlamelin. Inoculation of JC-1 on seeds of oilseed rape caused a suppressive effect on infection by L. biglobosa on the cotyledons of the resulting seedlings, suggesting that JC-1 can trigger ISR. Endophytic growth, accumulation of anthocyanins, up-regulated expression of CHI (for chalcone isomerase) and PR1 (for pathogenesis-related protein 1), and down-regulated expression of NECD3 (for 9-cis-epoxycarotenoid dioxygenase) were detected to be associated with the ISR. This study provided new insights into the biocontrol potential and modes of action of S. lamellicola. [ABSTRACT FROM AUTHOR]

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]

Aims: The study aimed at identifying volatile organic compounds (VOCs) produced by the non-pathogenic Fusarium oxysporum ( Fo) strain CanR-46, and to determine the anti-fungal spectrum and the control efficacy of the Fo-VOCs. Methods: The Fo-VOCs were identified by GC-MS. The antifungal activity of the Fo-VOCs was tested using the double-dish method. Soil contaminated with Verticillium dahliae was fumigated with the Fo-VOCs to test the control efficacy. The GFP-tagged derivative strain CanR-46GFP was tested to colonize cotton roots for prevention of infection by V. dahliae. Results: Nineteen VOCs were identified with eremophila-1 (10),11-diene being the most abundant. The Fo-VOCs inhibited mycelial growth of 14 fungal species including V. dahliae, delayed V. dahliae conidial germination, suppressed V. dahliae germ-tube elongation, and caused V. dahliae hyphal shriveling and collapse. Four synthetic chemicals, 5-hexenoic acid, limonene, octanoic acid and 3,4-2H-dihydropyran, in the Fo-VOCs profile showed antifungal activity against V. dahliae. The Fo-VOCs significantly ( P < 0.05) reduced severity of Verticillium wilt of cotton compared to the control. CanR-46GFP extensively colonized the cotton root tissues, thus effectively prevented V. dahliae infection. Conclusions: The VOC-producing fungus F. oxysporum CanR-46 is a promising biocontrol agent against V. dahliae. [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: 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]

Sclerotinia minor is a phytopathogenic fungus causing sclerotinia blight on many economically important crops. Here, we have characterized the biological and molecular properties of a novel endornavirus, Sclerotinia minor endornavirus 1 (SmEV1), isolated from the hypovirulent strain LC22 of S. minor. The genome of SmEV1 is 12,626 bp long with a single, large open reading frame (ORF), coding for a putative protein of 4020 amino acids. The putative protein contains cysteine-rich region (CRR), viral methyltransferase (MTR), putative DEXDc, viral helicase (Hel), and RNA-dependent RNA polymerase (RdRp) domains. The putative protein and the conserved domains are phylogenetically related to endornaviruses. SmEV1 does not contain a site-specific nick characteristic of most previously described endornaviruses. Hypovirulence and associated traits of strain LC22 and SmEV1 were readily cotransmitted horizontally via hyphal contact to isolates of different vegetative compatibility groups of S. minor. Additionally, SmEV1 in strain LC22 was found capable of being transmitted vertically through sclerotia. Furthermore, mycelium fragments of hypovirulent strain LC22 have a protective activity against attack by S. minor. Taken together, we concluded that SmEV1 is a novel hypovirulence-associated mycovirus with a wide spectrum of transmissibility, and has potential for biological control (virocontrol) of diseases caused by S. minor. [ABSTRACT FROM AUTHOR]