Advances in fundamental and applied studies in China of fungal biocontrol agents for use against arthropod pests

Entomopathogenic fungi, such as Beauveria bassiana and Metarhizium anisopliae, are environmentally friendly biocontrol agents (BCAs) against various arthropod pests. We provide an overview to the past-decade advances in fungal BCA research and application in China. Since 1960s, fungal BCAs have been mass-produced for application and at present, thousands of tons of their formulations are annually applied to control forest, agricultural, greenhouse and grassland insect pests throughout the country. Apart from technical advances in mass production, formulation and application of fungal BCAs, basic studies on the genomics, molecular biology, genetic engineering and population genetics of fungal entomopathogens have rapidly progressed in the past few years in China. The completed genomic studies of M. anisopliae, Metarhizium acridum, B. bassiana and Cordyceps militaris provide profound insights into crucial gene functions, fungal pathogenesis, host–pathogen interactions and mechanisms involved in fungal sexuality. New knowledge gained from the basic studies has been applied to improve fungal virulence and stress tolerance for developing more efficacious and field-persistent mycoinsecticides by means of microbial biotechnology, such as genetic engineering. To alleviate environmental safety concerns, more efforts are needed to generate new data not only on the effects of engineered BCAs on target and non-target arthropods but also on their potential effects on gene flow and genetic recombination before field release.     

Fungal antagonists of the plant pathogen Rhizoctonia solani: selection, control efficacy and influence on the indigenous microbial community

A broad spectrum of fungal antagonists was evaluated as potential biocontrol agents (BCAs) against the soil-borne pathogen Rhizoctonia solani using a new combination of in vitro and in vivo assays. The in vitro characterisation of diverse parameters including the ability to parasitise mycelium and to inhibit the germination of Rhizoctonia sclerotia at different temperatures resulted in the selection of six potential fungal antagonists. These were genotypically characterised by their BOX-PCR fingerprints, and identified as Trichoderma reesei and T. viride by partial 18S rDNA sequencing. When potato sprouts were treated with Trichoderma, all isolates significantly reduced the incidence of Rhizoctonia symptoms. Evaluated under growth chamber conditions, the selected Trichoderma isolates either partly or completely controlled the dry mass loss of lettuce caused by R. solani. Furthermore, the antagonistic Trichoderma strains were active under field conditions. To analyse the effect of Trichoderma treatment on indigenous root-associated microbial communities, we performed a DNA-dependent SSCP (Single-Strand Conformation Polymorphism) analysis of 16S rDNA/ITS sequences. In this first assessment study for Trichoderma it was shown that the pathogen and the vegetation time had much more influence on the composition of the microbiota than the BCA treatment. After evaluation of all results, three Trichoderma strains originally isolated from Rhizoctonia sclerotia were selected as promising BCAs.     

Regulating biocontrol agents: a historical perspective and a critical examination comparing microbial and macrobial agents

Ever since the inclusion of microbial biocontrol agents (MBCAs) within the regulatory frameworks initially designed for chemical pesticides, there has been awareness that these frameworks are not optimal for assessment and registration of new microbial biocontrol products. It is often claimed that the regulatory situation has contributed to a relatively slow uptake of microbial biocontrol in practice. In contrast to the MBCAs, non-indigenous invertebrate biocontrol agents (IBCAs) are regulated in many countries through quarantine and other biosecurity related legislation for prevention of introduction of alien organisms, whereas use of indigenous IBCAs are generally unregulated. In this study, we investigate what scientific support there is for performing evaluations of these two main groups of biocontrol agents (BCAs) within different frameworks. We compare potential risks of MBCAs and IBCAs, present a retrospective analysis of the development and implementation of the regulatory frameworks, and compare current requirements for MBCAs with those for other applications with microorganisms. One conclusion is that the ecological risks are of similar types between the two groups of BCAs, and that for both groups the environmental safety is most pertinently evaluated according to biological and ecological principles. The main difference between MBCAs and IBCAs with respect to human health is that the former may cause infectious disease. However, we found no evidence that this hazard is more serious for microorganisms for biocontrol than for microbes used in other types of applications, which generally have substantially lower regulatory demands than those for MBCAs. Several international initiatives have produced helpful guidelines and recommendations for simplified assessments and authorisations of BCAs. Still, we conclude that as long as MBCAs are evaluated within systems initially developed for chemicals, the risk for inappropriate emphasis of chemical hazards and therefore unnecessarily complicated assessments will be maintained. Therefore, this study supports the idea that development of new systems for the regulatory oversight of MBCAs, possibly a mutual framework covering all living BCAs, should be considered. Research issues that need to be further explored are to what extent utilisation of MBCAs actually results in increased exposure of non-targets to microorganisms, the biogeography and microbial ecology of representative MBCAs, and finally development of better methodology for determining potential human toxicity and pathogenicity of candidate MBCAs.     

A screening strategy of fungal biocontrol agents towards Verticillium wilt of cotton

Three hundred and seventy-three fungal isolates were obtained from the endorhiza, rhizosphere, and bulk soil of field-grown cotton plants. One hundred and five of them produced obvious inhibition zones against Verticillium dahliae Kleb., so they were selected as antagonists towards this pathogen. An assessment system was established to evaluate these 105 antagonists for their biocontrol potential and plant growth-promoting potential. Their biocontrol potential was assessed according to their in vitro antagonistic activity against V. dahliae and activities of fungal cell wall degrading enzymes including protease, cellulase, and chitinase. Their plant growth-promoting potential was assessed according to their in vitro activities of solubilizing phosphate and fixing nitrogen. Thirty-three antagonists received at least three points of the total value of assessed biocontrol potential and plant growth-promoting potential and were tested for their biocontrol efficacy and growth-promoting effect on cotton under greenhouse conditions. Twelve of them achieved positive biocontrol efficacy ranging from 8.58% to 69.78%; the conventional correlation coefficient of the biocontrol efficacy of these antagonists with their assessed biocontrol potential was 0.926. By using the screening strategy developed in this study, Fusarium oxysporum strain By125, Nectria haematococca Bx247, and Phomopsis sp. By231 were identified as potential BCAs for controlling Verticillium wilt in cotton, for they achieved biocontrol efficacy of 63.63–69.78% towards this disease and increased biomass by 18.54–62.63% under greenhouse conditions. The present study also demonstrated that the endorhiza of field-grown cotton plants may be a richer source of potential BCAs against Verticillium wilt than the rhizosphere and bulk soil.     

Evaluation of biocontrol agents and potassium silicate for the management of powdery mildew of zucchini

Investigations were conducted under greenhouse and field conditions to evaluate the effects of potential biocontrol agents (BCAs) and soluble silicon (Si) on powdery mildew of zucchini caused by Podosphaera xanthii. Five BCAs were applied as foliar sprays to zucchini leaves and Si was drenched weekly into the rhizosphere of these plants.In the greenhouse, all BCAs provided significant control of powdery mildew with fungal isolates, reducing disease levels by up to 90%. Si alone reduced powdery mildew by as much as 35% and improved the efficacy of most of the biocontrol agents. Higher disease pressure reduced the efficacy of Si on powdery mildew but did not affect the performance of the BCAs. In the field, a disease reduction of 10–70% was achieved by BCAs and Si. Lower temperatures and high humidity ranges were suitable for optimal performances. The efficacy of the bacterial BCA, Serratia marcescens – B15 and silicon diminished at temperatures above 25 °C. The fungal BCAs (Clonostachys rosea – EH and Trichothecium roseum – H20) were better suited to higher temperatures (25–30 °C) and were tolerant of low RH values. Application of K₂SiO₂ to zucchini roots increased the level of Si in the leaves, which was responsible for suppression of the disease.     

Biocontrol of strawberry fruit infected by Botrytis cinerea: Effects on the microbial communities on fruit assessed by next‐generation sequencing

Fruit grey mould, caused by the fungus Botrytis cinerea, is known to be a harmful disease of strawberry at postharvest stage. However, effects of an application of biological control agents (BCAs) on strawberry fruit in terms of shift in the microbial community are still unknown. The present research aimed to investigate the effects of an application of BCAs on postharvest microbial populations present on strawberry fruits. Strawberry plants were sprayed with three kinds of BCA, RhizoVital 42 fl. (Bacillus amyloliquefaciens FZB42), Trianum‐P (Trichoderma harzianum T22) and Naturalis (Beauveria bassiana ATCC 74040), targeting Botrytis cinerea fungus. Control plots were composed of water and fungicide treatments. Microbial communities (bacteria and fungi) were analysed via next‐generation sequencing on an Illumina MiSeq. Analysis of 16S RNA and ITS rRNA sequences indicated that the BCAs application modified both bacterial and fungal community compositions and diversity. An application of two BCAs together had more effects on microbial community composition than a single application. These results suggest that BCAs can modify bacterial and fungal community composition and diversity on strawberry fruits, which may consequently improve the efficiency and establishment of these products on control of postharvest diseases of fruits, such as grey mould.     

Biological control in the microbiome era: Challenges and opportunities

Biocontrol research has long been focused on the study of single strains of biocontrol agents (BCAs) and on their interaction with pathogens and host plants. Further focus on plant-associated microbial communities was suggested several years ago, but significant advances only occurred recently. The advent of high-throughput sequencing (or next-generation sequencing – NGS) technologies is now driving a paradigm change that allows researchers to integrate microbial community studies into the traditional biocontrol approach. This integration could answer old scientific questions, and will raise new biocontrol hypotheses. Microbial communities could impact disease control through their interaction with host plants, pathogens, and BCAs. A better understanding of these interactions will provide unexpected opportunities to develop innovative biocontrol methods against plant pathogens. For example, formulation or timing of BCA application can be improved, “helper” microbial strains can be selected, or molecules driving the microbiota to a pathogen-resistant composition (“prebiotic” approach) can be developed. The five main challenges of microbiome implementation in biocontrol research are also described, i.e. (i) the management of technical errors and biases, (ii) the growing importance of bioinformatics, (iii) the adaptation of experimental schemes, (iv) the appropriate interplay between NGS and other technologies, and (v) the need to complete current genome databases.     

Using combinations of biocontrol agents to control Botrytis cinerea on strawberry leaves under fluctuating temperatures

Experiments were conducted with Botrytis cinerea on strawberry leaves to investigate where combinations of commercially available biological control agents (BCAs) might control B. cinerea more effectively than individual BCAs. Specifically, we studied the persistence of biocontrol activities, spread of BCAs among leaves, and biocontrol efficacy in relation to application regimes: mixed versus single BCA, pre-versus post-inoculation application, and sequential versus simultaneous application. Three BCA products (Sentinel, Serenade and Trianum) were used for this study. Overall, Serenade did not significantly reduce sporulation of B. cinerea on strawberry leaf discs whereas Sentinel and Trianum gave a similar and significant biocontrol efficacy. Biocontrol efficacy remained almost unchanged 10 days after application at 20/20°C (day/night) or 24/16°C temperature regimes. In contrast, reduced biocontrol efficacy at 26/14°C suggests BCA survival was reduced under these conditions. Incidence of B. cinerea sporulation on leaf discs was ca. 60% higher on leaves that emerged after the BCA application than on leaves directly exposed to BCA, indicating insufficient amount of the BCA had managed to spread to new leaves. Combinations of BCAs, whether applied simultaneously or sequentially (48 h apart), did not improve disease control over the most effective BCA within the combination applied alone. This indicated possible antagonism or interference between the BCAs. Results suggested that there was significant antagonism for most combinations of the three BCAs tested and the degree of antagonism increased as the time from BCA application to pathogen introduction lengthened.     

Potential of soil-derived fungal biocontrol agents applied as a soil amendment and a seed coating to control Verticillium wilt of sugar beet

Verticillium dahliae (Vd) is an emerging threat to sugar beet production. Control measures such as fungicides are not available and the utilisation of resistant cultivars is very limited. Hence, we explored the potential of two soil-derived fungal biocontrol agents (BCAs), Fusarium oxysporum F2 (FoF2) and Verticillium tricorpus 1808 (Vt1808), against Verticillium wilt of sugar beet. Pathogenicity tests revealed that Vd caused over 90% disease incidence and severity and led to a significant yield reduction, whereas BCAs neither inhibited nor promoted plant growth. Germination rate was higher in BCA-treated seeds compared to untreated ones. Viability of both BCAs was significantly reduced after six months of storage in liquid methylcellulose formulation (MC), while BCA concentrations remained stable on stored seeds treated with MC. In contrast, Vt1808 produced in sand-rye flour formulation remained 100% viable after storage. Similarly, post-storage analysis of the FoF2 talcum powder formulation revealed improved colony forming units, but increments were not significant. In vitro, both BCAs caused no growth inhibition zones and only insignificant Vd growth reductions were observed. In the greenhouse, soil amendment with a higher dose of FoF2:Vt1808 mixture resulted in substantial reductions of disease severity on the crown (33.3%), disease incidence (55.6%) and disease severity (68.8%) on the beet as well as an improved yield (32.9%). In contrast, seed coating did not reduce symptoms on the crown and on the beet. In the field, both BCAs did not provide significant disease reductions. Nevertheless, a promising result was achieved with the application of FoF2. Despite the need for improvement of the biocontrol activity, the results of this study demonstrate the suitability of the optimised BCA formulations for utilisation in commercial sugar beet production.     

Biotechnological approaches to develop bacterial chitinases as a bioshield against fungal diseases of plants

Fungal diseases of plants continue to contribute to heavy crop losses in spite of the best control efforts of plant pathologists. Breeding for disease-resistant varieties and the application of synthetic chemical fungicides are the most widely accepted approaches in plant disease management. An alternative approach to avoid the undesired effects of chemical control could be biological control using antifungal bacteria that exhibit a direct action against fungal pathogens. Several biocontrol agents, with specific fungal targets, have been registered and released in the commercial market with different fungal pathogens as targets. However, these have not yet achieved their full commercial potential due to the inherent limitations in the use of living organisms, such as relatively short shelf life of the products and inconsistent performance in the field. Different mechanisms of action have been identified in microbial biocontrol of fungal plant diseases including competition for space or nutrients, production of antifungal metabolites, and secretion of hydrolytic enzymes such as chitinases and glucanases. This review focuses on the bacterial chitinases that hydrolyze the chitinous fungal cell wall, which is the most important targeted structural component of fungal pathogens. The application of the hydrolytic enzyme preparations, devoid of live bacteria, could be more efficacious in fungal control strategies. This approach, however, is still in its infancy, due to prohibitive production costs. Here, we critically examine available sources of bacterial chitinases and the approaches to improve enzymatic properties using biotechnological tools. We project that the combination of microbial and recombinant DNA technologies will yield more effective environment-friendly products of bacterial chitinases to control fungal diseases of crops.     

Screening of antagonistic bacterial strains against Meloidogyne incognita using protease activity

Twenty-six bacterial strains that had demonstrated antagonism to some fungal and bacterial pathogens were evaluated for their ability to inhibit Meloidogyne incognita Kofoid & White. The inhibition rates of egg-hatching and second-stage juveniles (J2) mortality of M. incognita by these strains ranged from ?16.5 to 87.4% and from 1.3 to 77.8%, respectively. The 12 strains causing J2 mortality over 40% were chosen for greenhouse experiments in which their biocontrol efficacy reached 33.3–65.6%. On the other hand, among the 26 strains, 20 demonstrated in vitro protease activity and 14 revealed chitinase activity. Significantly, strains Bacillus sp. AR156 and GJ24 in greenhouse tests showed the strongest protease activities. The analyses of the relationships of the efficacy of the 12 strains with their protease and chitinase activities, respectively, indicated that biocontrol efficacy was highly correlated with protease activity (r=0.92, P<0.001) but barely correlated with chitinase activity. The strong positive correlation between protease activity and efficacy suggests that in vitro protease activity could be used as a parameter for selecting biological control agents (BCAs) against root-knot nematodes. Consistently, the biocontrol efficacy of AR156, GJ24, abamectin reached 74.3, 73.4, and 40.9% in the field in Huai-an, Jiangsu; and 71, 69.9, and 37% in Zao-zhuang, Shandong, respectively. The fact that the strains with high protease activities also had significantly higher biocontrol efficacy than abamectin in the field implies that in vitro protease activity may be adopted as a reliable new parameter for speeding up the process of screening the biological control agents (BCAs).     

Seedling vaccination by stem injecting a conidial suspension of F2, a non-pathogenic Fusarium oxysporum strain, suppresses Verticillium wilt of eggplant

Several bacterial and fungal strains have been evaluated as biocontrol agents (BCAs) against Verticillium dahliae. In these studies, the BCAs were applied as a root drenching inoculum; however, this application method may have an adverse effect on the native, beneficial for the plants, microbial community. In the present study, it was evaluated whether endophytic application by stem injecting a conidial suspension of the non pathogenic Fusarium oxysporum F2 strain, isolated from a V. dahliae suppressive compost amendment, could reduce significantly Verticillium wilt symptom development in eggplants. It was revealed that stem injection of F2 seven days before transplanting the seedlings to soil infested by V. dahliae microsclerotia resulted in reduced disease severity compared to the control treatment. To visualise F2 ramification into the plant vascular system eggplant stems were injected with an EGFP transformed F2 strain. It was shown that F2 colonises effectively the plant vascular tissues over a long period of time as it was assessed by F2 DNA levels. In parallel, qPCR analysis showed that the application of F2 reduced significantly the amount of V. dahliae DNA in the stem tissues compared to the control treatment.     

Screening tomato-associated bacteria for biological control of grey mold on tomato

Aiming at discovering effective biocontrol agents (BCAs) against grey mold on tomato caused by Botrytis cinerea Pers., we selected 819 bacterial isolates from the surface as well as the interior of the roots, stems, and leaves of tomato plants grown in B. cinerea-infested fields. In a dual-culture assay, 116 isolates (14.16%) showed antagonism against B. cinerea and fewer ones against five additional tomato-associated fungal pathogens – Pythium ultimum, Phytophthora capsici, Fusarium oxysporum f. sp. lycopersici, Sclerotinia sclerotiorum and Ralstonia solanacearum. Thirty-one isolates with antagonism to B. cinerea and at least one of the five additional pathogens were assessed for their efficacy in controlling grey mold on tomato in a greenhouse test. Thirteen of them attained the efficacy over 50% and were subjected to the second greenhouse test, in which 12 isolates consistently accomplished the biocontrol efficacy over 50%, with isolates ABc28 and ABc22 achieving the efficacy of 66.71% and 64.90%, respectively. Under greenhouse conditions, the above two as well as isolates ABc2, ABc11 and ABc17 increased tomato biomass by more than 20% in comparison with the control. The 12 antagonistic isolates accomplishing the biocontrol efficacy over 50% in both greenhouse tests were considered potential BCAs against grey mold, which were identified as Pseudomonas spp., Pantoea spp., Bacillus spp. and Chryseobacterium spp. Ten of them were found to produce at least one of the three hydrolytic enzymes (protease, cellulase and chitinase) and/or siderophore, which might be involved in their mechanisms of suppressing the disease. Based on the origin of these 12 strains, the leaf tissue, especially the leaf interior, of tomato plants grown in a B. cinerea-infested field appears to be a good source of potential BCAs against grey mold.     

Molecular characterization and identification of biocontrol isolates of Trichoderma spp

The most common biological control agents (BCAs) of the genus Trichoderma have been reported to be strains of Trichoderma virens, T. harzianum, and T. viride. Since Trichoderma BCAs use different mechanisms of biocontrol, it is very important to explore the synergistic effects expressed by different genotypes for their practical use in agriculture. Characterization of 16 biocontrol strains, previously identified as "Trichoderma harzianum" Rifai and one biocontrol strain recognized as T. viride, was carried out using several molecular techniques. A certain degree of polymorphism was detected in hybridizations using a probe of mitochondrial DNA. Sequencing of internal transcribed spacers 1 and 2 (ITS1 and ITS2) revealed three different ITS lengths and four different sequence types. Phylogenetic analysis based on ITS1 sequences, including type strains of different species, clustered the 17 biocontrol strains into four groups: T. harzianum-T. inhamatum complex, T. longibrachiatum, T. asperellum, and T. atroviride-T. koningii complex. ITS2 sequences were also useful for locating the biocontrol strains in T. atroviride within the complex T. atroviride-T. koningii. None of the biocontrol strains studied corresponded to biotypes Th2 or Th4 of T. harzianum, which cause mushroom green mold. Correlation between different genotypes and potential biocontrol activity was studied under dual culturing of 17 BCAs in the presence of the phytopathogenic fungi Phoma betae, Rosellinia necatrix, Botrytis cinerea, and Fusarium oxysporum f. sp. dianthi in three different media.     

Bacillus thuringiensis C25 suppresses popcorn disease caused by Ciboria shiraiana in mulberry (Morus australis L.)

The mulberry tree is an important crop for silkworm farming and for the health care industry. In Asia, the annual productivity of mulberry fruits is greatly reduced due to popcorn disease mainly caused by Ciboria shiraiana, a sclerotia-forming fungal pathogen. To date, the development of efficient biocontrol agents (BCAs) against this disease has been hampered by the recalcitrance of C. shiraiana to in vitro culturing methods. Here, we established alternative in vitro antifungal assays that directly monitored the effects of BCAs on the growth of C. shiraiana apothecia and further reported that Bacillus thuringiensis C25 suppressed popcorn disease in field conditions. Initially, from mulberry drupelets showing the popcorn disease symptoms, we confirmed the infection of C. shiraiana and observed its morphology in asexual stage. Then, apothecia of C. shiraiana were induced from the sclerotia collected from the disease-infested orchard. Two bacterial isolates, Enterobacter sp. C5 and B. thuringiensis C25, strongly suppressed the elongation and fresh weight accumulation of apothecia stalks, the width of hymenium, and ascus and ascospore formation of C. shiraiana. In addition, both bacterial isolates degraded the ultrastructure of hymenium cells in C. shiraiana apothecia. Ultimately, treatment of mulberry trees with B. thuringiensis C25 mitigated the incidence of popcorn mulberry disease under field conditions. In conclusion, B. thuringiensis C25 is the first reported BCA shown to efficiently control mulberry popcorn disease. Our results also support that B. thuringiensis exerts diverse biocontrol roles in addition to insecticidal behaviour.     

Mechanisms of action and biocontrol potential of Trichoderma against fungal plant diseases - A review

Plant diseases are among the major causes of the low productivity of crops, causing yield losses of up to 30%, heralding an enormous threat to global food security. Indiscriminate use of chemical-based fungicides for controlling fungal diseases has raised severe concerns about ecosystem health. Moreover, pathogens have become insensitive against these chemicals necessitating excessive use of chemicals for adequate control. The resulting accumulation of these chemicals in the food chain has provoked numerous health complications. For combating the adversaries of chemical-based fungicides, biological control of fungal pathogens is proposed as an eco-friendly alternative. Among various biological controls, Trichoderma-based biological control agents (BCAs) are widely used in agriculture for controlling soil-borne pathogens. These BCAs are commercialized and known as; stimulators of resistance in plants, growth enhancers, bio-fertilizers, and bio-pesticides. Biological management of plant pathogens has yielded valuable results in the sustainability of ecosystems and compelling improvements in the quality and quantity of agricultural produce. These BCAs exhibit potential against pathogens, remarkably improve photosynthesis, plant growth, and nutrient use efficiency for impressive crop yields. Despite these peculiarities, Trichoderma's mechanisms against pathogens and their growth promotional effects are not thoroughly investigated, hence formulating the prime objective of the current review. Along with these, Trichoderma-based fungicides marketed in different geographical locations are encompassed in this review. Finally, the knowledge gaps and future research directions for improving the efficacy of Trichoderma-based BCAs are discussed.     

The fungal biocontrol agent Coniothyrium minitans: production by solid-state fermentation, application and marketing

Biological control agents (BCAs) are potential alternatives for the chemical fungicides presently used in agriculture to fight plant diseases. Coniothyrium minitans is an example of a promising fungal BCA. It is a naturally occurring parasite of the fungus Sclerotinia sclerotiorum, a wide-spread pathogen which substantially reduces the yield of many crops. This review describes, exemplified by C. minitans, the studies that need to be carried out before a fungal BCA is successfully introduced into the market. The main aspects considered are the biology of C. minitans, the development of a product by mass production of spores using solid-state fermentation technology, its biocontrol activity and marketing of the final product.     

Influence of different growth conditions on the survival and the efficacy of freeze-dried Pseudomonas fluorescens strain Pf153

High viability, storability and tolerance to variable environmental conditions are key factors in the development of microbial biological control agents (BCAs). The efficacy of microbial BCAs is influenced by the culture conditions and formulation process. Therefore, we investigated the influence of diverse growth conditions on the survival during freeze-drying and on the biocontrol efficacy of Pseudomonas fluorescens strain Pf153. Culture time, temperature and media, mild heat shock and pH change influenced the bacterium viability after freeze-drying. The best survival rate was reached by cultivation in King’s broth for 16 or 20 h. Growth temperatures of 25 and 30°C and a mild heat shock at 35°C for one hour influenced the survival rate positively. In all bioassays against Botrytis cinerea on Vicia faba leaves, Pf153 showed a significant increased efficacy compared to the untreated control. No differences of the efficacy between fresh and freeze-dried cells were observed. Furthermore, a growth temperature of 20°C increased the efficacy of Pf153 against B. cinerea. These results underline that the quality of the formulated product can be improved by manipulating the fermentation process.     

Effectiveness of 3 antagonistic bacterial isolates to control Rhizoctonia solani Kühn on lettuce and potato

Rhizoctonia solani causes yield losses in numerous economically important European crops. To develop a biocontrol strategy, 3 potato-associated ecto- and endophytically living bacterial strains Pseudomonas fluorescens B1, Pseudomonas fluorescens B2, and Serratia plymuthica B4 were evaluated against R. solani in potato and in lettuce. The disease-suppression effect of the 3 biocontrol agents (BCAs) was tested in a growth chamber and in the field. In growth chamber experiments, all 3 BCAs completely or significantly limited the dry mass (DM) losses on lettuce and the disease severity (DS) caused by R. solani on potato sprouts. Strain B1 showed the highest suppression effect (52% on average) on potato. Under field conditions, the DS on both crops, which were bacterized, decreased significantly, and the biomass losses on lettuce decreased significantly as well. The greatest disease-suppression effect on potato was achieved by strain B1 (37%), followed by B2 (33%) and then B4 (31%), whereas the marketable tuber yield increased up to 12% (B1), 6% (B2), and 17% (B4) compared with the pathogen control at higher disease pressure. Furthermore, in all experiments, B1 proved to be the most effective BCA against R. solani. Therefore, this BCA could be a candidate for developing a commercial product against Rhizoctonia diseases. To our knowledge, this is the first report on the high potential of endophytes to be used as a biological control agent against R. solani under field conditions.     

猕猴桃采后真菌病害生物防治研究进展

猕猴桃果实易受多种病原真菌的侵染,采后易发生软腐病、灰霉病、青霉病等严重危害果实品质的真菌性病害。传统的有效防治方法主要为采前使用化学杀菌剂,但其易污染环境并可能危害人类健康。目前,已有一系列生物防治方法被研究并报道,这些方法可有效防控猕猴桃采后真菌病害：第一类为天然抑菌物质,包括植物提取物和其他天然物质;第二类为拮抗微生物,包括生防酵母、生防细菌、生防木霉,生防菌也能与物理及化学方法协同发挥作用。本文概述了各类生物防治方法在猕猴桃采后真菌病害绿色防控方面的研究进展及生防机理,并提出了目前存在的问题,最后展望了这一领域今后的研究方向。