Yeasts occur in all environments and have been described as potent antagonists of various plant pathogens. Due to their antagonistic ability, undemanding cultivation requirements, and limited biosafety concerns, many of these unicellular fungi have been considered for biocontrol applications. Here, we review the fundamental research on the mechanisms (e.g., competition, enzyme secretion, toxin production, volatiles, mycoparasitism, induction of resistance) by which biocontrol yeasts exert their activity as plant protection agents. In a second part, we focus on five yeast species (Candida oleophila, Aureobasidium pullulans, Metschnikowia fructicola, Cryptococcus albidus, Saccharomyces cerevisiae) that are or have been registered for the application as biocontrol products. These examples demonstrate the potential of yeasts for commercial biocontrol usage, but this review also highlights the scarcity of fundamental studies on yeast biocontrol mechanisms and of registered yeast-based biocontrol products. Yeast biocontrol mechanisms thus represent a largely unexplored field of research and plentiful opportunities for the development of commercial, yeast-based applications for plant protection exist.

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Biocontrol Yeasts: Mechanisms and Applications

Damping-off is a disease that leads to the decay of germinating seeds and young seedlings, which represents for farmers one of the most important yield constraints both in nurseries and fields. As for other biotic stresses, conventional fungicides are widely used to manage this disease, with two major consequences. On the one hand, fungicide overuse threatens the human health and causes ecological concerns. On the other hand, this practice has led to the emergence of pesticide-resistant microorganisms in the environment. Thus, there are increasing concerns to develop sustainable and durable damping-off management strategies that are less reliant on conventional pesticides. Achieving such a goal requires a better knowledge of pathogen biology and disease epidemiology in order to facilitate the decision-making process. It also demands using all available non-chemical tools that can be adapted to regional and specific production situations. However, this still is not the case and major knowledge gaps must be filled. Here, we review up to 300 articles of the damping-off literature in order to highlight major knowledge gaps and identify future research priorities. The major findings are (i) damping-off is an emerging disease worldwide, which affects all agricultural and forestry crops, both in nurseries and fields; (ii) over a dozen of soil-borne fungi and fungus-like organisms are a cause of damping-off but only a few of them are frequently associated with the disease; (iii) damping-off may affect from 5 to 80% of the seedlings, thereby inducing heavy economic consequences for farmers; (iv) a lot of research efforts have been made in recent years to develop biocontrol solutions for damping-off and there are interesting future perspectives; and (v) damping-off management requires an integrated pest management (IPM) approach combining both preventive and curative tactics and strategies. Given the complex nature of damping-off and the numerous factors involved in its occurrence, we recommend further research on critical niches of complexity, such as seeds, seedbed, associated microbes and their interfaces, using novel and robust experimental and modeling approaches based on five research priorities described in this paper.

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Integrated management of damping-off diseases. A review

In plant-microbe interactions, plant sugars produced by photosynthesis are not only a carbon source for pathogens, but may also act as signals that modulate plant defense responses. Here, we report that decreasing sorbitol synthesis in apple (Malus domestica) leaves by antisense suppression of ALDOSE-6-PHOSPHATE REDUCTASE (A6PR) leads to downregulation of 56 NUCLEOTIDE BINDING/LEUCINE-RICH REPEAT (NLR) genes and converts the phenotypic response to Alternaria alternata from resistant to susceptible. We identified a resistance protein encoded by the apple MdNLR16 gene and a small protein encoded by the fungal HRIP1 gene that interact in both a yeast two-hybrid assay and a bimolecular fluorescence complementation assay. Deletion of HRIP1 in A. alternata enables gain of virulence on the wild-type control plant. Overexpression of MdNLR16 in two antisense A6PR lines increases resistance, whereas RNAi suppression of MdNLR16 in the wild-type control decreases resistance against A. alternata MdWRKY79 transcriptionally regulates MdNLR16 by binding to the promoter of MdNLR16 in response to sorbitol, and exogenous sorbitol feeding partially restores resistance of the antisense A6PR lines to A. alternata These findings indicate that sorbitol modulates resistance to A. alternata via the MdNLR16 protein that interacts with the fungal effector in a classic gene-for-gene manner in apple.

Sorbitol Modulates Resistance to Alternaria alternata by Regulating the Expression of an NLR Resistance Gene in Apple.

The population structure of Alternaria species associated with potato foliar diseases in China has not been previously examined thoroughly. Between 2010 and 2013, a total of 511 Alternaria isolates were obtained from diseased potato leaves sampled in 16 provinces, autonomous regions or municipalities of China. Based on morphological traits and molecular characteristics, all the isolates were identified as Alternaria tenuissima, A. alternata or A. solani. Of the three species, A. tenuissima was the most prevalent (75·5%), followed by A. alternata (18·6%) and A. solani (5·9%). Phylogenetic analysis based on sequences of the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) of representative Alternaria isolates showed that A. solani was distinct from the two small-spored Alternaria species. Phylogenetic analysis of the partial coding sequence of the histone 3 gene divided the same collection of isolates into three main clades representing A. tenuissima, A. alternata and A. solani, respectively. The pathogenicity of the isolates on detached leaves of potato cv. Favorite did not differ significantly between the three species or between isolates from different geographical origins. The results indicate that the population structure of Alternaria species associated with potato foliar diseases differs from that reported previously in China. This is the first report of A. tenuissima causing potato foliar diseases in China.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/ppa.12274

Characterization of Alternaria species associated with potato foliar diseases in China.

Cross-resistance, a phenomenon that a pathogen resists to one antimicrobial compound also resists to one or several other compounds, is one of major threats to human health and sustainable food production. It usually occurs among antimicrobial compounds sharing the mode of action. In this study, we determined the sensitivity profiles of Alternaria alternata, a fungal pathogen which can cause diseases in many crops to two fungicides (mancozeb and difenoconazole) with different mode of action using a large number of isolates (234) collected from seven potato fields across China. We found that pathogens could also develop cross resistance to fungicides with different modes of action as indicated by a strong positive correlation between mancozeb and difenoconazole tolerances to A. alternata. We also found a positive association between mancozeb tolerance and aggressiveness of A. alternata, suggesting no fitness penalty of developing mancozeb resistance in the pathogen and hypothesize that mechanisms such as antimicrobial compound efflux and detoxification that limit intercellular accumulation of natural/synthetic chemicals in pathogens might account for the cross-resistance and the positive association between pathogen aggressiveness and mancozeb tolerance. The detection of cross-resistance among different classes of fungicides suggests that the mode of action alone may not be an adequate sole criterion to determine what components to use in the mixture and/or rotation of fungicides in agricultural and medical sects. Similarly, the observation of a positive association between the pathogen’s aggressiveness and tolerance to mancozeb suggests that intensive application of site non-specific fungicides might simultaneously lead to reduced fungicide resistance and enhanced ability to cause diseases in pathogen populations, thereby posing a greater threat to agricultural production and human health. In this case, the use of evolutionary principles in closely monitoring populations and the use of appropriate fungicide applications are important for effective use of the fungicides and durable infectious disease management.

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Cross-resistance of the pathogenic fungus Alternaria alternata to fungicides with different modes of action

Potato (Solanum tuberosum L.), a worldwide, staple food crop, is susceptible to postharvest rots caused by a variety of fungal pathogens, including Fusarium spp., Alternaria spp., Phytophthora infestans, Helminthosporium solani, Rhizoctonia solani, and Colletotrichum coccodes. Rots resulting from infections by these pathogens cause a significant reduction in potato quality and marketable yield. Importantly, some of these decay fungi also produce mycotoxins that represent a potential risk to human health. In the present review, an overview and discussion are provided on the epidemiology and pathogenesis of decay fungi, especially Fusarium spp., that include recent data derived from genomic and phylogenetic analyses. The biosynthesis and functional role of fungitoxic metabolites such as trichothecene mycotoxins and fusaric acid, produced in rotted potatoes are also reviewed. Advances in pre- and postharvest measures for rot management, especially eco-friendly methods including physical control, biological control, the use of natural compounds, and other agricultural management practices are also reviewed. Lastly, novel approaches to control potato dry rot such as the use of mycoviruses and CRISPR technology are highlighted.

Pre- and postharvest measures used to control decay and mycotoxigenic fungi in potato (Solanum tuberosum L.) during storage.

Candida oleophila is an effective biocontrol control used to control postharvest diseases of fruits and vegetables. C. oleophila I-182 was an active agent used in the first-generation of yeast-based commercial product, Aspire™, for postharvest disease management. Several action modes, like competition for nutrients and space, induction of pathogenesis-related genes in host tissues, and production of extracellular lytic enzymes, have been demonstrated for the biological control activity exhibited by C. oleophila through which it inhibits postharvest pathogens. In the present study, the whole genome of C. oleophila I-182 was sequenced using PacBio and Illumina shotgun sequencing technologies, yielding an estimated genome size of 14.73 Mb. The genome size is similar in length to that of the model yeast strain Saccharomyces cerevisiae S288c. Based on the assembled genome, protein-coding sequences were identified and annotated with a gene description. The predicted genes were further assigned with gene ontology (GO) terms and clustered in special functional groups. A comparative analysis of C. oleophila proteome with the proteomes of 11 representative yeasts revealed 2 unique and 124 expanded families of proteins in C. oleophila. Availability of the genome sequence will facilitate a better understanding the properties of biocontrol yeasts at the molecular level.

/pdf/genome-sequence-assembly-and-characterization-of-the-3fecrbbjyv.pdf

Genome Sequence, Assembly, and Characterization of the Antagonistic Yeast Candida oleophila Used as a Biocontrol Agent Against Post-harvest Diseases.

Watermelon is an economically important crop in China and is commonly affected by Alternaria-like leaf blight that can result in significant economic losses. In this study, 830 Alternaria isolates, recovered from symptomatic watermelon leaves, were identified based on morphological traits, pathogenicity, and multilocus sequence analyses of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), histone 3 (HIS3), the internal transcribed spacer of ribosomal DNA (rDNA ITS), and the RNA polymerase II second largest subunit (RPB2). Watermelon isolates grouped to five Alternaria species and one unclassified Alternaria species. They were A. tenuissima, A. alternata, A. cucumerina, A. infectoria, A. gaisen, and Alternaria sp. Notably, A. tenuissima was the most prevalent (73.5%) of the six isolated species, followed by A. alternata (25.0%), A. cucumerina (1.1%), Alternaria sp. (0.2%), A. infectoria (0.1%), and A. gaisen (0.1%). Pathogenicity tests demonstrated that all six Alternaria species could produce brown necrotic lesions on detached leaves of watermelon. The average disease incidence (75.1%) and average disease index (60.8) of watermelon resulting from inoculation of leaves with A. cucumerina were significantly higher than levels resulting from A. alternata (52.9% and 37.2) and A. tenuissima (47.5% and 30.8). Inoculation with Alternaria sp. resulted in a disease incidence (70.0%) and disease index (51.5), which were lower than those of A. cucumerina. The disease incidence and disease index in watermelon leaves inoculated with the one isolate of A. infectoria and the one isolate of A. gaisen present in the inoculated leaves were 28.9% and 16.4, and 48.9% and 31.4, respectively. Results of the study indicate that Alternaria species associated with watermelon leaf blight in China are more diverse than that has been previously reported. This is the first report globally of A. infectoria, A. gaisen, and an unclassified Alternaria species as causal agents of leaf blight on watermelon.

Morphological and Molecular Characterization of Alternaria Species Causing Leaf Blight on Watermelon in China.

Two binucleate Rhizoctonia (BNR) isolates were recovered from potato cankered stems in Heilongjiang Province, northeastern China. Their cultural appearance on potato dextrose agar remained whitish as the cultures aged. White monilioid cells formed in the fluffy aerial hyphae, whereas no sclerotia appeared during the incubation. The two isolates could anastomose with each other, but they failed to anastomose with reference strains of BNR from AG-A to AG-Q, and AG-U. Analyses of restriction fragment length polymorphism (RFLP) of internal transcribed spacer of ribosomal DNA (rDNA-ITS) regions confirmed that these two isolates differed from the reference strains. The phylogenetic tree based on the sequences of rDNA-ITS regions showed that they were located in a distinct clade from other BNR AGs. These collective results suggested that the isolates recovered from potato in this study belonged to a new BNR AG designated as AG-W. Pathogenicity tests under glasshouse conditions revealed that both isolates were ab...

https://apsjournals.apsnet.org/doi/pdf/10.1094/PDIS-01-15-0036-RE

X. H. Wu

Papers

Characterization of Alternaria species associated with potato foliar diseases in China.

Pre- and postharvest measures used to control decay and mycotoxigenic fungi in potato (Solanum tuberosum L.) during storage.

Genome Sequence, Assembly, and Characterization of the Antagonistic Yeast Candida oleophila Used as a Biocontrol Agent Against Post-harvest Diseases.

Morphological and Molecular Characterization of Alternaria Species Causing Leaf Blight on Watermelon in China.

Characterization of a New Anastomosis Group (AG-W) of Binucleate Rhizoctonia, Causal Agent for Potato Stem Canker.