Canadian Journal of Plant Pathology ISSN: 0706-0661 (Print) 1715-2992 (Online) Journal homepage: https://www.tandfonline.com/loi/tcjp20 Biological control of powdery mildew on Cornus florida using endophytic Bacillus thuringiensis Emily RotichEMILY ROTICH, Margaret T. MmbagaMARGARET T. MMBAGA & Jacqueline JoshuaJACQUELINE JOSHUA To cite this article: Emily RotichEMILY ROTICH, Margaret T. MmbagaMARGARET T. MMBAGA & Jacqueline JoshuaJACQUELINE JOSHUA (2019): Biological control of powdery mildew on Cornusflorida using endophytic Bacillusthuringiensis, Canadian Journal of Plant Pathology, DOI: 10.1080/07060661.2019.1641555 To link to this article: https://doi.org/10.1080/07060661.2019.1641555 Accepted author version posted online: 08 Jul 2019. Published online: 31 Jul 2019. Submit your article to this journal Article views: 101 View related articles View Crossmark data Full Terms & Conditions of access and use can be found at https://www.tandfonline.com/action/journalInformation?journalCode=tcjp20 Can. J. Plant Pathol., 2019 https://doi.org/10.1080/07060661.2019.1641555 Disease control/Moyens de lutte Biological control of powdery mildew on Cornus florida using endophytic Bacillus thuringiensis EMILY ROTICH1, MARGARET T. MMBAGA2 AND JACQUELINE JOSHUA3 1College of Sciences and Mathematics, Department of Biology, Belmont University, Nashville, TN 37212, USA 2College of Agriculture, Human and Natural Sciences, Department of Agricultural and Environmental Sciences, Tennessee State University, Nashville, TN 37209, USA 3Division of Crop Science, Monsanto Company, Chesterfield, MO, 63017, USA (Accepted 4 July 2019) Abstract: Powdery mildew (Erysiphe pulchra) is a devastating disease in nursery production of flowering dogwood (Cornus florida L.). Disease management relies heavily on season-long use of chemical fungicides and there is a demand for alternative strategies. An endophytic bacterium IMC8 was isolated from disease-free C. florida stem tissue, and was shown to be an effective biological control agent against E. pulchra in growth chamber, greenhouse and shadehouse environments. This isolate, identified as Bacillus thuringiensis, exhibited normal growth in NaCl concentrations up to 6%, pH 5 to 11, and temperatures up to 50°C. It was compatible with the conventional fungicide thiophanate methyl and clarified hydrophobic extract of neem seed oil at label recommended concentrations. The isolate IMC8 produced volatile compounds identified by GC/MS, which included antifungal and antibacterial compounds and suggested antibiosis as a mode of action. Scanning electron microscopy revealed lysis of powdery mildew spores and hyphae, which suggested parasitism as a second mode of action. These observations suggested that the isolate IMC8 has potential as a biological control agent in the management of powdery mildew on C. florida, and would tolerate a wide range of salinity, pH and high temperature conditions. It could be included as a component of integrated pest management in rotation or tank mix with lower levels of conventional fungicides, or used with clarified hydrophobic extract of neem seed oil. Keywords: biological control agents, endophytes, Erysiphe pulchra, integrated disease management, mycoparasitism Résumé: L’oïdium (Erysiphe pulchra) est une maladie dévastatrice dans la production en pépinière de cornouiller à fleurs de Floride (Cornus florida). La gestion des maladies repose largement sur l’utilisation de produits chimiques pendant toute la saison. Couramment, il existe une demande pour des stratégies alternatives au produit chimique. La recherche d’agents de lutte biologique contre l’oïdium a permis l’isolement d’une bactérie endophyte, IMC8, à partir de tissu de C. florida indemne de maladie. L’isolat IMC8 s’est avéré être un agent de lutte biologique efficace contre E. pulchra dans les environnements de chambre de culture, de serre et serre d’ombre. Cet isolat identifié comme Bacillus thuringiensis a présenté une croissance normale dans une large gamme de niveaux de salinité et de pH, allant jusqu’à 6% de NaCl et pH entre 5 et 11, ainsi qu’à des températures allant jusqu’à 50°C. Il était compatible avec le fongicide classique thiophanate de méthyle et le biofongicide, l’extrait hydrophobe clarifiéd’huile de graines de neem, à des concentrations couramment utilisées dans les applications de lutte contre l’oïdium au pepinere commercial. L’isolat IMC8 a produit des composés volatils identifiés par GC/MS, parmi lesquels des composés antifongiques et antibactériens ont suggéré l’antibiose comme mode d’action. D’ailleurs, la microscopie électronique à balayage a révélé la lyse des spores et des hyphes de l’oïdium, suggérant le parasitisme comme second mode d’action. Ces observations suggèrent que l’isolat IMC8 présente un potentiel en tant qu’agent de lutte biologique dans la gestion de l’oïdium et qu’il tolérerait une large gamme de conditions salines, de pH et de températures élevées. Les résultats présentés suggèrent que l’isolat IMC8 peut constituer une alternative aux fongicides classiques pour la gestion de l’oïdium et réduire l’utilisation de fongicides dans la production en pépinière de C. florida, soit Correspondence to: Emily Rotich. E-mail: [email protected] © 2019 The Canadian Phytopathological Society Published online 31 Jul 2019 E. Rotich et al. 2 comme composante de la lutte intégrée en alternance, soit en mélange en réservoir de pulvérization avec des niveaux inférieurs de fongicides classiques, soit il peut être utilisé avec un extrait hydrophobe clarifiéd’huile de graines de neem. Mots clés: agents de lutte biologique, endophytes, Erisyphe pulchra, la lutte intégrée contre des maladies des plantes, mycoparasitisme few flowers for breeding purposes. In addition, break- Introduction down of resistance to powdery mildew or environmental Cornus florida (flowering dogwood) is a popular orna- effects which render the resistance ineffective at differ- mental tree throughout the southeastern United States. ent locations is a problem (Mmbaga and Sauvé 2004). Powdery mildew caused by Erysiphe (Sect. A few cultivars including ‘Cherokee Brave’, ‘Karen’s Microsphaera) pulchra (Cook & Peck, Braun & Appalachian Blush’, ‘Jean’s Appalachian Snow’, Takamatsu), is a devastating disease of C. florida ‘Kay’s Appalachian Mist’, and ‘Appalachian Joy’ have (Hagan and Mullen 1995; Mmbaga 1998, 2000;Li some degree of resistance to powdery mildew, but none et al. 2009). The white, powdery appearance caused by has a high level of resistance. An integrated disease dust-like mycelium on leaf surfaces reduces the aesthetic management system that combines different methods value of infected trees, as well as the rate of photosynth- would be ideal to manage powdery mildew infections esis, causing stunted growth and increasing the time (Witte et al. 2000; Li et al. 2009). needed for plants to reach optimal size for plant sale The use of biological and biorational products to sup- (Windham 1996; Mmbaga and Sauvé 2004; Agrios press fungal pathogens has been recommended in horticul- 2005; Shi et al. 2008). Severe infection with powdery tural crop production due to their lower level of toxicity mildew has also been shown to reduce flower set and the (Stimmel 1996). There are a few biorational products that amount of fruit formed (Heald 1999). are labeled for powdery mildew control and include plant Since the emergence of powdery mildew on flowering extracts from neem oil marketed as Triact 70, and neem dogwood in the early 1990s, management strategies have Gold as well as potassium bicarbonate salts marketed as relied heavily on conventional fungicides with applica- Armicarb and Kaligreen (Mmbaga and Sheng 2002). The tions starting soon after bud-break and continuing all incorporation of Armicarb and Triact 70 in fungicide rota- season-long (Windham 1994; Mmbaga 2000). tions has been reported to reduce conventional fungicide Fungicide applications such as wettable sulfur primarily usage by up to 56% (Mmbaga and Sheng 2002). Biological are used preventatively, but they are not effective against control strategies which use one living organism to control existing infections (Ash 1999). However, other fungi- the growth and proliferation of another are safer and inex- cides including thiophanate methyl and propiconazole pensive alternatives to conventional agrochemicals (Van are commonly used all season long as protective as Driesche and Bellow 1996; Hawley and Eitzen 2001; well as curative remedies. The season-long use of che- Gardener and Fravel 2002; Kiss 2004; Sharma et al. mical fungicides has significantly increased the cost of 2011). Biological-based disease management can be inte- production, leading to abandonment of C. florida pro- grated with other strategies such as host resistance and low duction by small-scale growers in favor of other orna- levels of fungicides. Since bio-control agents are living mentals (Li et al. 2009). In addition, there are concerns organisms, it is important to investigate the conditions over accidental exposure of humans and wildlife to these necessary for their optimal development, survival and effi- chemicals, environmental contamination, adverse effects cacy (Bélanger and Labbé 2002). Screening of diverse on microbial biodiversity, and beneficial non-target nat- epiphytes for biological control agents for dogwood pow- ural microflora that otherwise protect plants against other dery mildew identified bacteria, fungi and yeasts that sup- pathogens (de Jager et al. 1995; Sharma et al. 2011). pressed powdery mildew (Mmbaga et al.