Downloaded from www.microbiologyresearch.org by IP: 54.70.40.11 On: Mon, 17 Dec 2018 21:02:07 Low pH regulates the production of deoxynivalenol by Fusarium graminearum Donald M. Gardiner, Sheree Osborne, Kemal Kazan and John M. Manners Correspondence Donald M. Gardiner donald.gardiner@csiro.au CSIRO Plant Industry, Queensland Bioscience Precinct, 306 Carmody Road, Brisbane, Queensland 4067, Australia Received 24 March 2009 Revised 24 May 2009 Accepted 3 June 2009 Fusarium graminearum, which causes the globally important head blight disease of wheat, is responsible for the production of the harmful mycotoxin deoxynivalenol (DON) in infected grain. The production of DON by F. graminearum occurs at much higher levels during infection than during axenic growth, and it is therefore important to understand how DON production is regulated in the fungus. Recently, we have identified amines as potent inducers of in vitro DON production in F. graminearum. Although amines strongly induced expression of the key DON biosynthesis gene TRI5 and DON production to levels equivalent to those observed during infection, the timing of this induction suggested that other factors are also likely to be important for the regulation of DON biosynthesis. Here we demonstrate that low extracellular pH both promotes and is required for DON production in F. graminearum. A combination of low pH and amines results in significantly enhanced expression of the TRI5 gene and increased DON production during axenic growth. A better understanding of DON production in F. graminearum would have implications in developing future toxin management strategies. INTRODUCTION Fusarium graminearum (Gibberella zeae Schwabe) and closely related Fusarium spp. cause fusarium head blight (FHB) in wheat (Triticum aestivum L.) and other small- grain cereals, and ear rot in maize (Goswami & Kistler, 2004). FHB is one of the most economically important diseases of wheat worldwide and causes yield losses, grain quality reduction and the contamination of grain with trichothecene mycotoxins (Goswami & Kistler, 2004). Trichothecenes such as deoxynivalenol (DON) are potent, non-specific inhibitors of eukaryotic protein synthesis and are produced by the fungus during infection as part of its virulence arsenal. Isolates of F. graminearum unable to produce trichothecenes show reduced aggressiveness towards wheat (Bai et al., 2002; Jansen et al., 2005; Proctor et al., 1995). Trichothecenes are also harmful to humans and animals, and maximum allowable levels of trichothecenes in final food products are highly regulated (Anonymous, 2005). Therefore, understanding the regu- lation of mycotoxin production in the pathogen by endogenous and exogenous cues is important to enable the design of novel control strategies for Fusarium-incited diseases and also for better management of grain storage. The conditions required for the production of many known fungal secondary metabolites appear very specific. For example, the production of fumonisin by Fusarium verticillioides during infection of maize ears is triggered by amylopectin, a component of starch (Bluhm & Woloshuk, 2005). In Aspergillus nidulans, endogenous and exogenous oxylipids differentially regulate sterigmatocystin and peni- cillin production (Tsitsigiannis & Keller, 2006). A number of recent reports have also implicated various compounds, such as H 2 O 2 , NaCl, azole fungicides, magnesium, carbon source and a large number of phytochemicals, in both positive and negative regulation of trichothecene produc- tion in vitro (Boutigny et al., 2008; Jiao et al., 2008; Ochiai et al., 2007; Pinson-Gadais et al., 2008; Ponts et al., 2006). However, in these studies the amount of DON produced in culture does not appear to approach the levels observed in infected heads, where concentrations can exceed 500 p.p.m. on a fresh weight basis (Boddu et al., 2006; Mudge et al., 2006). In Fusarium sporotrichioides and F. graminearum, amines, such as the plant stress metabolite putrescine, have been shown to be involved in the stimulation of trichothecene production in vitro to levels similar to those observed in infected plants (Gardiner et al., 2009). In addition to specific inducing molecules, the production of secondary metabolites is usually determined by generic factors and associated cellular regulatory systems, such as the global nitrogen, carbon and pH regulatory systems, common to most fungi (Yu & Keller, 2005). For example, nitrogen and carbon source and low pH play key roles in regulation of sterigmatocystin/aflatoxin production in Aspergillus spp. (Kachholz & Demain, 1983; Keller et al., 1997a; Woloshuk et al., 1997). In A. nidulans, low pH is Abbreviations: DON, deoxynivalenol; RFU, relative fluorescence units. Supplementary material is available with the online version of this paper. Microbiology (2009), 155, 3149–3156 DOI 10.1099/mic.0.029546-0 029546 G 2009 Commonwealth Scientific and Industrial Research Organisation Printed in Great Britain 3149