Baker, Daniel

Fitt, Bruce

Fortune, James

Huang, Yongju

Karandeni Dewage, Chinthani Shanika

Ritchie, Faye

Stanley, James

English

University of Hertfordshire Research Archive

Management of Diseases and Pests of Oilseed Rape Jellis & Fitt (Eds) 2021 Agrifood Charities Partnership & University of Hertfordshire 25  Chemical warfare: the fungal quest to conquer oilseed rape  By JAMES FORTUNE, DANIEL BAKER, JAMES STANLEY,                                    CHINTHANI KARANDENI DEWAGE, FAYE RITCHIE, BRUCE D L FITT                and YONG-JU HUANG  Centre for Agriculture, Food and Environmental Management, School of Life and Medical Sciences, University of Hertfordshire, Hatfield, Hertfordshire, AL10 9AB, UK Corresponding Author Email: y.huang8@herts.ac.uk  Key words: Phoma stem canker, Leptosphaeria maculans, Leptosphaeria biglobosa, sirodesmin PL, interspecific interactions, phomamide  Introduction   Phoma stem canker, caused by Leptosphaeria maculans and L. biglobosa, causes an average yield loss of > £70M annually in UK oilseed rape (www.cropmonitor.co.uk) (Zhang et al., 2014). Previous studies had shown that L. biglobosa ascospores were released later than those of  L. maculans (Huang et al., 2011). However, more recent investigations that have used qPCR analysis have reported that ascospores of both species are more frequently released at similar times (Javaid et al., 2018). L. maculans produces sirodesmin PL, a non-host selective epipolythiodioxopiperazine; L. biglobosa does not (Pedras & Yu, 2009). Sirodesmin PL has an inhibitory effect on L. biglobosa (Elliott et al., 2007). There has been limited work investigating the interaction between L. maculans and L. biglobosa at key stages of their life cycles. Therefore, this study aims to provide a better understanding of the unknown interactions between L. maculans and L. biglobosa and investigate the changes in phytotoxin production as a result of increased interspecific competition.   Materials and methods   L. maculans and L. biglobosa were cultured in liquid culture, either individually or dual cultured with a competing pathogen. After 14 days, a secondary metabolites ethyl acetate extraction was done for each treatment, to investigate the effect of secondary metabolites on the colony growth of L. maculans and L. biglobosa.  Fungal plugs (8mm diameter) of L. maculans or L. biglobosa were inoculated onto clarified V8 juice agar plates. Each fungal plug was inoculated with the corresponding secondary metabolite extract from each treatment or ethyl acetate. Each treatment was replicated five times; the ethyl acetate control was replicated three times. Colony diameters for L. maculans and L. biglobosa were recorded at 7 days post inoculation and converted to colony areas. To investigate the changes in phytotoxin production, the secondary metabolites extracted from each treatment were analysed to identify differences in composition using HPLC and LC-MS.  Results    Analysis of interspecific interactions between the pathogens in vitro confirmed that different mechanisms of interspecific competition were used to out-compete each other. The secondary metabolites produced by L. maculans inhibited L. biglobosa colony growth. This inhibition was not observed when L. biglobosa was inoculated with secondary metabolites extracted from Management of Diseases and Pests of Oilseed Rape Jellis & Fitt (Eds) 2021 Agrifood Charities Partnership & University of Hertfordshire 26  the co-culture of L. maculans and L. biglobosa. There were three unique maxima found only in the secondary metabolite extracts that inhibited L. biglobosa colony growth. Using HPLC and LC-MS, these maxima were identified as sirodesmin PL precursors deacetylsirodesmin PL and phomamide, sirodesmin PL and an unknown compound. When L. maculans and L. biglobosa were co-inoculated, sirodesmin PL and its precursors were not produced. Additional maxima on the HPLC chromatograph were not found. Results of this study suggest that L. biglobosa must inhibit the formation of sirodesmin-precursor. Due to sirodesmin having an antagonistic effect on L. biglobosa, it is thought that this interference must happen very early in L. maculans-L. biglobosa interactions, before the production of sirodesmin. Considering application of the results for control of phoma stem canker in field conditions, if L. maculans and L. biglobosa ascospores are released at the same time, phoma leaf spot lesions may appear later or be smaller, allowing fungicides to be applied later.   Acknowledgements  This research was supported by the Hertfordshire Science Partnership (supported by the Hertfordshire Local Enterprise Partnership and The European Regional Development Fund), Chadacre Agricultural Trust, Felix Thornley Cobbold Agricultural Trust, the UK Biotechnology and Biological Sciences Research Council (BBSRC, M028348/1 and P00489X/1), the Innovate UK (102100 and 102641), AHDB Cereals & Oilseeds (RD-2140021105), the Department for Environment, Food and Rural Affairs (Defra, OREGIN). The authors thank the technical support team at University of Hertfordshire for their support and assistance.   References  Elliott CE, Gardiner DM, Thomas G, Cozijnsen A., Van De Wouw A, Howlett BJ. 2007. Production of the toxin sirodesmin PL by Leptosphaeria maculans during infection of Brassica napus. Molecular Plant Pathology 8:791-802  Huang YJ, Hood JR, Eckert MR, Stonard JF, Cools HJ, King GJ et al. 2011. Effects of fungicide on growth of Leptosphaeria maculans and L. biglobosa in relation to development of phoma stem canker on oilseed rape (Brassica napus). Plant Pathology 60:607-620. Javaid A, Gajula LH, Fitt BDL, Huang YJ. 2018. Investigating regional differences in proportions of Leptosphaeria maculans and Leptosphaeria biglobosa in southern England. International Congress of Plant Pathology 2018, Boston, Massachusetts, U.S.A. August 1st 2018. (Poster presentation. Abstract, Group 2: Pathogen Dispersal and Survival. 1043-P, p96). Pedras MSC, Yu Y. 2009. Mapping the sirodesmin PL biosynthetic pathway - A remarkable intrinsic steric deuterium isotope effect on a 1H NMR chemical shift determines beta-proton exchange in tyrosine. Canadian Journal of Chemistry 87:564–570. Zhang X, White RP, Demir E, Jedryczka M, Lange RM, Islam M et al. 2014. Leptosphaeria spp., phoma stem canker and potential spread of L. maculans on oilseed rape crops in China. Plant Pathology 63:598-612.   

Chemical warfare: the fungal quest to conquer oilseed rape

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Chemical warfare: the fungal quest to conquer oilseed rape

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