Investigation of the mechanisms of sexual reproduction in Zymoseptoria tritici and their consequences on STB dynamics

ISSDC9 Paris, 7-9 April 2016 Investigation of the mechanisms of sexual reproduction in Zymoseptoria tritici and their consequences on STB dynamics F Suffert*, D Morais, G Delestre, S Gélisse, V Laval, I Sache INRA BIOGER, France Epidemiology of Wheat Diseases group * frederic.suffert@versailles.inra.fr @cropsafexpert

Introduction Asexual stage: well described, visible, quantifiable (aggressiveness traits) (Suffert et al., 2013) efficacy of fungicide programs expression of host resistance disease forecast models Sexual stage: cryptic, difficult to quantify (but crucial to investigate dynamic of epidemics)

Introduction Evidence of sexual reproduction biology (Sanderson, 1972) population genetic (e.g., Zhan et al., 1998) Ascospores produced on wheat debris = main form of primary inoculum (Suffert et al., 2011; Suffert & Sache, 2011)

Introduction Why investigating sexual reproduction in STB? Applied interest Control of primary inoculum Academic interest Ecology of a fungal pathogen with dual reproduction mode - role of ascospores during epidemics - management of wheat debris in no-till farming (biocontrol?) - population genetics - trade-off between pathogenicity and transmission

1. Contribution of the ascospores to the early epidemic stages ascospores produced from - distant, old debris - local basal leaves M. Duvivier ascospores wind-dispersed from distant debris pycnidiospores splash-dispersed from basal leaves - upwards movement (Shaw, 1987) - sideways movement from older to younger leaves (Lovell et al., 2004) G. Garin T. Vidal Wheat canopy = target or source of ascospores? Erroneous models? if - epidemiological framework not clearly posed - biological knowledge is missing

Effect of the presence of infected debris on early STB dynamic strong, although transient and dependant of their fungal load (year effect) (Suffert & Sache, 2011; Morais et al., 2015; 2016a; 2016b) infected debris (wheat monoculture) no debris 0 +++ 8-year field experiment

Amount of airborne ascospores characterized at early epidemic stages, coupling spore trapping and qpcr the quantity of primary inoculum is rarely limiting (Morais et al., 2016a) Amount of DNA daily collected by spore trap 29 30 31 32 Ct value Ct 33 34 35 36 37 38 39 40 0 20 40 60 80 100 120 140 160 180 number of days 9 oct. 2011 1er fév. 2012 nombre de jours LOQ LOD up LOD low 121 ascospores 19 ascospores 13 ascospores

Comparison of pathogenicity of ascospores and pycnidiospores the latent period was significantly shorter (60 dd) after infection with pycnidiospores than ascospores 100 472.7 dd 536.2 dd 80 % of lesions LES(t)/LESmax (%) 60 40 36.8 20 ascospore pycnidospore 0 300 400 500 600 700 800 time (ddpi) latency (Morais et al., 2015)

2. Epidemiological determinants of sexual reproduction, from leaf to canopy What is determining sexual reproduction at leaf/ plant/ canopy scales?

2. Epidemiological determinants of sexual reproduction, from leaf to canopy Who? - effect of parental genotype? - crossing VR x AVR strains? (Ware, 2006) Mat1.1 Mat1.2

2. Epidemiological determinants of sexual reproduction, from leaf to canopy Who? t - effect of parental genotype? - crossing VR x AVR strains? (Ware, 2006) Where? - effect of infection site? (upper leaves, basal leaves, stem) When? - effect of co-infection timing? - effect of age of wheat tissue?

2. Epidemiological determinants of sexual reproduction, from leaf to canopy Who? - effect of parental genotype? - crossing VR x AVR strains? (Ware, 2006) Where? - effect of infection site? (upper leaves, basal leaves, stem) When? - effect of co-infection timing? - effect of age of wheat tissue? How? - effect of lesion density (coalescence)?

Experiments in sexual reproduction crossing in semi-controlled conditions - on seedlings (Kema et al., 1996): collection of progeny, for population genetics heritability of traits -on adult plants: for quantitative epidemiology quantity of ascospores = proxy of sexual reproduction intensity (Suffert et al., accepted)

Significant effect of parental genotype some are more capable for sexual reproduction than others nb ascospore / g 200 150 100 50 I07 I08 I09 I12 I24 I25 I28 I03 I04 I05 parental genotype Detrimental effect of co-infection delay on ascosporogenesis competition effect for resources? nb ascospor re / g 35 30 25 20 15 10 5 a a b c 0 Δ0(mix) D0-mix Δ0(succ) D0-succ. Δ15days D15 Δ22days D22 (Suffert et al., accepted) delay in co-infection

Higher STB severity is associated with higher levels of ascospore production at the plant scale (corroborated at field scale ; Cowger et al., 2002) nb ascospore / g 2500 2000 1500 1000 500 0 SEV max SEV opt 0 20 40 60 80 100 disease severity (%) threshold effect of lesion density on the intensity of sexual reproduction (competition effect for resources?) (Suffert et al., in prep.)

Sites of sexual reproduction shift from basal to upper parts correlatively to co-infection timing 70 Sept. Jan. Nov. 60 50 40 30 20 10 wheat stubble (after harvest) 10 100 1000 10000 nb ascospores / cm (Suffert et al., in prep.)

3. Sexual reproduction, transition between two epidemic seasons articulation annual vs. inter-annuel dynamics Trade-off between pathogenicity (asexual multiplication) and transmission(sexual reproduction on debris)? spor rulation index 100 high pathogenicity 80 medium 60 40 20 low 10 parental isolates 28 crossings 0 350 400 450 500 550 latency (dpi) no effect of parental aggressiveness traits on the amount of ascospores (Suffert et al., accepted)

However, increase of aggressiveness over the course of epidemics (Suffert et al., 2015) A-L. Boixel Trade-off pathogenicity/transmission likely due to: - Allee effect (on the upper part of plant, the low lesion density decreases the likelihood of mating between compatible strains)

Take-home message Z. tritici is a relevant model to investigate causes and consequences of sexual reproduction on STB epidemics (1) the epidemiological determinants of sex can be studied using crossing on adult plant number of ascospores = proxy of life history trait to be studied (2) complexity of biotic/abiotic factors, spatial/temporal scales which impact sexual reproduction knowledge needed on nature, amount and efficiency of primary inoculum (3) possible trade-off between pathogenicity(asexual stage) and transmission (sexual stage) to be investigated

Thanks for your attention David Morais Valérie Laval Florence Carpentier Ivan Sache Sandrine Gélisse Ghislain Delestre Thierry Marcel Henriette Goyeau PLANTFOODSEC - UE FP7 EMPHASIS- H2020 Lydie Kerdraon Anne-Lise Boixel SEPTOVAR - LabEx BASC

References Suffert F, Sache I, Lannou C (2011) Early stages of septoria tritici blotch epidemics of winter wheat: Build-up, overseasoning, and release of primary inoculum. Plant Pathology 60: 166-177. http://dx.doi.org/10.1111/j.1365-3059.2010.02369.x Suffert F, Sache I (2011). Relative importance of different types of inoculum to the establishment of Mycosphaerella graminicola in wheat crops in north-west Europe. Plant Pathology 60: 878-889. http://dx.doi.org/10.1111/j.1365-3059.2011.02455.x Suffert F, Sache I, Lannou C (2013) Assessment of quantitative traits of aggressiveness in Mycosphaerella graminicola on adult wheat plants. Plant Pathology 62: 1330-1341. http://dx.doi.org/10.1111/ppa.12050 Suffert F, Ravigné V, Sache I (2015) Seasonal changes drive short-term selection for fitness traits in the wheat pathogen Zymoseptoria tritici. Applied and Environmental Microbiology 81: 6367-6379. http://dx.doi.org/10.1128/aem.00529-15 Morais D, Laval V, Sache I, Suffert F (2015) Comparative pathogenicity of sexual and asexual spores of Zymoseptoria tritici (Septoria tritici blotch) on wheat leaves. Plant Pathology 64, 1429 1439. http://dx.doi.org/10.1111/ppa.12372 Morais D (2016) Components of the early stages of septoria tritici blotch epidemics (Zymoseptoria tritici): quantity, efficiency and origin of primary inoculum. PhD Thesis, 204 p., available on https://hal.archives-ouvertes.fr/tel- 01142864/document Morais D, Sache I, Suffert F, Laval V (2016a) Is onset of Septoria tritici blotch epidemics related to local availability of ascospores? Plant Pathology 65, 250-260. http://dx.doi.org/ 10.1111/ppa.12408 Morais D, Laval V, Sache I, Suffert F (2016b) Inferring the origin of primary inoculum of Zymoseptoria tritici from differential adaptation of resident and immigrant populations to wheat cultivars. European Journal of Plant Pathology 145: 393-404. http://dx.doi.org/10.1007/s10658-015-0853-y Suffert F, Delestre G, Carpentier F, Walker AS, Gazeau G, Gélisse S, Duplaix C (-) Fashionably late partners have more fruitful encounters: impact of the timing of co-infection and pathogenicity on sexual reproduction in Zymoseptoria tritici. Fungal Genetics and Biology, accepted.