Mycorrhiza induced resistance in potato plantlets challenged by Phytophthora infestans

Abstract

Biological control of soil-borne pathogens by arbuscular mycorrhizal (AM) fungi has been repeatedly demonstrated. However, their role in the control of above-ground hemibiotrophic pathogens is less conclusive. Here, we investigated in vitro the impact of an AM fungus on Phytophthora infestans in potato plants. The leaf infection index was decreased in mycorrhizal potato plants. Real-Time Quantitative PCR revealed the induction of two pathogenesis related genes (PR1 and PR2) in the leaves of mycorrhizal plants shortly after infection with P. infestans. These results suggested a systemic resistance in mycorrhizal plants, related to the priming of the two PR genes in potato.

Introduction

Phytophthora infestans (Mont.) de Bary is a hemibiotrophic pathogen with biotrophic behaviour during early infection and necrotrophic behaviour in the later stage of colonization. This pathogen induces late blight and is considered one of the most devastating microbes that cause disease in potato crops [1], [2]. Annual losses in Europe (i.e. costs of control and damage) are estimated above 1 billion € [3]. Late blight is mainly controlled by repeated pesticide applications, which is time-consuming, costly and harmful for the environment [4], [5]. The control of this disease is also conducted via integrated management practices including sanitation, utilization of certified seeds, breeding programs for resistant cultivars [4], [6] and fertilization management [7]. In the recent years, natural compounds such as plant extracts as well as microorganisms have been tested in the context of organic potato production [8]. Biological control agents such as Pseudomonas species, fungal antagonists and arbuscular mycorrhizal (AM) fungi [9], [10], [11] have been used against Phytophthora disease. For example under greenhouse conditions, Andreote et al. [10] obtained a diminution of 45% of the lesion after 5 days of infection by P. infestans in potato plants inoculated with Pseudomonas putida strain P9.

AM fungi are soil microorganisms developing obligate symbiotic associations with the roots of more than 80% of higher plants including the vast majority of agricultural crops [12]. The fungal symbiont receives carbohydrates from the plant, required to complete its life cycle, in exchange for nutrients (i.e. such as phosphate) and water transferred to the plant [12]. This bi-directional exchange results in increased growth of the plant and profuse development of the AM fungus. In parallel, AM fungi have been widely reported to confer bio-protection to plants, in particular against soil-borne pathogens. Numerous studies reported reduction in incidence and/or severity caused by fungi such as Rhizoctonia solani, Fusarium oxysporum or Verticillium Wilt and by fungal-like Oomycetes such as Phytophthora sp., Pythium sp. or Aphanomyces euteiches [13], [14].

In contrast, few studies were conducted on the above-ground control of pathogens by AM fungi and results were less conclusive [13]. Improved tolerance was observed in mycorrhizal plants against biotrophic pathogens such as powdery mildew (Blumeria sp.) and rust fungi (Oidium, Uromyces) resulting in increased plant mass and yield in comparison to non-mycorrhizal plants [13], [15]. The positive effect of AM fungi on plant resistance has also been reported against necrotrophic pathogens such as Alternaria solani in tomato [16], [17] and Botrytis cinerea in roses [18] and tomato [19]. With hemibiotrophic fungi and particularly Colletotrichum orbiculare, the impact of AM fungi was more controversial. Lee et al. [20] observed a decrease of the anthracnose disease on the leaves of cucumber plant colonized with Glomus intraradices, whereas, no significant effect on disease development was observed in cucumber plants inoculated with Glomus mosseae [21]. Finally, in the case of P. infestans, a delay in the progression of disease was observed on potato plants colonized with a commercial AM fungus inoculated in the field [9].

The modulation of plant defence by AM fungi has been proposed as one of the principal mechanism responsible for the control of above-ground pathogens [14]. In particular, the signalling pathways of jasmonic acid (JA), ethylene (ET) and salicylic acid (SA) which play major roles in the regulation of plant defence seems to be involved [22]. If the induction of defence mechanism has been clearly observed in AM fungal colonized areas of the roots, and also on the whole root system since long time [23], [24], [25], the induction of plant defence has only been recently reported in the leaves of mycorrhizal plants [14], [26]. In a recent study, Jung et al. [19] demonstrated that the induced systemic resistance conferred by G. mosseae against B. cinerea in tomato leaves was associated with a marked induction of a JA marker gene (Pin II) in the mycorrhizal plants. They argued that the Mycorrhiza Induced Resistance (MIR) against B. cinerea could be associated to a priming of JA-dependent defences.

In the present study, the effect of the AM fungus Glomus sp. MUCL 41833 on the increased resistance of potato plants against the hemibiotrophic pathogen P. infestans MUCL 43257 was evaluated. Analyses on leaf infection index and AM fungal root colonization were coupled to the Real-Time Quantitative PCR technology to estimate the impact of the AM fungus on the early-development of the late blight disease. In particular, the expression of defence genes dependent on the SA pathway: Gluthatione-S-transferase 1 (GST1) and acidic Pathogenesis Related 2 (PR2); dependent on the JA pathway: Lipoxygenase (Lox), dependent on the ET pathway: Ethylene Response Factor 3 (ERF 3) and dependent on the JA/ET pathways: basic Pathogenesis Related 1 (PR1) and Phenylalanine ammonia lyase (PAL) were visualised. This study was conducted under in vitro culture conditions using the Mycelium Donor Plant (MDP) in vitro culture system [27], which allowed to synchronize the development of the AM fungus in the potato roots.